Peripheral Nerve Diseases: Handbook of Clinical Neurophysiology, Volume 7

Handbook of Clinical Neurophysiology Series Editors

Jasper R. Daube Department of Neurology, Mayo Clinic, 200 First Street SW, Rochester, MN 55905, USA and

François Mauguière Functional Neurology and Epilepsy Department, Hôpital Neurologique Pierre Wertheimer, 59 Boulevard Pinel, F-69394 Lyon Cedex 03, France

Volume 7 Peripheral Nerve Diseases Volume Editor

Jun Kimura Professor Emeritus, Department of Neurology, Kyoto University School of Medicine, Japan and Professor, Division of Clinical Electrophysiology, Department of of Neurology, University of Iowa College of Medicine, 200 Hawkins Drive, Iowa City, IA 52242, USA

Edinburgh London New York Oxford Philadelphia St Louis Sydney Toronto 2006

Radarweg 29, 1043 NX, Amsterdam, The Netherlands © 2006, Elsevier B.V. All rights reserved. The right of Jun Kimura to be identified as editor of this work has been asserted by him in accordance with the Copyright, Designs and Patents Act 1988. No part of this publication may be reproduced, stored in a retrieval system, or transmitted in any form or by any means, electronic, mechanical, photocopying, recording or otherwise, without either the prior permission of the publishers or a licence permitting restricted copying in the United Kingdom issued by the Copyright Licensing Agency, 90 Tottenham Court Road, London W1T 4LP. Permissions may be sought directly from Elsevier’s Health Sciences Rights Department in Philadelphia, USA: phone: (+1) 215 238 3804, fax: (+1) 215 238 3805, e-mail: [email protected]. You may also complete your request on-line via the Elsevier homepage (http://www.elsevier.com), by selecting ‘Customer Support’ and then ‘Obtaining Permissions’. This edition published 2006 ISBN 0 444 51358 2 ISSN 0 156 74 23 1 British Library Cataloguing in Publication Data A catalogue record for this book is available from the British Library Library of Congress Cataloging in Publication Data A catalog record for this book is available from the Library of Congress Note Knowledge and best practice in this field are constantly changing. As new research and experience broaden our knowledge, changes in practice, treatment and drug therapy may become necessary or appropriate. Readers are advised to check the most current information provided (i) on procedures featured or (ii) by the manufacturer of each product to be administered, to verify the recommended dose or formula, the method and duration of administration, and contraindications. It is the responsibility of the practitioner, relying on their own experience and knowledge of the patient, to make diagnoses, to determine dosages and the best treatment for each individual patient, and to take all appropriate safety precautions. To the fullest extent of the law, neither the publisher nor the authors assume any liability for any injury and/or damage to persons or property arising out of or related to any use of the material contained in this book. The Publisher

Printed in the Netherlands

For Elsevier: Commissioning Editor: Michael Parkinson Project Development Editor: Lynn Watt Project Manager: Anne Dickie Design Director: George Ajayi

List of Contributors

James W. Albers

Department of Neurology, University of Michigan Health System, Ann Arbor, MI, USA.

Amer Al-Shekhlee

Case Western Reserve University, University Hospitals of Cleveland, OH, USA.

Michael J. Aminoff

Department of Neurology, School of Medicine, University of California, San Francisco, CA, USA.

Edward A. Aul

University of Iowa, Roy J. and Lucille A. Carver College of Medicine, Department of Neurology, Iowa City, IA, USA.

Tulio E. Bertorini

University of Tennessee Health Science Center and Wesley Neurology Clinic, P.C., Memphis, TN, USA.

Juan M. Bilbao

Department of Neuropathology, Sunnybrook Health Sciences Center, University of Toronto, Toronto, Canada.

Hugh Bostock

Sobell Department, Institute of Neurology, University College London, Queen Square, London, UK.

Kevin B. Boylan

Department of Neurology, Mayo Clinic, Jacksonville, FL, USA.

Mark B. Bromberg

Department of Neurology, University of Utah, Salt Lake City, UT, USA.

William F. Brown

McMaster University Medical Center, Hamilton, Ontario, Canada.

David Burke

Institute of Clinical Neurosciences, University of Sydney and Royal Prince Alfred Hospital, Sydney, Australia.

William W. Campbell

Neurology Department, Uniformed Services University of the Health Sciences, Bethesda, MD, USA.

Phillip F. Chance

Neurogenetics Laboratory, Chief, Division of Genetics and Developmental Medicine, Department of Pediatrics, University of Washington School of Medicine, Seattle, WA, USA.

Vinay Chaudhry

Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, MD, USA.

David R. Cornblath

Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, MD, USA.

Giorgio Cruccu

Dipartimento Scienze Neurologiche, Viale Università 30, Roma, Italy.

Elaine S. Date

Division of Physical Medicine & Rehabilitation, Department of Orthopedic Surgery, Stanford University Medical Center, Stanford, CA, USA.

J. Gert van Dijk

Department of Neurology and Clinical Neurophysiology, Leiden University Medical Centre, 2300 RC Leiden, The Netherlands.

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LIST OF CONTRIBUTORS

Günther Deuschl

Department of Neurology, Christian-Albrechts-Universität Kiel, Niemannsweg 147, Kiel, Germany.

John England

Chairman of Neurology, Deaconess Billings Clinic, Neuroscience Department, Billings, MT, USA.

Roberto Guiloff

Neuromuscular Unit, West London Neurosciences Centre, Charing Cross Hospital, London, UK.

Chelsea Grow

George Washington University, Washington, DC, USA.

Mark C. Hannibal

Neurogenetics Laboratory, Division of Genetics and Developmental Medicine, Department of Pediatrics, University of Washington School of Medicine, Seattle, WA, USA.

Richard A.C. Hughes

Department of Clinical Neurosciences, Guy’s, King’s and St Thomas’ School of Medicine, London, UK.

Ryuji Kaji

Department of Clinical Neuroscience, Graduate School of Medicine, University of Tokushima, Tokushima, Japan.

Praful Kelkar

Department of Neurology, University of Iowa Hospital and Clinics, Iowa City, IA, USA.

John J. Kelly

George Washington University, Washington, DC, USA.

Matthew C. Kiernan

Institute of Neurological Sciences, Prince of Wales Hospital, Prince of Wales Medical Research Institute and University of New South Wales, Sydney, Australia.

Byung - Jo Kim

Department of Neurology, Korea University Medical Center, Seoul, Korea.

Jun Kimura

Department of Neurology, University of Iowa Hospital and Clinics, Iowa City, IA, USA.

Christian Krarup

Department of Clinical Neurophysiology NF3063, Rigshospitalet, Copenhagen, Denmark

Mark E. Landau

Clinical Neurophysiology, Walter Reed Army Medical Center, Washington DC, USA.

Daniel Larriviere

Department of Neurology, University of Virginia, Charlottesville, VA, USA.

Richard J. Lederman

Department of Neurology, S-91, Cleveland Clinic Foundation, Cleveland OH, USA.

Kerry H. Levin

Department of Neurology, Cleveland Clinic Foundation, Cleveland, OH, USA.

Cindy S.-Y. Lin

Sobell Department, Institute of Neurology, University College London, London, UK.

William J. Litchy

Consultant in Neurology, Department of Neurology, Mayo Clinic Rochester, MN, USA.

Janice M. Massey

Department of Medicine, Division of Neurology, Duke University Medical Center, Durham, NC, USA.

Daniel L. Menkes

Department of Neurology, University of Tennessee Health Sciences Center at Memphis, Link Building, Memphis, TN, USA.

Gyl Midroni

Clinical Neurophysiology, St Michael’s Hospital, Toronto, Canada.

Suraj Muley

Department of Neurology, VAMC, University of Minnesota, Minneapolis, MN, USA.

LIST OF CONTRIBUTORS

xi

Nicholas M.F. Murray

The National Hospital for Neurology and Neurosurgery, London, UK.

Anh X. Nguyen

Neurology, CentraCare Clinic, Saint Cloud, MN, USA.

Hiroyuki Nodera

Department of Clinical Neuroscience, Graduate School of Medicine, University of Tokushima, Tokushima, Japan.

J. Payan

D’ Aumale Cottages, Twickenham, Middlesex, UK.

Lawrence H. Phillips II

Department of Neurology, University of Virginia, Charlottesville, VA, USA.

David C. Preston

Case Western Reserve University, University Hospitals of Cleveland, OH, USA.

Scott Riggins

Deaconess Billings Clinic, Neuroscience Department, Billings, MT, USA.

Lawrence R. Robinson

Department of Rehabilitation Medicine, University of Washington, Seattle, WA, USA.

Howard W. Sander

Weill Medical College, Cornell University, Peripheral Neuropathy Center, New York, NY, USA.

Barbara E. Shapiro

Neuromuscular Research, University Hospitals of Cleveland, Case Western Reserve University School of Medicine, Cleveland, OH, USA.

Wilhelm J. Schulte-Mattler Neurologische Klinik und Poliklinik, Universität Regensburg, Regensburg, Germany. Benn E. Smith

Mayo Clinic College of Medicine, Department of Neurology, Mayo Clinic, Scottsdale, AZ, USA.

S Veronica Tan

The National Hospital for Neurology and Neurosurgery, London, UK.

James W. Teener

Department of Neurology, 1C325/0032 University Hospital, University of Michigan Health System, Ann Arbor, MI, USA.

W. Trojaborg

Helleruplund Alle 3, Hellerup, DK 2900, Denmark.

A. Truini

Department of Neurological Sciences, La Sapienza University, Rome, Italy.

Josep Valls-Solé

Unitat d’EMG,Servei de Neurologia, Hospital Clínic, Facultad de Medicina, Institut d’Investigació Biomèdica August Pi i Sunyer (IDIBAPS), Barcelona 08036, Spain.

David B. Voduˇsek

Division of Neurology, University Medical Center, Zaloska Cesta 7, 1525 Ljubljana, Slovenia.

Francis O. Walker

Department of Neurology, Wake Forest University School of Medicine, Winston-Salem, NC, USA.

Bradley V. Watson

London Health Sciences Centre, London, Ontario, Canada.

Asa J. Wilbourn

EMG Laboratory, S90, Cleveland Clinic Foundation, Cleveland, OH, USA.

Thoru Yamada

Department of Neurology, University of Iowa Hospital and Clinics, Iowa City, IA USA.

Malcolm Yeh

Department of Neurology, University of Iowa Hospital and Clinics, Iowa City, IA, USA.

Foreword

Clinical neurophysiology encompasses the application of a wide variety of electrophysiologic methods in the analysis of normal function and in the diagnosis and treatment of diseases involving the central nervous system, peripheral nervous system, autonomic nervous system and muscles. The steady increase in growth of subspecialty knowledge and skill in neurology has led to a need for the compilation of the whole range of physiologic methods applied in each of the major categories of neurologic disease. While some of these methods are applied to a single category of disease, most are useful in multiple clinical settings. Each volume has been designed to serve as the ultimate reference source for academic clinical neurophysiologists and subspecialists involved in neurology. It will provide information needed to fully understand the physiology and pathophysiology of disorders in their patients. As such, these volumes will also serve as a major teaching aid for trainees in each of the subspecialties. The Handbook volumes cover all the clinical disorders served by clinical neurophysiology, including the muscle and neuromuscular junction diseases, epilepsy, surgical epilepsy, motor system disorders, peripheral nerve disease, autonomic dysfunction, sleep disorders, oculomotor/balance disorders, somatosensory system disorders, behavioral disorders, visual and auditory system disorders, and the monitoring of neural function. Each volume will focus on the advances in one of these major areas of clinical neurophysiology. Each volume will include critical discussions of fresh ideas in basic neurophysiology, approaches to characterization of disease type, localization, severity and prognosis with detailed discussion of advances in techniques to accomplish these. It is recognized that some techniques will be discussed in more than one volume, but with different focuses in each of them. Each volume will also include an overview of the field; followed by a section that includes a detailed description of each of the CNP techniques used in the disorders, and a third section discussing electrophysiologic findings in specific diseases. The latter will include how to evaluate each along with a comparison of the relative contribution of each of the methods. A final section will discuss ongoing research studies, and anticipated future advances. Our increasing understanding of peripheral nerve diseases and their presentations as large fiber, small fiber, peripheral neuropathy, polyradiculopathy and mononeuropathy makes them a particularly appropriate volume in this series. We are privileged to have Jun Kimura, one of the world’s leaders in the clinical neurophysiology of peripheral nerve diseases as the volume editor. He has done a superb job of assembling the other world leaders in the description of the methods and in their application to individual categories of motor neuron disease. The volume covers diseases from the spinal anterior horn cell to the neuromuscular junction. Special emphasis is on clinical disorders, and the advances in techniques for diagnosing and understanding their pathophysiology. Wherever possibly applicable, the information presented focuses on evidence based medicine; the specificity and sensitivity of each test is provided when known, along with comparison of their relative values. Jasper R. Daube François Mauguière Series Editors

Preface

The International Federation of Clinical Neurophysiology (IFCN) has recognized the unmet need of neurologists who must practice electrodiagnostic medicine without specialized training. This discipline covers important areas of neurological diagnosis as the only technique currently available to document abnormalities of function rather than structure. Yet, there is no textbook that addresses all the issues faced by physicians who must deliver electrodiagnostic service. We have thus undertaken the task of creating a series of volumes on neurophysiological subspecialty topics utilizing the very latest information on various aspects of technology. This series constitutes one of the most ambitious projects of the IFCN attempting to review all the neurophysiologic disciplines of clinical interest. As part of this effort, our volume provides readers with a comprehensive source of reference for electrophysiologic investigations in the evaluation of peripheral nerve disorders. This volume consists of three parts: Section I Anatomy and Histology, Section II Methodology and Technique in Assessing the Peripheral Nerve, and Section III Clinical Application. The authors assigned to the first section wrote an overview on histological and physiological properties of the peripheral nervous system. This basic information should prepare readers for the subsequent presentations on techniques and the interpretation of findings. The chapters in the second section deal with general technical principles followed by detailed methods to test individual nerves, providing step-by-step instructions. Solely based on the information given herein, readers should be able to select the most appropriate technique to study the patient under consideration and interpret the data within the clinical context. The third section primarily relates to electrophysiologic abnormalities expected in each disease process, summarizing important findings for each of the techniques described earlier. This portion includes little methodological description to avoid overlap with technical chapters, and only an abbreviated clinical note pertinent to electrophysiologic findings without a full description of disease entities per se, which is outside the scope of this book. Although I have practiced only at the University of Iowa in the US and at Kyoto University in Japan, I am well acquainted with the variety of techniques widely used in European countries from my experience gained while serving on the Executive Committee of the IFCN. I have, therefore, tried to assemble an international slate of authors from different corners of the world to contribute to this volume. All of them are neurologists, physiatrists, or neurophysiologists practicing the art of clinical electrophysiology as it pertains to the assessment of the peripheral nerve. It has been my pleasure to work with so many distinguished experts in this endeavor, which will help improve the practice of clinical electrophysiology globally. I hope the perusal of this text will make the study of the next patient easier every step of the way. In electrodiagnostic assessments, we always consider nerve conduction and various other studies within the context of clinical signs and symptoms. Thus, most authors initially found it difficult to restrict the writing either to the technical domain without mentioning clinical features or to electrophysiologic abnormalities without stating the details of the methodology. Consequently, most chapters had to undergo one or two revisions after careful review. Fortunately, the authors graciously agreed to do the necessary extra work to tailor the contents to suit our needs, and to avoid major redundancy among the chapters dealing with technical and clinical aspects. Needless-to-say, certain aspects of information are repeated to make the discussion cohesive, but otherwise each chapter, standing on its own, delivers a focused description of the intended objectives.

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PREFACE

For those authors who completed the final version more than a year ago according to the original schedule, I thank you for your timely submission. For those who just recently finished the task for a number of justifiable reasons, I apologize for my constant reminders despite your effort, and thank you for your patience and hard work. A few authors dropped out after the initial agreement to participate in the project. When we did lose contact, despite our best efforts either by mail, telephone, or through mutual friends, we recruited other colleagues to step in and finish the task in the interest of time. It was my intention not to hold up publication of the volume beyond the reasonable deadline for missing chapters. Despite short notice, substitute authors were able to meet the deadline in an excellent form to bail us out. Thus, all chapters were written in a satisfactory manner, ranging over one and half years from the original target date of September 2003 to the actual date of culmination in February 2005. I wish to take this opportunity to thank all the contributors for their willingness to write and rewrite their chapters and their patience and constructive effort to achieve our goal. Jasper Daube, the Series Editor in charge of this volume, reviewed all the chapters, providing useful suggestions. I am also indebted to Sheila Mennen who coordinated our effort from the Iowa Office with the help of Leigha Rios and Wendy Sebteka to maintain a steady progress even when I worked in Kyoto or elsewhere. Regardless of my whereabouts, we were able to maintain uninterrupted correspondence by mail, which helped move the project forward in a steady, albeit slow, pace. I have used up all the favors that I could possibly ask of my friends. In fact, I am now so deeply in debt that I will be unable to edit another multi-author book for a while. But, on this sunny winter day, when we celebrate the conclusion of the project, the future can wait. Jun Kimura Volume Editor

Peripheral Nerve Diseases Handbook of Clinical Neurophysiology, Vol. 7 J. Kimura (Ed.) © 2006 Elsevier B.V. All rights reserved

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CHAPTER 1

Anatomy and histology of peripheral nerve Benn E. Smith* Mayo Clinic College of Medicine, Department of Neurology, Mayo Clinic, 13400 East Shea Boulevard, Scottsdale, AZ 85259, USA

1.1. Introduction Over the last 100 years the emphasis of human and mammalian neuroanatomy has traditionally been the central nervous system (CNS). The topic of this chapter is an introduction to the structure of “the other nervous system,” which is composed of delicate cables forming a dense tapestry of fibers coursing through every corner of the body, some axons destined to communicate with muscle fibers, others projecting to the vast cutaneous network of skin terminals, and the small diameter myelinated and unmyelinated fibers that comprise the peripheral autonomic nervous system. Neurons in the peripheral nervous system (PNS), like some cells that reside entirely within the brain and spinal cord, often traverse great distances. In the typical adult human a single dorsal root ganglion cell extends from its pacinian corpuscle in the hallux, past the cell body in the vicinity of the fifth lumbar intervertebral foramen, and up the fasciculus gracilis to its synapse with the second order neuron in the medulla. It can span 1.5 m or more, and have a diameter of only 10 to 20 microns. To illustrate these proportions on a somewhat larger scale, if the axon was 1 m in diameter, its extent from sensory nerve ending to rostral end would be approximately 100 km or more. While the cellular inhabitants of the cerebral cortex consist of as many as 1010 neurons of several different varieties, by comparison the cast of cellular characters in the PNS is relatively small. These include: (1) neurons (the anterior horn cell, dorsal root ganglion cell, and autonomic ganglion cell) and their axons; (2) the Schwann

* Correspondence to: Benn E. Smith, Assistant Professor in Neurology, Mayo Clinic College of Medicine, Consultant, Department of Neurology, Mayo Clinic, 13400 East Shea Boulevard, Scottsdale, AZ 85259, USA. E-mail address: [email protected] Tel.: +480-301-8100; fax: +480-301-8451.

cell (which together with the axon constitutes the nerve fiber, some myelinated, others still ensheathed by Schwann cells but without myelin, being therefore unmyelinated); (3) connective tissue elements (including cells and cytoskeletal scaffolding forming the endoneurium, perineurium, and epineurium), and (4) cells that comprise and travel through blood vessels. Each of these components will be described in the following pages. The unusual architecture of nerve fibers is well suited to their primary function of relaying information from one part of the body to another. Neural signals may be propagated slowly or very rapidly, with conduction velocities ranging from less than 1 to greater than 100 m/s, the peripheral axon also serves the organism by transporting small molecules, vesicles, and larger organelles both away from and toward the cell body (Brown, 2003). Multiple systems exist within the nerve fiber to mediate axonal transport (Chao, 2003). A variety of substances and cellular commodities are translocated in the slow systems (moving at 1–2 mm daily, while others are conveyed via the fast systems with velocities exceeding 400 mm per day. The functions of these transport systems are thought to include: (1) maintenance of the metabolic integrity of long cellular processes far removed from the protein synthetic machinery of the nucleus; (2) ferrying of substances to the nerve terminal to be used in synaptic functions and functions such as neuromuscular transmission; and (3) mediating so-called “trophic influences” on postsynaptic neurons or other cells with which neural contact is maintained. The neurotrophins, reviewed by Kernie and Parada, are important in peripheral development, critical to the reparative process after nerve trauma, and play a sustentacular role on postsynaptic cells (Kernie and Parada, 2000). In contrast, in unmyelinated nerve fibers, conduction velocities of propagated nerve impulses are very slow, in the range of 1 m/s. There are examples of rapidly conducting unmyelinated fibers, but upper end velocities

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can only be achieved when the diameter of the axon is substantially increased, as in the case of the invertebrate giant squid axon. In order to achieve faster conduction velocities without resorting to massively enlarged axon diameters, the mammalian solution is myelination. While the myelinated fiber may be studied readily by light microscopy, the size of the unmyelinated fiber has made the details of its morphology inaccessible until the advent of electron microscopy (Fig. 1.1). Similarly, until recent times, clinical neurophysiologic methods have largely focused on functions mediated by myelinated fibers. Advances in sensory and autonomic physiology have witnessed the development of clinically informative techniques which interrogate functions mediated by unmyelinated fibers (Low et al., 2003). In addition, immunohistochemical analyses of skin nerves have opened a new avenue to the morphologic study of small myelinated and unmyelinated fibers in health and disease (Polydefkis et al., 2003). Although it is a simple matter to distinguish myelinated fibers from unmyelinated fibers on histologic grounds at the electron micro-

BENN E. SMITH

scopic level, it is not yet possible to categorize an individual myelinated fiber as motor or sensory by morphology alone. The same is true of unmyelinated fibers. One cannot distinguish afferents from efferents by inspecting their single profiles on electron micrographs. Different classes of myelinated and unmyelinated fibers have different conduction velocities, which allow them to be classified roughly into functional groups (see below, Section 5.). 1.2. Myelinated nerve fiber anatomy Myelinated axons in the PNS are lengthy thin cytoplasmic processes that extend from parent nerve cell bodies residing either in the central nervous system, as in the case of motor neurons in the spinal cord and brain stem, or in peripheral sensory and autonomic ganglia. Many of these cells, therefore, extend from the CNS into the periphery, or, like the dorsal root ganglion cell, send long axons both centrally (dorsal column sensory axons) and peripherally (primary sensory

Fig. 1.1 A region of myelinated fibers and unmyelinated fibers from a transverse section of the tibial nerve of a normal baboon. Osmium tetroxide fixation × 7200. LMF = large myelinated fiber; SMF = small myelinated fiber; UMF = unmyelinated fiber (Thomas et al., 1993 with permission from Elsevier).

ANATOMY AND HISTOLOGY OF PERIPHERAL NERVE

afferent nerve fibers). The only peripheral nerve cells that can truly be said to reside entirely outside the CNS are the postganglionic autonomic neurons. As large myelinated nerve fibers are most accessible to morphologic, electrophysiologic, and molecular biologic study, they will be the main emphasis for the remainder of this chapter. As mentioned previously, the myelinated peripheral nerve fiber consists of the axon and multiple Schwann cells, the latter forming a series of internodes like elongated beads on a string (the axon), interrupted by narrow gaps between beads, the nodes of Ranvier. This cylindrical myelin sheath typically invests the axon from a point near the cell body, extending peripherally to within a few microns of the nerve terminals of the axon. The cytoplasm of each myelin-forming Schwann cell–other Schwann cells invest but do not form myelin around unmyelinated peripheral axons, with cytoplasmic profiles being stained histochemically by glial fibrillary acid protein (Cheng and Zochodne, 2002) – forms a narrow band outside of the myelin sheath referred to as the abaxonal cytoplasm. The outermost turn of the abaxonal cytoplasm forms the outer mesaxon (Fig. 1.2). This abaxonal compartment is home to a number of organelles and cytoskeletal ele-

Fig. 1.2 Transmission electron micrograph of a transverse section of a small myelinated fiber from 5 day postnatal mouse sciatic nerve. aSc = adaxonal Schwann cell cytoplasm; im = inner mesaxon; ml = myelin lamellae; om = outer mesaxon; sm = Schwann cell surface membrane (Reproduced from Thomas, Ochoa,1984 with permission from Elsevier).

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ments, which are excluded from the region of compact myelin, including an elongated cigar-shaped Schwann cell nucleus, often located approximately midway between the nodes that define the borders of a given internode. Another layer of Schwann cell cytoplasm, the adaxonal compartment, can be found juxtaposed between the innermost myelin turn and the underlying axolemma. The tongue of Schwann cell cytoplasm which envelopes the axon most closely is the inner mesaxon (Fig. 1.2). The thin gap (usually 15–20 nm) between the axonal membrane and the adaxonal Schwann cell layer is the periaxonal space. Other interruptions in the compact spiral structure of the myelin sheath include pockets of cytoplasm form microvilli and terminal loops in the nodal and paranodal regions as well as usually diagonally-oriented SchmidtLanterman incisures which form irregularly occurring periodic conical gaps in the myelin along most internodes. A number of detailed and more lengthy reviews of the morphology of the myelinated axon and Schwann cell may provide additional useful information (Thomas and Ochoa, 1984; Williams et al., 1989; Peters et al., 1991; Thomas et al., 1993; Midroni and Bilbao, 1995). 1.2.1. Axonal anatomy The mammalian axon is clothed within an asymmetric trilaminar plasma membrane known as the axolemma, which ultrastructural studies have shown to be approximately 7–9 nm thick. Both the inner and outer leaflets are osmiophilic, the inner being more dense. An electron lucent clear zone separates the two. Numerous rounded 7–12 nm particles are evident, particularly on the inner surface of the axolemma by freeze-fracture studies particularly in and around the nodes of Ranvier (Thomas and Ochoa, 1984). These are thought to represent transmembrane proteins such as ion channels. The axonal cytoplasm, also called axoplasm, is rich with cytoskeletal elements, membrane bound organelles, and other structures of varying electron density. Organelles seen by ultrastructural investigations have included mitochondria, smooth endoplasmic reticulum, rare ribosomes, multivesicular bodies, dense core vesicles, glycogen particles, and polyglucosan bodies. Transmission electron microscopy has demonstrated microfilaments, microtubules, and intermediate filaments—also called neurofilaments (Thomas and Ochoa, 1984). The cross-sectional area of an axon is

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one of the main characteristics by which it is identified and classified. The caliber of axons in peripheral nerve fibers follows specific patterns and allows functional categorization of axons. In adult mammalian nerve, the normal range of axon caliber varies by a factor of 10 000, ranging from the most miniscule unmyelinated axon to the largest myelinated axon (Hoffman and Griffin, 1993). Axon diameter has physiologic and structural implications for the peripheral nervous system. The caliber of the axon influences myelin thickness and perhaps myelinogenesis in the developing nerve. The link between axon diameter and nerve conduction velocity in meters per second is equivalent to six times the diameter in μm (CV = 6 D) (Hursh, 1939). For smaller myelinated nerve fibers, the factor is less (CV = 3.5 D to 4.5 D) (Bessou and Perl, 1969; Boyd and Davey, 1968) and for unmyelinated fibers the factor is still lower (CV = 1.7 D) (Gasser, 1955). A reflection of this diameter dependency of CV is the latency of the monosynaptic reflex arc, which is increased when axon caliber is diminished (Farel, 1978). Recruitment order of myelinated nerve fibers also relates to axonal diameter (Henneman, 1965). In addition, changes in axonal cross-sectional area are frequently encountered in peripheral nervous system disorders where they may be a product of secondary demyelination. 1.2.2. Schwann cell anatomy Proceeding from the outer aspect of the Schwann cell inward, the first structure encountered is the basal lamina. The entire Schwann cell is surrounded by a basement membrane, which is continuous across the node of Ranvier, forming an elongated tube the length of the nerve fiber, thought to be important in guiding regenerating nerve fibers following injury to the axon. Like the axolemma, the plasma membrane of the Schwann cell is trilaminar, but unlike the membrane delimiting the axon, its inner and outer layers are of similar electron density. The Schwann cell which invests myelinated nerve fibers is perhaps best considered unfurled from its spiraled grasp on the axon. As Thomas and Ochoa point out, in this configuration it is a flattened, trapezoid-shaped cell that consists of three different zones (Thomas and Ochoa, 1984). The innermost or smallest dimension is the adaxonal cytoplasmic region. The outmost, somewhat wider segment is the abaxonal cytoplasm that invests the compact myelin sheath and extends the nodal microvilli from its distal and proximal margins. The intermediate portion contains compact myelin, with a lateral rim

BENN E. SMITH

of cytoplasm helically attached to the axolemma in the vicinity of the paranode. Schmidt-Lanterman incisures criss-cross the compact myelin, sometimes forming cytoplasmic conduits between the adaxonal and abaxonal cytoplasm (Fig. 1.3). The most conspicuous occupant of the outer region of the Schwann cell is it nucleus. Almost invariably oriented along the axis of its axon, it is often surrounded by a pocket of cytoplasm containing a variety of organelles such as mitochondria, endoplasmic reticulum, Golgi apparatus, centrioles, and π (pi) granules. In addition to functioning as the myelin forming cells of the PNS, Schwann cells appear to play a significant role in peripheral neural development (Mirsky et al., 2002). 1.2.2.1. Myelin sheath The characteristic ultrastructural feature of myelinated nerve fibers in transverse sections is a very regular compact spiral of alternating dark and light lamellae. On closer inspection, aldehyde fixed osmicated preparations demonstrate a radial periodicity of 12 to 17 nm, while the periodicity is somewhat greater (16 to 19 nm) by x-ray diffraction in unfixed nerve tissue (Finean, 1958). The chemical explanation for the alternating dark and light bands of peripheral myelin developed slowly over the last century. Once the main components of brain myelin were found to be protein and lipid, x-ray diffraction analysis led to the concept that myelin might consist of alternating layers of protein and lipid. The model of the plasma membrane as a lipid bilayer with hydrophobic lipid moieties directed inward and hydrophobic protein components directed outward has prepared the way for an understanding of images seen on high-resolution electron microscopy. Fixation by osmium tetroxide for electron microscopy resulted in the osmium becoming attached to the polar groups, with dark electron-dense regions interpreted as indicating polar regions in myelin where lipid and protein interact, while the light electron-lucent region was thought to represent the hydrophobic fatty acid chains of the lipid bilayer (Bischoff, 1967). Further studies have confirmed this model (Napolitano, 1969). 1.2.2.2. Schmidt-Lanterman incisures Although originally described in living nerve fibers (Schmidt, 1874; Lanterman, 1877; Nageotte, 1910), many early researchers thought these structures to be artifacts of preparation. Schmidt-Lanterman incisures are now recognized as being normal elements of myelinated nerve. Being more numerous in fibers with thick myelin, they are often not present in paranodal

ANATOMY AND HISTOLOGY OF PERIPHERAL NERVE

7

Fig. 1.3 Diagram of a Schwann cell unfurled. The white regions are those containing cytoplasm while the stippled areas semicompact myelin and the black territories compact myelin. Note the cytoplasmic channels (“incisures”) some of which are complete and others of which are blind ends (Mugnaini et al., 1977).

and perinuclear regions of compact myelin (Fig. 1.4). A number of investigators have found that incisures are more frequent in developing or remyelinating nerve (Hiscoe, 1947; Ghabriel and Allt, 1981). Although the function of Schmidt-Lanterman incisures has not been established, including the contention that they are involved in peristaltic movement of axoplasm or transport of metabolites from the Schwann cell to the axon, some experiments have demonstrated the translocation of tracers such as ruthenium red first in the cytoplasmic spiral, then in the adaxonal Schwann cell cytoplasm, and finally in the axon itself (Thomas and Ochoa, 1984). 1.2.2.3. Node of Ranvier In 1871, Ranvier described regular indentations in peripheral nerve fibers as an “annular constriction of the nerve tube, of an elegant form.” It was not until well into the following century that the nature of the nodes was

determined to be a segmental interruption in the myelin sheath (Ranvier, 1871) (Fig. 1.5). Berthold has reviewed the ultrastructural maturation of nodes of Ranvier in the developing cat (Berthold, 1996). 1.2.2.3.1. The nodal axon. On approaching the node of Ranvier the axon narrows in caliber to somewhere between 1/3 and 1/6 of the internodal axonal cross sectional area. The effect seems to be more pronounced for larger axons. A particularly unusual finding at the node is the high density of particles on the outer cytoplasmic leaflet by freeze-fracture, as great as 1200/μm2 (Schnapp and Mugnaini, 1978). It has been suggested that these particles may correspond to voltage-gated sodium channels. As to the mechanism of clustering the particles in this particular site, Rosenbluth has suggested that protein synthesis occurs in the cell body, with mature proteins being transported in the

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BENN E. SMITH

Fig. 1.4 Transmission electron micrograph of a Schmidt-Lanterman incisure. ax = axon; ml = myelin lamellae; mt = microtubule; pv = pinocytotic vesicle; Sc = Schwann cell cytoplasm (Reproduced from Thomas, Ochoa, 1984 with permission from Elsevier).

anterograde direction floating in the axolemma, and being sequestered in the nodal region, trapped by the myelin attachment on either side of the node (Rosenbluth, 1976). The inner cytoplasmic leaflet of the nodal axolemma (P face) also demonstrates a high density of particles. Whether these may be associated with ATPase activity or other ion channels has not yet been established. It is

known that the nodal axon has few mitochondria, leading to the concept that some of the energy for nerve conduction may arise from the adjacent Schwann cell (Williams and Landon, 1964). 1.2.2.3.2. The paranodal region. The morphology of the myelin sheath and its associated myelin is nothing short of spectacular in the paranodal region. The first

Fig. 1.5 Transmission electron micrograph of an osmicated mouse nerve, longitudinal section, illustrating a node of Ranvier. ax = axon; m = mitochondrion; mvb = multivesicular body; tl = terminal loops of Schwann cell cytoplasm (Reproduced from Thomas, Ochoa, 1984 with permission from Elsevier).

ANATOMY AND HISTOLOGY OF PERIPHERAL NERVE

departure from the compacted myelin in the internode is a series of broad large folds, typically three to six in number, which extend for a distance of approximately 40 μm from the node on large myelinated fibers. The axon within the region of myelin expansion conforms to the shape of the myelin folding. The valleys between the myelin folds on the outer aspect of the myelin layer contain Schwann cell cytoplasm packed with large numbers of mitochondria (Berthold, 1967). Of note, the large myelin expansions on either side of the node are not symmetrical. The expansion on the proximal side, closer to the cell body, is larger. It is apparent from the nerve compression experiments of Ochoa and colleagues and the morphologic research on inherited tendency to pressure palsies by Dyck and co-workers that the terminal rim of cytoplasm of the myelin lamellae is firmly attached to the axolemma on both sides of the node (Ochoa et al., 1972; Yoshikawa and Dyck, 1991). 1.2.2.3.3. The nodal gap. As the myelin tapers toward the node, the lamellae separate, forming multiple small rounded bulbs of Schwann cell cytoplasm, like a collar of clusters of grapes applied to the axon. These nodal processes are 70–100 nm in diameter. Histochemical studies have shown that the node is bathed in polyanionic materials which might form a cation reservoir at the node. This could be important in buffering the ionic currents that accompany saltatory conduction. 1.2.3. Molecular biology 1.2.3.1. Structural proteins of peripheral nerve The major peripheral nerve structural proteins occur in high concentrations. These molecules have no known enzymatic properties, but do possess the capacity to polymerize into filamentous structures (Pleasure, 1984). Interstitial collagen fibrils, that abound in nerves, are composed entirely of tropocollagen. The main function of collagen that constitutes 25 to 50% of total peripheral nerve protein, is to enable nerve trunks to resist stretch and mechanical forces. Tissue culture studies have shown that interstitial collagen is also important for normal myelin development. Type IV collagen is a major constituent of the basal lamina, which invests perineurial cells, Schwann cells, and axons. At least four different varieties of intermediate filaments (IF) have been found in peripheral nerve. Ten nanometer (nm) neurofilaments, identified only in

9

nerve cell bodies and axons, are polymers of peptides with a range of molecular weights between 68 and 210 kD. There are two types of IF in Schwann cells. One consists of polymers of vimentin, while another is likely related to desmin, a component of IF more often associated with muscle. Myelinating Schwann cells control the number and phosphorylation of axonal neurofilaments, which are direct determinants of axonal caliber (Martini, 2001). Some satellite cells in peripheral autonomic ganglia may contain IF composed of peptides antigenically similar to glial fibrillary acid protein (GFAP), and therefore bear a similarity to IF of CNS astrocytes. The role each of these intermediate filament subtypes plays is unknown. As a cytoskeletal-associated protein, periaxin may function by regulating the shape of the Schwann cell and by mitigating gene expression during formation of the myelin sheath by facilitating integration of extracellular signalling through the cytoskeleton (Takashima et al., 2002). Microfilaments of 6 nm diameter are characteristically detected in large numbers in the growth cones of advancing axons. Sensitive immunohistochemical techniques also demonstrate these actin microfilaments to be widely distributed in most cell types, including mature neurons and Schwann cells. Actin microfilaments play an essential role in cell motility and have been implicated in axonal transport. Microtubules consist of 25 nm cylinders formed by polymerization of tubulin dimers. These structures are abundant in axons and also present in Schwann cells, fibroblasts, and other cells in peripheral nerve. Microtubules play a major cytoskeletal function as well as participating heavily in axoplasmic transport. In recent years, a number of neuropeptides have been demonstrated in association with cutaneous peripheral nerve terminals in the epidermis, dermis, and specialized sensory end organs (Scholzen et al., 1998). These neuropeptides include calcitonin gene-related polypeptide (CGRP), neurokinin A (NKA), neuropeptide Y (NPY), substance P (SP), and vasoactive intestinal polypeptide (VIP) among others. Connexin 32 (CX32) is a gap junction protein present in uncompacted myelin which in Schwann cells is used to join adjacent membranes in paranodal loops and Schmidt-Lanterman incisures. CX32 forms tiny conduits that facilitate transfer of ions and other small molecules between Schwann cells and axons. When the

10

gene for CX32 is defective, X-linked Charcot– Marie–Tooth (CMTX) results (Bergoffen et al., 1993). 1.2.3.2. Glycoproteins of peripheral nerve myelin Most of the studies of glycoproteins in peripheral nerve have focused on peripheral nerve myelin, as these have significance to a number of peripheral neuropathies. Preparations of isolated myelin from peripheral nerve have been found to contain a prominent glycoprotein, P0 (“P zero”), a homophilic adhesion molecule with a molecular weight of 28 to 30 kD. This protein represents 50 to 60% of the total myelin protein in the peripheral nervous system and is thought to be the major protein mediating myelin compaction. Myelin P0 undergoes post-translational fucosylation, phosphorylation, and sulfation. Its carbohydrate content constitutes 6.3% by weight and exists as a monosaccharide having one sialic acid, one galactose, one fucose, three mannose, and three N-acetylglucosamine residues per mole of protein (Poduslo, 1984). The P0 gene is encoded by chromosome 1q21-23. Different mutations in the P0 gene may result in divergent morphologic and physiologic effects on the myelin sheath. Depending on where the point mutation occurs, the result can be Charcot–Marie–Tooth 1B, Dejerine-Sottas syndrome, congenital hypomyelination, or CMT2 (Zhou and Griffin, 2003). Peripheral myelin protein 22 (PMP22) is a transmembrane glycoprotein found in compact myelin, which appears to function in the formation and maintenance of myelin, but which may also be involved with Schwann cell proliferation, differentiation, and apoptosis (Fabbretti et al., 1995; Zoidl et al., 1995; Jetten and Suter, 2000). While abnormal duplication of a 1.5 megabase domain of chromosome 17p11.2 that contains the PMP22 gene leads to Charcot–Marie–Tooth 1A (CMT1A), point mutations in the PMP22 have been associated with a variety of other inherited neuropathies including CMT1 phenotype, hereditary neuropathy with liability to pressure palsies (HNPP), and rarely Charcot–Marie–Tooth 2 (Zhou and Griffin, 2003). A deletion of the 1.5 megabase stretch of 17p11.2 is the most common cause of HNPP. Two basic proteins have been found in peripheral nerve myelin. The first, called P1, has a molecular weight of 18.5 kD and is similar if not the same as myelin basic protein in the CNS. It induces experimental allergic encephalomyelitis (EAE) when injected into experimental animals. The second is P2, which is found in very low concentrations in adult Schwann cells

BENN E. SMITH

(Webster, 1993). P2 injected experimentally results in experimental allergic neuritis (EAN). Myelin-associated glycoprotein (MAG) is a high molecular weight glycoprotein and a member of the immunoglobulin superfamily. One function of MAG is to serve as a signal to modulate the caliber of myelinated axons (Yin et al., 1998). This glycoprotein localizes to Schmidt-Lanterman incisures, paranodal regions, and periaxonal Schwann cell regions, and has been implicated in some autoimmune neuropathies. 1.2.3.3. Lipids of peripheral nerve myelin Although there are some differences that appear to correspond to age, studies of peripheral nerve myelin lipids, endoneurium, or whole nerve from several mammalian species reveal many similarities. The major lipids of the membranes are cholesterol, cerebroside, ethanolamine plasmalogen, sphingomyelin, phosphatidylcholine, and phosphatidylserine, the most abundant being cholesterol (Yao, 1984). The class of lipids of greatest clinical interest to physicians involved with neuromuscular disease, the glycolipids (or gangliosides), has been the subject of a recent review (Willison and Yuki, 2002). Advancing knowledge of these molecules is improving the localization of specific glycolipids to peripheral nerve with increasing precision (Gong et al., 2002). Patients with Guillain–Barré syndrome associated with preceding Campylobacter jejuni infection have been found to have high titers of IgG antibodies to GM1, GD1b, GD1a, and GalNAc-GD1a. Elevated anti-GQ1b antibodies are consistently associated with the Miller Fisher syndrome. 1.3. Development and aging: fiber diameter composition of peripheral nerve The fetal human sural nerve has been studied extensively and is useful for comparison with adult nerve fibers because this nerve is sometimes biopsied in children and adults for diagnostic purposes (Ochoa, 1971; Shield et al., 1986). In the 9-week old human foetus the sural nerve has small fascicles containing large bundles of axons surrounded by Schwann cell processes. A basal lamina surrounds these processes with narrow clefts containing collagen fibrils. By week 16, the axon bundle size diminishes and the number of bundles increases as does the Schwann cell number. The unimodal fiber diameter histogram peak has increased to 2.4 μm. Studies of the developing trochlear nerve showed that the axonal outgrowth occurs at different times, with some fibers undergoing

ANATOMY AND HISTOLOGY OF PERIPHERAL NERVE

degeneration before and others after myelination (Mustafa and Gamble, 1978). Myelination appears to have begun in human sural nerve by week 18 (Ochoa, 1971). At this stage, a number of axons between 1.0 and 3.2 μm in diameter had paired 1:1 with Schwann cells and had formed small spirals of cytoplasm. During the postnatal period the number of myelinated fibers rises from nearly 4000 to about 12000 by age 5 years (Gutrecht and Dyck, 1970). Sural nerve conduction velocities increase from 35 m/s during months 1–6 to 40 m/s between months 7–12 and further to between 48 and 62 m/s from age 25–60 months, the normal range being 43 to 65 m/s between ages 121 and 180 months (Kuntz NL [1983] Childhood sensory nerve conductions studies [unpublished data]). The adult normal value range is 43 to 59 m/s (age 20–40). Conduction velocity slows progressively beginning at 20–30 years of age, by age 80 having slowed by approximately 10 m/s (Daube, 1996). In normal peripheral nerve, the contour of the compound action potential (the waveform recorded from a nerve following application of an external electrical impulse of sufficient energy to depolarize the vast majority of myelinated fibers in that nerve) is determined by the relative numbers of excitable and conducting nerve fibers demonstrating various conduction velocities. Because conduction velocity and response amplitude are proportional to diameter of the fiber, by using appropriate proportionality factors, one can make a reasonable prediction of the fiber diameter spectrum in normal nerve from the contour

11

of the compound action potential. This principle was first applied by Gasser and Erlanger (1927), and more recently further explored by Lambert and Dyck (1984). The diameter histogram plotted per mm2 of fascicular area of normal sural nerve tissue of infants or small children show a unimodal histogram with a peak in the 3 to 5 μm range. Similar plots made from normal adult human sural nerve demonstrates a bimodal histogram with peaks at 2.5 and 8.0 μm. (Fig. 1.6), the normal distribution of myelinated fibers in the adult being from 2 to 22 μm (O’Sullivan, 1968). 1.4. Fascicular anatomy of peripheral nerve trunks Peripheral nerve trunks contain multiple fascicles (Fig. 1.7). While there appears to be a high degree of somatotopy in peripheral nerves, because of the exchange between fibers between fascicles as they course from proximal to distal, partial nerve lesions may produce restricted deficits that might confuse the clinician attempting to localize the lesion (Stewart, 2003). The same is true of the sympathetic trunk (Aguayo et al., 1976). 1.5. Nerve terminal anatomy 1.5.1. At the neuromuscular junction Each motor unit is composed of one anterior horn cell, its single axon, the terminal branches which fan

Number of axons

Child

Adult

100

100

90

90

80

80

70

70

60

60

50

50

40

40

30

30

20

20

10

10 0.4 0.8 1.2 1.6 2.0 2.4 2.8 3.2 μm diameter

0.4 0.8 1.2 1.6 2.0 2.4 2.8 3.2 μm diameter

Fig. 1.6 Frequency distribution of unmyelinated fibers from a boy aged 15 years (left) and an adult patient with isoniazid neuropathy (Ochoa, 1970 with permission from Elsevier).

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BENN E. SMITH

Fig. 1.7 Transverse section of mammalian peripheral nerve consisting of ten fascicles fixed with osmium tetroxide. The myelinated fibers stand out as dark rings (Stewart, 1987).

Synapsin-I,II Rab 3a Rabphilin 3a Cysteine string protein SV2 Synaptobrevin Synaptotagmin Synaptophysin Vesicular ACh transporter

S-100 N-CAM

Choline acetyltransferase

nn Schwa Cell

Nerve Terminal

a CGRP

from the axon near or in the muscle it innervates, the neuromuscular junctions formed by those terminal branches, and the postsynaptic skeletal muscle fibers supplied by the anterior horn cell. Although the vast majority of muscle fibers in normal individuals are innervated by a single anterior horn cell, in extraocular muscles some muscle fibers may receive synaptic input from multiple anterior horn cells (Meiggioli et al., 2004). As a myelinated fiber approaches its muscle, the myelin sheath is lost and the terminal branches, invested by Schwann cell cytoplasm, near the muscle membrane. Each fine terminal branch expands into a terminal bouton or swelling, which comes to rest in a small depression in the muscle membrane. The basal lamina of the Schwann cell is continuous with that of the sarcolemma. The highly intricate subcellular design of the presynaptic nerve terminal, the synapse, the postsynaptic nerve terminal, the muscle cell, and the organelles that populate these structures are well beyond the focus of this chapter. The reader is encouraged to peruse Engel and Frazini-Armstrong’s authoritative text which describes muscle development, muscle contraction, neuromuscular transmission, and structural details of the neuromuscular junction in a well-illustrated and comprehensive review (Engel and FranziniArmstrong, 2004). A number of proteins at the neuromuscular junction have both physiologic and pathophysiologic significance for clinical neuromuscular disorders (Fig. 1.8).

Acetylcholine

1.5.2. In the skin Liminin-a2,a4,a5,b2,g1 Collagen IV-a3,a4,a5 AChE-T, Q Agrin Neuregulin GalNAcβ-X N-CAM Na channels Ankyrin Dystrophin aDystrobrevin-2 Syntrophin-α1,β1

Rim K+ channels Ca++ channels Syntaxin SNAP 25

Muscle Fiber

Rapsyn Utrophin aDystrobrevin-1 β2-Syntrophin

AChR-α,β,δ,e Intergrin a7A,B,b1 Dystroglycan a,b Sarcoglycan-a,β,g,δ MuSK ErbB2,3,4 GalNAc transferase

Fig. 1.8 Molecular components of the neuromuscular junction of functional, structural, and pathophysiologic significance. The elements for which knockout mice have been generated are indicated in boldface type (Sanes et al., 1998).

Primary afferent nerve fibers in mammals range in axon diameter from 0.2 to 20 μm. Some of the smallest fibers are unmyelinated. There is an uneven distribution of peripheral nerve fiber diameters (Fig. 1.9) and their corresponding conduction velocities (Table 1.1). When a single afferent nerve is supramaximally stimulated, the compound nerve action potential recorded at a distance of several centimeters shows a response consisting of multiple peaks (Fig. 1.10). These peaks are designated with uncial Roman letters for their major components and minuscule Greek letters for their subgroups (Gasser and Erlanger, 1927; Gasser, 1941). In muscle nerves, the peaks are often labelled by Roman numerals (Table 1.1). Although the peaks were originally associated with specific sensory modalities (Aαβ with tactile sensation and Aδ with sharp or stinging pain), more recently it has been established that primary sensory

ANATOMY AND HISTOLOGY OF PERIPHERAL NERVE

Aδ

13

Aαβ

60

Number of fibers

50 40 30 20 10

0 0.8

2.4 4.0

5.5 7.1 8.7 10.3 11.8 13.4 15.0 16.5 diameter (in μm)

afferents may better be categorized by the type of environmental stimulus that excite the cell preferentially, that is, at the lowest threshold, the so-called adequate stimulus (Light and Perl, 1993). In this paradigm, three categories of cutaneous sensory units emerge, low threshold mechanoreceptors, thermoreceptors, and nociceptors, which are described in greater detail elsewhere (Light and Perl, 1993). Quantitative immunohistochemical pathology has recently been used to study human myelinated and unmyelinated nerve terminals as well as sensory receptors in health and disease (Fig. 1.11). 1.6. Connective tissue elements While the electrically excitable tissues within peripheral nerve, that is the axon and myelin sheath, have been the focus of academic study and clinical interest for many years, perhaps chiefly among those with an inclination toward physiology, the interstitial eleTable 1.1

Fig. 1.9 Primary afferent nerve fiber diameter histogram of 294 randomly selected fibers from two feline sural nerves as assessed by electron microscopy (Burgess et al., 1968).

ments, that is connective tissues, have been relatively neglected. Peripheral neuropathologists have emphasized that information gleaned from surveying interstitial abnormalities in nerve is often hidden from the techniques employed by the neurophysiologist. In addition, investigations in recent years have emphasized the considerable importance of connective tissue structures in the normal function of peripheral nerves. These have been reviewed in detail by Thomas and Olsson (Thomas and Olsson, 1984). The original observations of Key and Retzius and also by Ranvier (Ranvier, 1878) formed the basis for all that has followed. Key and Retzius were the first to suggest that the peripheral connective tissues be subdivided into epineurium, perineurium, and endoneurium. This terminology still remains the most satisfactory (Key and Retzius, 1876). Some hold that, for morphologic reasons, the perineurium should not be considered as a connective tissue, but rather as part of the membranous covering of the whole of the entire nervous system, both centrally and peripherally. 1.6.1. Epineurium

Primary afferent nerve fibers classified by conduction velocity and fiber diameter. (Reproduced from Light, Perl, 1993 with permission from Elsevier).

Cutaneous nerve

Muscle nerve

Aαβ

Group I

Aδ C

Group II Group III Group IV

Conduction velocity in cat (m/s)

Diameter (mm)

72–130 35–108 36–72 3–30 0.2–2

12–22 6–18 6–12 3–7 0.25–1.35

The epineurium is thought of as a condensation of loose connective tissues that surround the perineurial ensheathment of the fascicles of uni- and multifascicular nerves. Despite being continuous with the surrounding connective tissues, the physical attachment is fairly loose, so that the nerve trunks maintain mobility except where tethered by large blood vessels or exiting branches. Where nerves cross joints greater proportions of epineurial connective tissue are present. In human nerve trunks, Sunderland and Bradley found that the epineurium usually constitutes anything from 30 to 75% of the cross-sectional

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BENN E. SMITH

Fig. 1.10 Drawing of the compound nerve action potential recorded from a mammalian saphenous sensory nerve with the Aαβ, Aδ, and C fiber components labeled (Gasser, 1941).

area, although extremes of 22 and 88% were documented (Sunderland, Bradley, 1949). As a general trend, the more fascicles a nerve trunk carries, the greater the contribution of epineurium to its crosssectional area. The collagen bundles are scattered throughout the epineurium. These fibers are primarily oriented along the axis of the nerve trunk. Elastic fibers, also mainly oriented longitudinally, have been identified

by light and electron microscopy, particularly in proximity to the perineurium. Apart from fibroblasts, which comprise the main cells constituting epineurial structures, the epineurium may contain mast cells, and even the occasional single immune system representative such as a lymphocyte or macrophage. Variable quantities of epineurial fat may be present, particularly in the larger proximal nerve trunks (Fig. 1.12).

Fig. 1.11 Confocal microscope images of cutaneous nerves stained with protein gene product (PGP) (green) and myelin basic protein (MBP) (red). The box in panel B encloses a single internode bordered on each side by a node of Ranvier (Nolano et al., 2003).

ANATOMY AND HISTOLOGY OF PERIPHERAL NERVE

The vasa nervorum enter the epineurium at various levels. On entering the nerve, these vessels communicate with a complex longitudinal anastomotic network of arterioles and venules. The epineurium also contains lymphatic vessels, which are not thought to be present within the fascicles. Sunderland surveyed the literature on the lymph drainage of nerves and concluded that an epineurial lymphatic capillary network is drained by lymphatic channels that accompany the arteries of the nerve trunk, which coalesce to form larger channels, and ultimately drain into regional lymph nodes (Sunderland, 1965). 1.6.2. Perineurium Although Key and Retzius recognized that the essential component of perineurium is a lamellated arrangement of flattened cells (Fig. 1.13), a disagreement arose concerning the embryological origin of these cells and whether there may be a separate connective tissue layer external to the lamellated cellular layer that can be distinguished from the epineurium. There is a general agreement that the perineurium provides a covering for both the somatic and peripheral autonomic nerves and their ganglia (Shantha, 1968). The perineurial lamellae are composed of concentric layers of flattened polygonal cells, the morphology of which was skillfully illustrated by Key and Retzius in their original descriptions, and subsequently confirmed

Fig. 1.12 Drawing of a transverse section through the edge of the rabbit brachial plexus showing the arrangement of epineurium, perineurium, and endoneurium (Key and Retzius, 1876).

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by electron microscopy. While the number of lamellae is generally proportional to the diameter of the fascicle, up to 15 layers may be present around the fascicles of large mammalian nerve trunks. Perineurial cells are bounded on both sides by basal lamina, although gaps in this membrane over short lengths are sometimes seen. The basal lamina is sometimes quite thick, sometimes as much 0.5 μm in human nerves. The cytoplasm is scant except in the perinuclear region. Organelles such as endoplasmic reticulum and mitochondria are predominantly found huddled near the nucleus, and glycogen particles often abound (Fig. 1.13). Thomas and Cravioto were among the first to observe that contiguous perineurial cells either overlapped or interdigitated with one another, but without interposition of basal lamina between adjacent membranes. At some sites, contiguous perineurial cells are linked by “tight junctions,” where the extracellular space appears to be obliterated (Thomas, 1963; Cravioto, 1966). Tight junctions are important in perineurial cells as evidenced by the abundant expression of claudin-1, a major structural protein of the tight junction, which interacts with occludin and ZO-1 (Folpe et al., 2002). Other markers expressed by perineurial cells include epithelial membrane antigen and glucose transporter 1 (Hirose et al., 2003). The existence of such cell relationships may be related to the diffusion barrier property of the perineurium. The finding of endocytotic vesicles,

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BENN E. SMITH

and neuromuscular junctions, however, the perineurial sleeve terminates with an open end. From detailed studies of the mouse neuromuscular junction, Saito and Zacks demonstrated that terminal perineurial cell processes form a bell-shaped cuff that does not directly contact the innervated muscle fiber (Saito and Zacks, 1969). Instead, between the termination of the perineurial sheath and the basal lamina covering the muscle fiber there is an open gap of 1 to 1.5 μm. The conclusion is that this arrangement provides a point of communication between the endoneurial space and the exterior. 1.6.3. Endoneurium

Fig. 1.13 Drawing of the surface perspective of the canine forelimb nerve demonstrating the mosaic pattern of polygonal perineurial cells (Key and Retzius, 1876).

particularly in the outer lamellae, may indicate a transport system across perineurial cells. Histochemical analysis suggests that perineurial cells contain a wide range of phosphorylating enzymes and high titers of both ATPase and creatine phosphatase activity (Shanthaveerappa and Bourne, 1962). On this basis, these cells could be considered well equipped to act as a metabolically active diffusion barrier. The nature of the junctional contacts between the perineurial cells have been further defined by the methodology of freeze-fracture. At the points of contact between adjacent perineurial cells extensive zonulae occludentes (tight junctions) are confirmed. The perineurium is penetrated by blood vessels, which link the longitudinal anastomotic network of arterioles and venules in the epineurium with the longitudinal intrafascicular capillary (microvessel) network. The consistently observed arrangement is that vessels carry in with them a perineurial “sleeve” for some distance, which does not come in close contact with the vascular wall, but rather terminates and provides a communication between the endoneurial and epineurial compartments. A perineurial investment follows the exiting terminal nerve branches into the periphery. Here the smallest nerve twigs may be ensheathed by a single layer of perineurial cells (the sheath of Henle). This layer then merges with the capsules of the muscle spindles and the encapsulated end organs. At unencapsulated endings

The endoneurium is the collective compartment within each fascicle of peripheral nerve, occupied by sparse connective tissue elements, nerve fibers, microvessels, and occasional single cells. Collagen fibrils in the endoneurium are similar in appearance and dimension to those of the perineurium, being oriented longitudinally, and demonstrating condensations in the proximity of nerve fibers and endoneurial microvessels. There are sizable spaces within nerve fascicles, which ultrastructural studies demonstrate to consist of a finely granular heterogeneous material without a specific identifiable profile, even by electron microscopy. A few morphologic patterns can, however, be discerned, including a space of variable thickness immediately subjacent to the perineurium. Nerve fibers tend to be grouped into small loose bundles. The cross-sectional area bordered by perineurium is termed the transverse endoneurial area. This has been estimated to range from 0.6 to 1.2 mm2 in distal human sural nerve. Of the total, the space taken up by dense collagen fibrils is 0.3 mm2. The remaining endoneurium populated by myelinated and unmyelinated nerve fibers is 0.2 to 0.4 mm2 (Thomas and Olsson, 1984). Each nerve fiber is clothed with a basal lamina that follows the contour of the nodes of Ranvier and is continuous across them, covering both the Schwann cell nodal and paranodal apparatus as well as the intervening extracellular space. Slightly external to this, a thin band of collagen fibrils surround the larger myelinated nerve fibers; these typically show a circular and oblique orientation. Still further from the nerve fiber are longitudinally oriented collagen fibrils comprising the outer endoneurial sheath. No clear demarcation between the inner and outer endoneurial sheaths is apparent around the smaller myelinated and unmyelinated nerve fibers, where collagen fibrils are less densely packed and oriented parallel to the nerve fiber axis.

ANATOMY AND HISTOLOGY OF PERIPHERAL NERVE

The most numerous cells inhabiting the endoneurium are endoneurial fibroblasts and Schwann cells. Occasional cells with the cytologic and histochemical features of macrophages and lymphocytes are also seen in normal mammalian and human nerve, often in a perivascular or subperineurial distribution. Mast cells may also occur in the endoneurium, being much more commonly encountered in rat nerve than in human, rabbit, or guinea pig nerve. 1.6.4. Blood vessels Not only does peripheral nerve cell architecture possess unique features distinct from that of the CNS, the PNS vascular supply demonstrates structural peculiarities providing advantages in supplying oxygen, nutrients, and waste removal for the structures they perfuse. The diffusion and transport properties of vasa nervorum endothelium appears to be of particular functional significance. There are two main structural characteristics that distinguish the peripheral nerve blood supply. These are: (1) the dense branching anastomotic network of vessels that weaves in and out of the epineurial and perineurial compartments; and (2) the system of endoneurial microvessels, which perfuses the endoneurial space. Large segmental vessels give off nutrient arteries, which, in turn, branch out to form multiple arteries that enter the epineurium. These arteries then contribute to the anastomotic epineurial/perineurial network, which ultimately is the source of the tiny terminal microvessel branches in the endoneurium. Although these vessels have the diameter of small arterioles and capillaries, they are referred to simply as “microvessels” (Olsson, 1984). Because small terminal nerve branches are devoid of endoneurial blood vessels but still often possess a perineurial sheath, they are thought to receive nutrients and other vital substances through diffusion. It may also be that very small caliber nerve trunks are partly supplied by this mechanism and therefore not completely dependent on endoneurial vessels. The arrangement of venous drainage of the nerve appears to mirror that of the arterial supply. The small and large vessels in the epineurium are invested with rather loose connective tissue. The vessel wall proper is indistinguishable from vessels in other organs, demonstrating endothelial cells, pericytes, basement membrane, and in the larger channels, circularlyoriented smooth muscle cells. Still larger vessels in the epineurium show longitudinally arrayed smooth muscle

17

elements. Transmission electron microscopy of endoneurial endothelial cells confirms the presence of numerous tight junctions between neighbouring cells. This arrangement is presumed to form an osmotic barrier which is relatively impermeable to protein and smaller molecules, which is referred to as the blood-nerve barrier (Allt and Lawrenson, 2000). When considered with the perineurium and the nerve-cerebrospinal fluid barrier, the blood-nerve barrier provides a relatively protected environment for the peripheral nerve (Olsson et al., 1971). In peripheral nerve, particularly in the course of certain inflammatory demyelinating neuropathies, a staggering array of proinflammatory cytokines are elaborated or stimulated by resident and recruited immunoreactive cells such as macrophages, lymphocytes, mast cells, and perhaps Schwann cells and neurons. These substances include tumour necrosis factor alpha and beta, interleukin 1, 2, and 6, leukemia inhibitory factor, vascular endothelial growth factor, interferon gamma, and transforming growth factor beta (Creange et al., 1997) 1.6.5. Central-peripheral transition The cells and sparse connective tissue of spinal root endoneurium are very similar to those in distal peripheral nerve, although the number and density of collagen fibrils is less and they are not nearly as organized at the root level. Where spinal roots enter and exit the spinal cord, an irregular but well-defined transition from peripheral nerve to central nervous system tissue can be identified. This has been called the Obersteiner-Redlich zone. As the posterior root axons enter this zone near the dorsal aspect of the cord, Schwann cells give rise to oligodendrocytes, and the peripheral myelin sheath is replaced by central nervous system myelin. The central portion of the root is limited at its periphery by a layer of astrocytes enclosed by basement membrane. This basal lamina extends beyond the spinal cord to ensheath nerve fibers in the peripheral segment of the root and to form the perimeter of the most proximal stretch of peripheral endoneurium. Collagen fibers are found only on the peripheral side of the junctional zone (Berthold et al., 1993). As they leave the confines of the central nervous system, spinal roots enter and course through the subarachnoid space covered by a delicate multilamellar nerve root sheath (Haller, 1971). These fibers tent and then penetrate dura mater at the subarachnoid angle. Outside of the dural sac nerve roots gain their full investment of

18

BENN E. SMITH

DM

A

SA PM EP P RS Subarachnoid space

PM

EP

A Connective tissue space DM

epineurium, perineurium, and endoneurium, which continues distally as nerve trunks take their various and sometimes convoluted courses into the periphery. As dorsal root fibers approach the spinal cord, the epineurium fuses with dura and the endoneurium continues as far as the transition between roots and the dorsal root entry zone of the spinal cord (Fig. 1.14). The transition between central and peripheral myelin, the former produced by oligodendrocytes in the CNS and the latter by Schwann cells in the periphery, is not clearly demarcated at the point of emergence of the cranial nerves and spinal roots from the neuraxis. At some sites, and especially in the cranial nerves, tongues of glial tissue (glial bundles) extend a considerable distance outside the CNS. Seen in the oculomotor and other cranial nerves, this phenomenon is particularly pronounced in the seventh cranial nerve (Thomsen, 1887). As myelinated fibers exit the central nervous system they gradually become smaller in diameter. Like microscopic palm trees rooted in the brainstem or spinal cord, myelinated fibers progressively taper the further they proceed into the periphery. This phenomenon has been documented for a number of different nerves in various species including man, and is not explainable on the basis of branching, in that nerves

Fig. 1.14 Diagram of the relationships between the meninges of the spinal cord and the peripheral nerve connective tissue elements. A = arachnoid; DM = dura mater; EP = epineurium; En = endoneurium; P = pia mater; RS = root sheath (Haller and Low, 1971).

which have no branches also taper along their proximal to distal course (Smith, 1992). References Aguayo, AJ, Bray, GM, Terry, LC and Sweezy, E (1976) Three dimensional analysis of unmyelinated fibers in normal and pathologic autonomic nerves. Neuropathol. Exp. Neurol. 35: 136–151. Allt, G and Lawrtenson, JG (2000) The blood nerve barrier: enzymes, transporters and receptors – a comparison with the blood-brain barrier. Brain Res. Bull., 52: 1–12. Arroyo, EJ and Scherer, SS (2000) On the molecular architecture of myelinated fibers. Histochem. Cell Biol., 113: 1–18. Bergoffen, J, Scherer, SS, Wang, S, Scott, MO, Bone, LJ, Paul, DL, Chen, K, Lensch, MW, Chance, PF and Fischbeck, KH (1993) Connexin mutations in X-linked Charcot–Marie–Tooth disease. Science, 262: 2039–2042. Berthold, C-H and Skoglund, S (1967) Histochemical and ultrastructural demonstration of mitochondria in the paranodal region of developing feline spinal roots and nerves. Acta Soc. Med. Ups. 72: 37–70.

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Berthold, C-H, Carlstedt, T and Corneliuson, O (1993) The central-peripheral transition zone. In: PJ Dyck, PK Thomas, JW Griffin, PA Low and JF Poduslo (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, p. 243. Berthold, C-H (1996) Development of nodes of Ranvier in feline nerve: an ultrastructural presentation. Microscop. Res. Tech., 34: 399–421. Bessou, P and Perl, ER (1969) Response of cutaneous sensory units with unmyelinated fibers to noxious stimuli. J. Neurophysiol., 32: 1025–1043. Bischoff, A and Moor, H (1967) Ultrastructural differences between the myelin sheaths of peripheral nerve fibers and CNS white matter. Z. Zellforsch., 81: 303–310. Boyd, IA and Davey, MR (1968) Composition of Peripheral Nerves. Edinburgh, Livingstone. Brown, A (2003) Axonal transport of membranous and nonmembranous cargoes: a unified perspective. J. Cell Biol., 160: 817–821. Burgess, PR, Petit, D and Warren, RM (1968) Receptor types in cat hairy skin supplied by myelinated fibers. J. Neurophysiol., 31: 833–848. Chao, MV (2003) Retrograde transport redux. Neuron, 39: 1–2. Cheng, C and Zochodne, DW (2002) In vivo proliferation, migration and phenotypic changes of Schwann cells in the presence of myelinated fibers. Neuroscience, 115: 321–329. Cravioto, H (1965) The role of Schwann cells in the development of human peripheral nerves. An electron microscopic study. J. Ultrastruct. Res., 12: 634–651. Creange, A, Barlovatzmeimon, G and Gherardi, RK (1997) Cytokines and peripheral nerve disorders. Europ. Cytokine Network, 8: 145–151. Daube, JR (1996) Compound muscle action potentials. In: JR Daube (Ed.), Clinical Neurophysiology, F.A. Davis, Philadelphia, p. 214. Engel, AG and Franzini-Armstrong, C (2004) Myology. McGraw-Hill, San Francisco, pp. 3–595. Fabbretti, E, Edomi, P, Brancolini, C and Schneider, C (1995) Apoptotic phenotype induced by overexpression of wild-type gas3/PMP22: its relation to the demyelinating peripheral neuropathy CMT1A. Genes Dev., 9: 1846–1856. Farel, PB (1978) Reflex activity of regenerating frog spinal motorneurons. Brain Res., 158: 331–341. Finean, JB (1958) X-ray diffraction studies of the myelin sheath in peripheral and central nerve fibers. Exp. Cell Res. Suppl., 5: 18–32.

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Folpe, AL, Billings, SD, McKenney, JK, Walsh, SV, Nusrat, A and Weiss, SW (2002) Expression of claudin-1, a recently described tight junction-associated protein, distinguishes soft tissue perineurioma from potential mimics. Am. J. Surg. Path., 26: 1620–1626. Gasser, HS and Erlanger, J (1927) The role played by the sizes of the constituent fibers of a nerve trunk in determining the form of its action potential wave. Am. J. Physiol., 80: 522. Gasser, HS (1941) The classification of nerve fibers. Ohio J. Sci., 41: 145. Gasser, HS (1955) Properties of dorsal root unmedullated fibers on the two sides of the ganglion. J. Gen. Physiol. 38: 709–728. Ghabriel, MN and Allt, G (1981) Incisures of Schmidt-Lanterman. Prog. Neurobiol. 17: 25–58. Gong, Y, Tagawa, Y, Lunn, MPT, Laroy, W, HefferLauc, M, Li, CY, Griffin, JW, Schnaar, RL and Sheikh, KA (2002) Localization of major gangliosides in the PNS: implications for immune neuropathies. Brain, 125: 2491–2506. Gutrecht, JA and Dyck, PJ (1970) Quantitative teased fiber and histologic studies of human sural nerve during postnatal development. J. Comp. Neurol. 138: 117–129. Haller, FR and Low, FN (1971) The fine structure of the peripheral nerve root sheath in the subarachnoid space in the rat and other laboratory animals. Am. J. Anat., 131: 1–19. Henneman, E, Somjen, G and Carpenter, DO (1965) Excitability and inhibitability of motorneurons of different sizes. J. Neurophysiol., 28: 599–620. Hirose, T, Tani, T, Shimada, T, Ishizawa, K, Shimada, S and Sano, T (2003) Immunohistochemical demonstration of EMA/Glut1-positive perineurial cells and CD34-positive fibroblastic cells in peripheral nerve sheath tumors. Modern Pathol., 16: 293–298. Hiscoe, HB (1947) Distribution of nodes and incisures in normal and regenerated nerve fibers. Anat. Rec., 99: 447. Hoffman, PN and Griffin, JW (1993) The control of axon caliber. In: PJ Dyck, PK Thomas, JW Griffin, PA Low, JF Poduslo (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, pp. 389–402. Hursh, JB (1939) Conduction velocity and diameter of nerve fibers. Am. J. Physiol., 127: 131. Jetten, AM and Suter, U (2000) The peripheral myelin protein 22 and epithelial membrane protein family. Prog. Nucleic Acid Res. Mol. Biol., 64: 97–129.

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Kernie, SG and Parada, LF (2000) The molecular basis for understanding neurotrophins and their relevance to neurologic disease. Arch. Neurol., 57: 654–657. Key, A and Retzius, G (1876) Studien in der Anatomie des Nervensystems und des Bindegewebes. Samson & Wallin, Stockholm. Lambert, EH and Dyck, PJ (1984) Compound action potentials of sural nerve in vitro in peripheral neuropathy. In: PJ Dyck, PK Thomas, EH Lambert, R Bunge (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, p. 1030. Lanterman, AJ (1877) Über den feineren Bau der markhaltigen Nervenfasern. Arch. Mikrosk Anat. Entwicklungsmech. 13: 1. Light, AR and Perl, ER (1993) Peripheral sensory systems. In: PJ Dyck, PK Thomas, JW Griffin, PA Low, JF Poduslo (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, pp. 149–165. Low, PA, Vernino, S and Suarez G (2003) Autonomic dysfunction in peripheral nerve disease. Muscle Nerve, 27: 646–661. Martini, R (2001) The effect of myelinating Schwann cells on axons. Muscle Nerve, 24: 456–466. Meiggioli, MN, Howard, JF and Harper, CM (2004) Neuromuscular Junction Disorders. Marcel Dekker, New York, p. 3. Midroni, G and Bilbao, JM (1995) Normal anatomy of peripheral (sural) nerve. In: Biopsy Diagnosis of Peripheral Neuropathy. Butterworth-Heinemann, Boston, p. 13. Mirsky, R, Jessen, KR, Brennan, A, Parkinson, D, Dong, Z, Meier, C, Parmantier, E and Lawson, D (2002) Schwann cells as regulators of nerve development. J. Physiol. Paris, 96: 17–24. Mugnaini, E, Osen, KK, Schnapp, B and Friedrich, VL (1977) Distribution of Schwann cell cytoplasm and plasmalemmal vesicles (caveolae) in peripheral myelin sheaths. An electron microscopic study with thin sections and freeze-fracturing. J. Neurocytol., 6: 647–668. Mustafa, GY and Gamble, HJ (1979) Changes in axonal numbers in developing human trochlear nerve. J. Anat., 128: 323–330. Nageotte, J (1910) Incisures de Schmidt-Lanterman et protoplasma des cellules de Schwann. C R Acad Sci (Paris), 68: 39. Napolitano, L and Scallen, TJ (1969) Observations on the fine structure of peripheral nerve myelin. Anat. Rec, 163: 1–6. Nolano, M, Provitera, V, Crisci, C, Stancanelli, A, Wendelschafer-Crabb, G, Kennedy, WR and

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Santoro, L (2003) Quantification of myelinated endings and mechanoreceptors in human digital skin. Ann. Neurol., 54: 197–205. Ohcoa, J (1970) Isoniazid neuropathy in man: quantitative electron microscopic study. Brain, 93: 831–850. Ochoa, J (1971) The sural nerve of the human foetus: electron microscope observations and counts of axons. J. Anat., 108: 231–245. Ochoa, J, Fowler, JT and Gilliatt, RW (1972) Anatomical changes in peripheral nerves compressed by a pneumatic tourniquet. J. Anat., 113: 433–455. Olsson, Y, Kristensson, K and Klatzo, J (1971) Permeability of blood vessels and connective tissue sheaths in the peripheral nervous system to exogenous proteins. Acta Neuropathol., Suppl. 5: 61–69. Olsson, Y (1984) Vascular permeability in the peripheral nervous system. In: PJ Dyck, PK Thomas, EH Lambert, R Bunge (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, p. 579. O’Sullivan, DJ and Swallow, M (1968) The fibre size and content of the radial and sural nerves. J. Neurol. Neurosurg. Psychiatry, 31: 464–470. Peters, A, Palay, SL and Webster, HdF (1991) The cellular sheaths of neurons. In: The Fine Structure of the Nervous System: Neurons and their Supporting Cells., Oxford University Press, New York, p. 212. Pleasure, D (1984) The structural proteins of peripheral nerve. In: PJ Dyck, PK Thomas, EH Lambert, R Bunge (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, p. 441. Poduslo, JF (1984) Glycoproteins of the peripheral nervous system. In: PJ Dyck, PK Thomas, EH Lambert, R Bunge (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, p. 494. Polydefkis, M, Griffin, JW and McArthur, J (2003) New insights into diabetic polyneuropathy. JAMA, 290: 1371–1376. Ranvier, L (1871) Recherches de l’histologie et la physiologie des nerfs. Arch. Physiol. I, Ser 4: 129. Ranvier, L (1878) Leçons sur l’histologie du systems nerveux. Paris, F. Savy. Rosenbluth, J (1976) Intramembranous particle distribution at the node of Ranvier and adjacent axolemma in myelinated axons of the frog brain. J. Neurocytol., 5: 731–745. Saito, A and Zacks, SI (1969) Ultrastructure of Schwann and perineurial sheaths at the mouse neuromuscular junction. Anat. Rec., 164: 379–390. Sanes, JR, Apel, ED, Burgess, RW, Emerson, RB, Feng, G, Gautam, M, Glass, D, Grady, RM, Krejci, E, Lichtman, JW, Lu, JT, Massoulie, J, Miner, JH,

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Moscoso, LM, Nguyen, Q, Nichol, M, Noakes, PG, Patton, BL, Son, YJ, Yancopoulos, GD and Zhou, H (1998) Development of the neuromuscular junction: genetic analysis in mice. J. Physiol. (Paris), 92: 167–172. Schmidt, HD (1874) On the structure of the dark or double-bordered nerve fiber. Mon Microsc. J. (Lond.), 11: 200. Schnapp, B and Mugnaini, E (1978) Membrane architecture of myelinated fibers as seen by freeze-fracture. In: SG Waxman (Ed.), Physiology and Pathobiology of Axons, Raven Press, New York, p. 83. Scholzen, T, Armstrong, CA, Bunnett, NW, Luger, TA, Olerud, JE and Ansel, JC (1998) Neuropeptides in the skin: interactions between the neuroendocrine and the skin immune systems. Exp. Dermatol., 7: 81–96. Shantha, TR and Bourne, GH (1968) The perineural epithelium – a new concept. In: GH Bourne (Ed.), The Structure and Function of Nervous Tissue, Academic Press, New York, Vol. 1, p. 379. Shanthaveerappa, TR and Bourne, GH (1962) The “perineural epithelium,” a metabolically active, continuous, protoplasmic cell barrier surrounding peripheral nerve fasciculi. J. Anat., 96: 527. Shield, LK, King, RH and Thomas, PK (1986) A morphometric study of human fetal sural nerve. Acta Neuropathol. (Berl.), 70: 60–70. Smith, BE and Dyck, PJ (1992) Subclinical histopathological changes in the oculomotor nerve in diabetes mellitus. Ann. Neurol., 32: 376–385. Stewart, JD (1987) The Structure of the Peripheral Nervous System. In: Focal Peripheral Neuropathies, Elsevier, New York, p. 6. Stewart, JD (2003) Peripheral nerve fascicles: anatomy and clinical relevance. Muscle Nerve, 28: 525–541. Sunderland, S (1965) The connective tissues of peripheral nerves. Brain, 88: 841–854. Sunderland, S and Bradley, KC (1949) The cross-sectional area of peripheral nerve trunks devoted to nerve fibers. Brain, 72: 428–449. Takashima, H, Boerkoel, CF, De Jonghe, P, Ceuterick, C, Martin, JJ, Voit, T, Schroder, JM, Williams, A, Brophy, PJ, Timmerman, V and Lupski, Jr (2002) Periaxin mutations cause a broad spectrum of demyelinating neuropathies. Ann. Neurol., 51: 709–715. Thomas, PK (1963) The connective tissue of peripheral nerve: an electron microscope study. J. Anat., 97: 35–44.

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Thomas, PK and Ochoa, J (1984) Microscopic anatomy of peripheral nerve fibers. In: PJ Dyck, PK Thomas, EH Lambert, R Bunge (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, p. 39. Thomas, PK and Ochoa, J (1984) Microscopic anatomy of peripheral nerve fibers. In: PJ Dyck, PK Thomas, EH Lambert, R Bunge (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, p. 41. Thomas PK and Ochoa J (1984) Microscopic anatomy of peripheral nerve fibers. In: PJ Dyck, PK Thomas, EH Lambert, R Bunge (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, p. 51. Thomas, PK and Ochoa, J (1984) Microscopic anatomy of peripheral nerve fibers. In: PJ Dyck, PK Thomas, EH Lambert, R Bunge (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, p. 57. Thomas, PK and Olsson, Y (1984) Microscopic anatomy and function of the connective tissue components of peripheral nerve. In: PJ Dyck, PK Thomas, EH Lambert, R Bunge (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, p. 97. Thomas, PK, Berthold, C-H and Ochoa, J (1993) Microscopic anatomy of the peripheral nervous system. In: PJ Dyck, PK Thomas, JW Griffin, PA Low, JF Poduslo (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, pp. 28–91. Thomas, PK, Berthold, C-H and Ochoa, J (1993) Microscopic anatomy of the peripheral nervous system. In: PJ Dyck, PK Thomas, JW Griffin, PA Low, JF Poduslo (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, p. 60. Thomsen, R (1887) Ueber eigenthumliche aus veranderten Ganglienzellen hervorgegagene Gebilde is den Stammen der Hirnnerven des Menschen. Virchows Arch., 109: 459. Webster, HD (1993) Development of peripheral nerve fibers. In: PJ Dyck, PK Thomas, JW Griffin, PA Low, JF Poduslo (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, pp. 243–266. Williams PL and Landon DN (1964) The energy source of the nerve fibre. New Sci., 21: 166. Williams, PL, Warwick, R, Dyson, M and Bannister, LH (1989) Structure of the peripheral nervous system. In: Gray’s Anatomy. Churchill Livingstone, New York, p. 896. Willison, HJ and Yuki, N (2002) Peripheral neuropathies and antiglycolipid antibodies. Brain, 125: 2591–2625. Yao, JK (1984) Lipid composition of normal and degenerating nerve. In: PJ Dyck, PK Thomas, EH

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Lambert, R Bunge (Eds.), Peripheral Neuropathy, W.B. Saunders, Philadelphia, p. 510. Yin, X, Crawford, TO, Griffin, JW, Tu, P, Lee, VM, Roder, J and Trapp, BD (1998) Myelin-associated glycoprotein is a myelin signal that modulates the caliber of myelinated axons. J. Neurosci., 18: 1953–1962. Yoshikawa, H and Dyck, PJ (1991) Uncompacted inner myelin lamellae in inherited tendency to pressure palsy. J. Neuropathol. Exp. Neurol., 50: 649–657.

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Zhou, L and Griffin JW (2003) Demyelinating neuropathies. Curr. Opin. Neurol., 16: 307–313. Zoidl, G, Blass-Kampmann, S, D’Urso, D, Schmalenbach, C and Muller, HW (1995) Retroviral-mediated gene transfer of the peripheral myelin protein PMP22 in Schwann cells: modulation of cell growth. EMBO J., 14: 1122–1128.

Peripheral Nerve Diseases Handbook of Clinical Neurophysiology, Vol. 7 J. Kimura (Ed.) © 2006 Elsevier B.V. All rights reserved

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CHAPTER 2

Physiology and function Christian Krarup* Department of Clinical Neurophysiology, the Neuroscience Center, Rigshospitalet, Denmark

2.1. Introduction The purpose of the clinical neurophysiological examination is to localize, identify, and specify pathophysiological abnormalities as consistent with disease affecting the peripheral nervous system (PNS), neuromuscular transmission or muscle and after integration with clinical and other paraclinical information to establish a specific diagnosis. To accomplish these goals, it is necessary to have a clear notion of the physiological characteristics of the individual systems, which methods are needed to study these systems, and how particular deviations from normal function reflect pathological abnormalities. In addition, recently it has become apparent that axon membrane function is fundamental for phenomena such as fasciculations, cramps and neuromyotonia (NewsomDavis, 1997; Mogyoros et al., 1997, 1998; Heidenreich and Vincent, 1998; Vincent et al., 1998; Kiernan et al., 2001b). These goals are by no means trivial since the pathophysiological correlates of pathological disturbances rarely are specific. It is, therefore, necessary to understand the limitations of the physiological methods and apply an integrated approach using different criteria to arrive at a comprehensive picture of the disorder. This chapter aims at discussing the physiology and function of peripheral nerve to assist the reader in understanding the methods and techniques covered in this section, which are used to investigate the different parts of the peripheral nervous system, whereas it is not intended to deal with these topics in detail. The main outcome of the neurophysiological studies is to determine whether there is loss of nerve fibers or whether their function is abnormal or both. The main parameters in routine

* Correspondence to: Christian Krarup MD DMSc FRCP Department of Clinical Neurophysiology NF3063, Rigshospitalet, Blegdamsvej 9, 2100 Copenhagen, Denmark E-mail address: [email protected]. Tel: +45 3545 3060; fax: +45 3545 3264.

electrophysiological studies include conduction velocity and amplitudes of the evoked responses, and it is necessary to consider to what extent abnormalities associated with these parameters may be used to determine loss or abnormalities of function. In recent years, we have seen a marked development of insights into basic function and structure of the axon, glial cells and ion-channels and methodologies have been developed to investigate abnormalities in these structures in patients. In the future, we are likely to see these methods as integrated tools in clinical settings. Several textbooks describe the techniques of conduction studies in motor and sensory nerve fibers (Kimura, 1989; Brown and Bolton, 1993; Binnie et al., 1995; Aminoff, 1998; Daube, 2002). 2.2. Anatomy, structure, and function of peripheral nerve Peripheral nerves are defined as the part of the nervous system situated outside the central nervous system (CNS) demarcated by the leptomeninges. The glia surrounding axons of the PNS is derived from Schwann cells, and in the CNS from oligodendrocytes (Kleitman and Bunge, 1995). The division of the nervous system into the PNS and the CNS is to some extent artificial since parts of peripheral neurons are situated within the CNS. Only the thin unmyelinated postganglionic autonomic fibers are located wholly outside the CNS (Berthold and Rydmark, 1995). Even though peripheral nerve disorders by definition are located to the PNS, a number also affect the central parts of the PNS (Asbury and Johnson, 1978; Berger and Schaumburg, 1995). 2.2.1. Classification of fibers of the PNS Peripheral nerve fibers are classified according to morphology and function. The afferent system consists of myelinated and unmyelinated fibers that innervate different types of sensory receptors and accordingly convey different sensory modalities (see Kandel et al., Principles of Neural Science, 4th edition, 2000 (Gardner

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et al., 2000)). Myelinated fibers are grouped according to diameters into types Aα-δ in cutaneous nerves (touch, vibration, temperature, pain), whereas they are named I-III in muscle nerves (proprioception, pressure, pain) (Pearson and Gordon, 2000), and unmyelinated fibers consist of type C in cutaneous nerves and type IV in muscle nerves that innervate pain and temperature receptors. It is now known that unmyelinated fibers have different subtypes (Serra et al., 1999). The efferent system is divided in myelinated α-, γ- and δ-fibers that innervate extrafusal muscle fibers, muscle spindles and both, respectively. Autonomic postganglionic fibers are unmyelinated. In myelinated fibers (Fig. 2.1), each axon is surrounded by the specialized membrane of trains of single Schwann cells wrapped around the axon in lamellae (Fig. 2.1A, insert), and each Schwann cell is divided from the next at the node of Ranvier. In contrast, unmyelinated fibers are arranged in groups of 1–50 axons ensheathed in Schwann cells (referred to as Remak fiber, Fig. 2.1B). Each myelinated fiber and group of unmyelinated fibers are delimited from the endoneurium by a basal lamina (Fig. 2.1) (Behse, 1990; Berthold and Rydmark, 1995). The number of unmyelinated fibers is much greater than myelinated fibers (20 000–50 000 in the sural nerve) but the proportion of unmyelinated to myelinated fibers varies in different types of nerves (Ochoa and Mair, 1969; Behse, 1990). The distribution of motor and sensory myelinated fibers in humans is bimodal, with motor fibers classified as small 1–8 μm γ-fibers, and large 8–15 μm α-fibers (Lee et al., 1975), and sensory fibers ranging from small 1 μm Ad fibers to large 18 μm Aα and Aβ fibers (Fig. 2.2) (Buchthal and Rosenfalck, 1966; Behse, 1990; Caruso et al., 1992; Berthold and Rydmark, 1995). Unmyelinated fibers have small diameters of 1–2 μm.

Fig. 2.1 Myelinated (A) and unmyelinated (B) fibers from the sural nerve. A single axon is surrounded by compact myelin lamellae (A, inset); several axons are ensheathed by Schwann cell process in unmyelinated fibers. BM, basal lamina; *, axons; SC, Schwann cell processes. In (A) the thick straight arrow points at the inner mesaxon; the curved arrow points at the outer mesaxon (Courtesy of H. Schmalbruch, University of Copenhagen).

CHRISTIAN KRARUP

Individual myelinated fibers are subdivided into specialized regions (Berthold and Rydmark, 1995; Arroyo and Scherer, 2000; Peles and Salzer, 2000; Scherer and Arroyo, 2002) consisting of the node of Ranvier with a width of about 1 mm and the internodal segment with a length of 300–2000 mm proportional to the fiber diameter (Vizoso and Young, 1948). The myelin thickness and the number of lamellae are also proportional to the fiber caliber, and the ratio of axon/fiber diameter (g-ratio) is reasonably constant about 0.6, at least among the larger myelinated fibers (Boyd and Davey, 1968; Arbuthnott et al., 1980b; Behse, 1990). A g-ratio of about 0.6 is recovered in regenerated nerve though the proportionality between fiber diameter and internodal length is not reestablished (Vizoso and Young, 1948; Hildebrand et al., 1987). A constant relationship between the surface area and the myelin volume was found in normal, regenerated and remyelinated fibers (Smith et al., 1982a). 2.2.2. Molecular organization of myelinated fibres The specialized organization has important implications for the function of the nerve fiber, and the nodes of Ranvier and paranodes are formed by interaction between myelin and axon (Scherer, 1996) that occur during development (Vabnick and Shrager, 1998). The distribution of current along the axon before, during and after the action potential is determined by its biophysical passive properties and by the function and distribution of ion-channels (Waxman and Ritchie, 1985; Arroyo et al., 1999; Arroyo and Scherer, 2000; Scherer and Arroyo, 2002, 2004), which have many features in common in most mammals, including human nerve fibers (Scholz et al., 1993; Reid et al., 1999). During acute demyelination, the ability to propagate action

PHYSIOLOGY AND FUNCTION

25

Number of fibers

Mean fiber diameter distribution from normal sural nerve (n =10) (Behse, 1990) 2000 1800 1600 1400 1200 1000 800 600 400 200 0

Mean number of fibers = 6940 (range 5075−9460)

1

2

3

4

5

6 7 8 Fiber diameter (μm)

9

10

11

12

13

Fig. 2.2 Fiber diameter distribution from 10 normal sural nerves. The total number of myelinated fibers was calculated from sampling of 200–300 fibers in whole nerve biopsies multiplied by the total fascicular area. The bars indicate the mean number of fibers in each bin (1 μm) and the error bars indicate the standard deviation Calculated from data in Behse (1990).

potentials is impaired and the block of conduction may occur (Lafontaine et al., 1982). Nevertheless, continuous conduction can occur along demyelinated internodes, indicating that the internodal membrane is excitable (Bostock and Sears, 1978). Conduction along demyelinated internodes is insecure, and broadening of the action potential improves the safety factor (Bostock et al., 1978, 1981). Consistent with this internodal excitability was the increased number of Na+ channels, probably formed de novo rather than through redistribution (England et al., 1991), and immunocytochemistry studies have shown that incorporation of channels occurs at sites of axon-Schwann cell interaction (Dugandzija-Novakovic et al., 1995). After demyelination, saltatory-like conduction was resumed before remyelination, which suggested that increased concentration of Na+ channels occurred at future nodes of Ranvier (Smith et al., 1982b). The inward current during the action potential is carried by Na+ ions through specific voltage-gated channels. At the node of Ranvier, Na+ channels are concentrated at a density of 1000–2000 channels/μm2, whereas the density at the internode is only 4% of this at about 25 channels/μm2 (Barchi, 1995; Waxman, 1995). Even though the total number of Na+ channels at the internode is larger than at the node (Shrager, 1989), their low density is insufficient to carry action potentials during acute demyelination. Na+ channels are closed at the resting membrane potential but open at depolarization to threshold (activation) reaching a high conductance, each channel carrying as much as 10 000 ions during a single impulse. During this

inward current, the membrane potential approaches the reversal potential of Na+ of about +40 mV but repolarizes due to rapid inactivation of the channel in spite of depolarization. The Na+ channel activation by depolarization is associated with movement of an intramembraneous gating-charge (Dubois and Schneider, 1981; Chiu and Ritchie, 1981b; Ritchie, 1995), which opens the channel and which is blocked during inactivation. Voltage-gated ion-channels are composed of subunits, and K+, Na+, and Ca++ channels form part of a superfamily with six membrane-spanning segments of the subunit. The Na+ channel consists of a pore forming α-subunit with more than 2000 aminoacids and a MW of about 260 kDa and 24 transmembraneous segments (S1-6) in four domains (I-IV) (Fig. 2.3). The S4 segment is the likely voltagesensitive factor associated with charge movement during depolarization and the channel protein is subject to conformational changes during depolarization (Catterall, 1992). Cytoplasmic parts of the channel are involved in inactivation, and in addition to the a-subunit, the structure includes β-subunits that have MW in the order of 40 kDa and provide auxiliary functions for the ion permeability. Thus the a-subunits allow Na+ flux but co-expression of β1- and β2-subunits increases the activation and the inactivation rates (Catterall, 2000). More than 10 genes encode for several isoforms of voltage-gated Na+ channels (Goldin, 1999; Goldin et al., 2000), denoted as Nav1 for the first gene subfamily, and include 9 types. Nav1.6 is the isoform most widely distributed in mammalian motor and sensory fibers of both the CNS and the PNS

26

CHRISTIAN KRARUP

A

Na+

Exracellular Direction of passive movement for Na+

ΔV Intracellular Closed channel

Open channel

B P segment I Out

II + +

1 2 3 4 5

+ + 6

IV

+ +

1 2 3 4 5

+ +

In

III

6

1 2 3 4 5

+ +

+ + 6

1 2 3 4 5

+ +

6

+ + COOH

NH2

C

II

Na+

III

SIDE VIEW

II

III

I

IV

TOP VIEW I

IV

Selectivity filter

COOH

NH2 +

Fig. 2.3 Diagrammatic representation of axonal Na channels. (A) closed and open voltage gated channels inserted in the axolemma. (B) the α-subunit consisting of four domains (I-IV) each with 6 (1-6) membrane spanning segments. The S4 segment is the likely voltage sensitive factor and the S5-6 segments the pore forming parts of the channel. (C) the folded subunit as arranged in the axolemma (With permission from Hammond: Cellular and Molecular Neurobiology, © Academic Press.).

(Caldwell et al., 2000; Kearney et al., 2002), and mice lacking the gene that encode for the Nav1.6 have severe neurological disease (Kearney et al., 2002). Nevertheless, nodes of mice lacking Nav1.6 can conduct action potentials and show Na+ channels that stain for a panspecific channel type, indicating that there are several isoforms at the node of Ranvier (Caldwell et al., 2000; Peles and Salzer, 2000). In developing fibers there is a shift from Nav1.2 to Nav1.6 isoform during myelination, whereas this switch does not occur in unmyelinated fibers (Boiko et al., 2001; Kaplan et al., 2001; Kazarinova-Noyes and Shrager, 2002). The great majority of Na+ channels are tetrodotoxin (TTX) sensitive. TTX-resistant Nav1.9 channels also occur in dorsal root ganglia and may be involved in pain sensation (Dib-Hajj et al., 2002; Fang et al., 2002). These channels are

down-regulated and the TTX-sensitive Nav1.3 channel is up-regulated after neural trauma (Cummins and Waxman, 1997; Cummins et al., 2001), indicating that the channel isoforms may be modified in the adult mammalian organism. Although most Na+ channels are rapidly inactivating, 1–3% remain active at more positive membrane potentials (so-called persistent Na+ channels) that may arise from different types of channels or be due to a modality shift of the same isoform (Brown et al., 1994; Crill, 1996; Baker and Bostock, 1997). The segregation of Na+ channels at the nodal membrane (Ritchie and Chiu, 1981) may be explained by different mechanisms including physical restriction, soluble factors, cytoskeletal anchoring or molecules in the extracellular matrix (Barchi, 1995; Waxman, 1995; Peles and Salzer, 2000). The nodal gap is limited by the paranodal loops, and the Na+

PHYSIOLOGY AND FUNCTION

27

channels form complexes with several membrane proteins and extracellular matrix components directly or through ankyrin G, and the beta-units may contribute to the anchoring and segregation of Na+ channels to the nodal membrane (Fig. 2.4) (Davis et al., 1996; Arroyo and Scherer, 2000; Catterall, 2000; Peles and Salzer, 2000; Kazarinova-Noyes and Shrager, 2002; Scherer and Arroyo, 2002). In myelinated amphibian and mammalian, including human nerve fibres, there exist one type of Na+ channel and at least three types of K+ channels (Jonas et al., 1989; Scholz et al., 1993; Barchi, 1995; Vogel and Schwarz, 1995; Reid et al., 1999), fast, intermediate and slowly activated K+ channels. Fast activated K+

channels are blocked by 4-aminopyridine (4AP), and are located in the juxtaparanode in mammalian fibers, whereas the slowly activated K+ channels are located in the node and the internode and may be blocked by tetraethylammonium (TEA). The fast-activated K+ channels in mammalian peripheral nerve belong to the Kv1.1 and Kv1.2 alpha-subunits and the Kvβ2 subunit and consist of 4 membrane-spanning domains similar in configuration to the Na+ channel (Fig. 2.3) localized beneath the myelin and together with the protein Caspr2 bind to the myelin protein P0 (Arroyo et al., 1999; Rasband and Trimmer, 2001; Scherer and Arroyo, 2002) and participate in the formation of axoglial junctions (Arroyo and Scherer, 2000).

Kv1.5

Compact Myelin

Slo1

Microvilli Adaxonal Membrane ?

actin

?

?

Paranodal Myelin Loop

TAG-1

?

Ezrin Radixin Moesin

KIR2.3 KIR2.1 Caspr2

?

Kvβ2

PDZ?

pr

NF Band 4.1B

Kv3.1b

KCNQ2

Ca s

n cti nta Co

Kv1.2

β1

AnkyrinG

Tenascin-R

Connexin29

NF186

Kv1.1

Nr-CAM

Connexin32

Tenascin-C

15

5

?

α

β2

Spectrin βIVΣ1 Na+ Channel

Fig. 2.4 Specialization at nodes and paranodes of myelinated nerve fibers in the CNS and the PNS. The voltage gated Na+ are concentrated mainly at the node and the K+ at the juxtaparanode. Na+ channels are linked to the spectrin cytoskeleton via ankyrinG. The precise role of other molecules at the node are yet to be defined: the β subunit may interact with tenascin. The paranodal loops form the axoglial junction and are bound to the axolemma by tight junctions (Caspr). K+ channels are located at the juxtaparanode and tethered to the cytoskeleton via Caspr2 molecules (From Scherer et al., (2004) with permission).

28

The myelin is organized into compact and non-compact regions at the internode and the paranode, respectively, and contain a non-overlapping set of proteins (Arroyo and Scherer, 2000). In the PNS, the most abundant protein P0 is involved in the compacting of myelin at the internode as does the next most abundant protein myelin basic protein, MBP (Monuki and Lemke, 1995). Peripheral myelin protein 22 (PMP-22) is a transmembraneous protein localized to compact myelin considered to be a regulator of axon-glial interactions (Naef and Suter, 1998). As P0, myelin-associated glycoprotein (MAG) belongs to the Ig superfamily of neural adhesion molecules and is localized to the non-compacted portion of myelin, including the Schmidt-Lantermann incisures, the paranodal loops, and the inner and outer mesaxon. In addition, the non-compacted myelin contains a number of other molecules, connexin 32 and E-cadherin that link to the actin cytoskeleton and may have junction properties and participate in the intracellular traffic. Mutations in the genes that code for these myelin proteins are responsible for several inherited neuromuscular disorders. 2.2.3. Ion-channels and membrane potential The Hodgkin–Huxley model of generation of the action potential followed by repolarization included successive activation-deactivation of Na+ channels and of K+ channels that repolarize (outward rectification) the membrane following the action potential (Hodgkin and Huxley, 1952). The model was applicable in unmyelinated and myelinated amphibian nerve fibers (Frankenhaeuser and Huxley, 1964). The Na+ channels are voltage-gated and activated by depolarization of the membrane potential by about 10–20 mV even though some Na+ channels are inactivated at the resting membrane potential of 70–80 mV. Voltage clamp studies and the lack of effect of 4-AP on the action potential, however, have shown that K+ ions do not contribute to the repolarization in mammalianmyelinated fibers (Chiu et al., 1979; Brismar, 1980; Sherratt et al., 1980; Bostock et al., 1981). In contrast, during paranodal demyelination with access to the axonal membrane sealed under the myelin, the function of fast-activated K+ channels became apparent (Bostock et al., 1978; Brismar, 1979; Chiu and Ritchie, 1980, 1981a), and insight into this has stimulated attempts to ascertain the effect of broadening the action potential in patients with demyelinating neuropathy to improve propagation (Russell et al., 1995; Franssen et al., 1999). These electrophysiological findings are in keeping with the segregation of fast activated K+ channels to the

CHRISTIAN KRARUP

paranode, whereas they in the main lack at the node. The function of the paranodal K+ channels is to stabilize the membrane potential at the node and thus dampen depolarizations and avoid spontaneous activity (Waxman, 1995). In regenerated myelinated fibers, fastactivated K+ channels appear to influence the recovery after activity, as indicated by the effect of blocking fastactivated K+ channels (Gordon et al., 1991). The repolarization following the action potential in myelinated mammalian fibers is, therefore, due to rapid inactivation of Na+ channels, and an ohmic leak through the nodal membrane. The repolarization is delayed by a capacitative current from the paranodal region that has been charged during the action potential via a low-resistive pathway underneath the myelin, the so-called Barrett–Barrett pathway (Barrett and Barrett, 1982). The discharge through the Barrett–Barrett resistance is associated with the depolarizing afterpotential, which is dependent on the membrane potential and lasts several ms and is the mechanism behind the supernormal period (Barrett and Barrett, 1982; Blight, 1985; Bowe et al., 1987; David et al., 1995; McIntyre et al., 2002). In addition to the fast inactivated Na+ channels, 2–3% of Na+ channels remain open during depolarization (so-called persistent Na+ channels). Persistent Na+ channels are more numerous in sensory rather than motor fibers and responsible for the longer strength-duration constant in sensory than motor fibers (Baker and Bostock, 1997; Bostock and Rothwell, 1997). The slow K+ channels at the node and internode are not activated during the action potential but cause a late hyperpolarization associated with late subexcitability (Bergmans, 1973; Bostock and Bergmans, 1994) and they are blocked by TEA (Baker et al., 1987; Kocsis et al., 1987). The K+ channels have outward rectification properties. In addition, there are ion-channels at the internode, which are permeable to both Na+ and K+, that open during hyperpolarization, that are blocked by Cs+ and have inward-rectifying properties; these channels may protect the axon to the hyperpolarizing effect of prolonged activity and the consequent reduction in excitability due to activation of the electrogenic Na+/ K+-ATPase pump (Bostock and Grafe, 1985; Baker et al., 1987). These characteristics of myelinated fibers are important not only to understand the normal nerve fiber function but also for the development of clinical deficits and the disturbances in electrodiagnostic studies in patients with disorders of myelin, axons and neurons (Bostock et al., 1995; Kiernan et al., 1996; Kaji et al., 2000; Daskalova and Stephanova, 2001; Kiernan et al., 2002a, 2002b; Kuwabara et al., 2002b; Kaji, 2003).

PHYSIOLOGY AND FUNCTION

29

2.2.4. Studies of axonal membrane properties in situ The central parameters obtained at nerve conduction studies include the amplitudes and conduction velocities of the CMAP and SNAP and abnormalities are usually interpreted in terms of axonal loss or demyelination. Nevertheless, the conditions of nerve conduction studies do not allow insight into functional aspects of active and passive axonal properties that may be important during motor control and sensory activity and may further explain symptoms, such as fasciculations, cramps and paresthesia (Baker, 2000). Furthermore, slowing of nerve conduction may occur in axonal disorders and A

B Threshold reduction (%)

80 Threshold reduction (%)

may be due to either atrophy or ion-channel abnormalities. Investigation of axonal ion-channel or passive properties, however, requires the use of specialized studies that have been developed to evaluate changes in excitability of axons in situ (Bostock et al., 1998; Kiernan et al., 2000). These methods have been developed to study the effects of accommodation and activity on the axon membrane potential and access internodal properties that are not explicitly tested during nerve conduction studies and reveal differences in motor and sensory fibers (Fig. 2.5) (Kiernan et al., 2001c). The internode as well as the node of Ranvier contain rectifying channels and the electrogenic Na+/K+-ATPase

60 40 20 0 −20

20 0 −20 − 40 − 60 − 80 −100 −120

0

200

0

D

0

− 50

−100 − 400

50 100 150 Conditioning-test delay (ms)

200

60

50 Threshold reduction (%)

Polarizing current (% Threshold)

C

50 100 150 Conditioning-test delay (ms)

− 300

−200

−100

Threshold reduction (%)

40 20 0 −20 − 40

0

10 Interstimulus interval (ms)

100

Fig. 2.5 Measurements of excitability in motor (filled symbols) and sensory (open symbols) nerve fibers from a normal median nerve. The motor responses were evoked by stimulation at wrist and recording at the abductor pollicis brevis muscle and the sensory responses by recording at digit 2. Excitability was tested using the qtrack method (see text) during subthreshold depolarization for 100 ms (A) and 200 ms (C), and during hyperpolarization for 100 ms (B) and 200 ms (C), and following a single supramaximal pulse (D). The effects of depolarization and hyperpolarization differed substantially in motor and sensory fibers suggesting higher activation of internodal K+ channels during depolarization and of inward rectifying internodal channels during hyperpolarization of the latter. Following a single pulse, the recovery cycle showed that the refractory period was longer, the supernormal period at 6–8 ms smaller, and the late subexcitability smaller in sensory than in motor fibers consistent with effects of persistent Na+ channels. (From Moldovan, Lozeron, Krarup, unpublished.).

30

pump that fundamentally impinge on the ability of the fiber to propagate repetitive action potentials, recovery of membrane potential after activity, prevention of ectopic activity, and redistribution of ions following activity (Bostock et al., 1983; Bostock and Grafe, 1985; Kiernan et al., 1996; Zhou et al., 1999; Kiernan et al., 2000; Burke et al., 2001; Kuwabara et al., 2002a). Bostock (for details see Chapter 17) extended studies in experimental animals (Baker et al., 1987; Baker and Bostock, 1989) to the development of thresholdtracking techniques in human nerve (Bostock and Baker, 1988; Bostock et al., 1998). Such studies have shown that the same rules regarding accommodation could be described in myelinated fibers from humans and rats (Baker and Bostock, 1989). It was already found by Bergmans (Bergmans, 1973) in studies conducted on single human motor nerve fibers that single or repetitive activity was followed by changes in excitability, which were interpreted as being due to changes in membrane potential and activation of specific ion-channels and these findings were subsequently confirmed and extended in experimental animals (Bostock and Grafe, 1985). The detection of excitability changes was made feasible by the threshold-tracking technique, which measures the stimulus current necessary to clamp the amplitude of the CAMP at a specified value on the steep part of the stimulus-response relationship curve (Bostock et al., 1998). A wealth of information regarding the behavior of normal and pathological nerve fibers have been obtained using this technique to study strength-duration relationships, recovery of excitability following activity, and accommodation during subthreshold de- and hyperpolarization (Fig. 2.5) (Kiernan et al., 2000; Burke et al., 2001). Thus, studies have shown differences in sensory compared to motor studies regarding strength-duration relationships, recovery of excitability and accommodation to hyperpolarization,which are consistent with differences in persistent Na+ channels at the node and a more effective inward rectification. Changes in excitability have been found in patients with diabetes mellitus, amyotrophic lateral sclerosis, multifocal motor neuropathy, Guillain–Barré syndrome, and chronic inflammatory demyelinating polyneuropathy that raise the possibility that this type of studies may provide supplementary information regarding pathogenesis and functional disturbances (Bostock et al., 1995; Horn et al., 1996; Kiernan et al., 1996; Cappelen-Smith et al., 2000; Kaji et al., 2000; Cappelen-Smith et al., 2001; Cappelen-Smith et al., 2002; Kiernan et al., 2002a; Kuwabara et al., 2002b). In patients with diffuse

CHRISTIAN KRARUP

demyelinating neuropathy such as Charcot-MarieTooth type 1 and some cases of CIDP, the accommodation during electrotonus suggested the presence of membrane hyperpolarization (Sung et al., 2004). Interestingly, in long-term regenerated nerve, similar findings were obtained (Moldovan and Krarup, 2004a, 2004b), and these similarities may be due to hyperpolarization associated with the raised Na+ ions at the increased number of nodes of Ranvier and short internodal length, and subsequent hyperactivity of the electrogenic sodium/potassium pump. 2.3. Physiological studies of the PNS The function of nerve fibers is to propagate action potentials and investigations are based on the ability to elicit and record sensory and motor responses (for details see Chapter 5, Nodera and Kaji). Different methods are used to study single nerve fibers or groups of nerve fibers, sensory or motor fibers, or myelinated or unmyelinated fibers. Each of these methods has specific requirements and limitations due to differences in characteristics of the nerve fibers and their signals that must be considered when used for clinical or research purposes. In the clinical electrodiagnostic setting, studies are confined to large myelinated motor and sensory fibers, and responses evoked by electrical stimulation are derived from groups of fibers. Thus, disorders confined to a small number of fibers or to small fibers in partial lesions may not be detected in these studies. Moreover, the location of abnormalities outside sites accessible to conduction studies or involvement of ion-channels requires methods specifically aimed at elucidating such questions. Finally, the activity studied in the diagnostic set-up is highly artificial with single, spaced stimuli to elicit synchronized responses, whereas the activity of the nervous system during movement or sensation is characterized by bursts of repetitive activity, and the interaction and integration of this activity are better associated with clinical symptoms than single electrical stimuli. 2.3.1. Stimulation Electrical depolarizing stimuli (square pulses, 0.1–0.5 ms in duration, constant current or voltage) are applied through a surface or needle cathode to the motor or sensory fibers with sufficient strength to ensure that all fibers are stimulated above the threshold (Fig. 2.6) (for details see Chapter 5, Kaji). Supramaximal levels at 10–20% above the maximal response are required to

PHYSIOLOGY AND FUNCTION

31

Maximal response

Supramaximal stimulation

25

Response (mV)

20

15 a Response = −c⫻Stimulus 1 + b ⫻e

10 Threshold stimulation 5

Fig. 2.6 Stimulus-response relationship: Tibial nerve in cats is stimulated at ankle and responses are recorded from the plantar muscles via subcutaneous needle electrodes. The 0.2 ms negative rectangular stimuli were increased in 6% steps with two responses averaged for each step (open circles). The stimulus-response relationship (continuous line) is plotted for the 0.5–95% of supramaximal response (broken line). The parameters of the fit (R × R = 0.997) are a = 23.2847, b = 0.4412, c = 7.1630 (From Moldovan unpublished observations).

0 1

1.5

2

2.5

3

3.5

4

Stimulus (mA)

avoid submaximal stimulation due to movement of the electrode, and corresponds in most instances to almost five times the threshold stimulus when using surface electrodes (Lange et al., 1992) but maximal stimulation may be difficult or impossible to attain in demyelinated nerve fibers with low excitability (Meulstee et al., 1997; Krarup, 1999). It is often assumed that the negative relationship between threshold and fiber diameter demonstrated in experimental nerve (Gasser and Erlanger, 1927; Gasser and Grundfest, 1939) also holds for the clinical study; however, the latency of the first recorded motor unit potential is often not the shortest indicating that the anatomical localization of the fibers with respect to the electrical field determines the sequence of excitation (Kimura, 1989, 1997). Moreover, the latency and conduction velocity are also influenced by the stimulus current due to the rate of depolarization and the utilization time (Krarup et al., 1992; Carp et al., 2003). Needle electrode stimulation may be necessary to obtain maximal responses in deeply seated nerves such as motor cervical or lumbosacral plexus, roots or spinal nerves (Inouye and Buchthal, 1977; Liguori et al., 1992). Alternative stimulation methods include magnetic pulse stimulation of spinal roots, plexus or peripheral nerves but require standardization that differ from the use of electrical stimulation (Ravnborg et al., 1990; Ravnborg and Dahl, 1991; Bischoff et al., 1995; Ravnborg, 1996). Needle-electrodes for stimulation have the advantage of lower stimulus strength associated with less displacement of the site of excitation (Podivinski, 1965; Wiederholt, 1970), and less risk of activation of nearby nerves.

Even when stimulating the sensory nerve fibers at distal sites, such as fingers or toes, the use of electrical stimulation does not allow the study of the very distal segment of sensory fibers at their junction with receptors, which might be expected to reveal abnormalities early in the course of axonal neuropathies. There has been strong interest in the microneurographical evaluation of the function of individual sensory nerve fibers innervating receptors in both normal and diseased nerves (Hagbarth, 2002), and such studies have served to clarify the adequate stimulus and distribution of different tactile receptors (Johansson et al., 1980; Johansson and Vallbo, 1979; Knibestöl and Vallbo, 1970). Sampling may, however, be biased in diseased nerve, even though microneurography may show abnormalities in neuropathy (Mackel, 1985). Useful information may be obtained by recording the SNAP obtained by stimulation of groups of tactile receptors (Fig. 2.7), and this method has been used in both normal and diseased nerve to identify different types of receptors and to localize primary pathological changes (Buchthal, 1982a, 1982b; Hashimoto et al., 1989; Caruso et al., 1993; Krarup-Hansen et al., 1993; Krarup and Trojaborg, 1994; Simonetti et al., 1998; Baba et al., 2001). Tactile stimulation does not allow supramaximal activation of all nerve fibers, and sampling bias may, therefore, be a source of error. 2.3.2. Recording of the action potential in situ The arrangement of regions of internodal segments surrounded by myelin with high resistance and low capacitance gives the nerve fibers of small diameters

32 Fig. 2.7 Electrical and tactile stimulation of sensory nerve: Median nerve sensory potentials were evoked by electrical (top traces) or tactile stimuli (bottom traces, rate of indentation 400 μm/ms, indentation depth 300 μm) applied at the tip of digit III and recorded at wrist and elbow. Amplitudes, latencies and conduction velocities are shown below traces. The bottom trace represents the tactile stimulus (From Krarup, 2004).

CHRISTIAN KRARUP

Electrical − 3 μV +

Dig3tip-elbow 8 μV, 67 m/s Dig3tip-wrist

5 μV 14 μV, 3.4 ms, 55 m/s 0

20 ms Tactile T400 − 0.5 μV +

Dig3tip-elbow 1.4 μV, 69 m/s Dig3tip-wrist 1.8 μV, 4.0 ms T400

400 μm/ms, 300 μm

0

of less than 10–12 μm unique capacity to propagate action potentials at high velocity by optimizing the space and the time constants to depolarize the nodal membranes rapidly and efficiently during saltatory conduction (Ritchie, 1995; Rushton, 1951; Tasaki and Matsumoto, 2002). The voltage and current flows that occur during the nerve action potential are based on the core-conductor or the cable theory (Aidley, 2001) implying that the axon behaves like a poorly insulated cable. The action potential is associated with local circuit currents that arise during the explosive inward flux of Na+ ions lasting few tenths of a millisecond and drives ions forward along the interior of the axon depolarizing the next segment of the membrane causing an outward capacitative current. The forward extent of this local current is determined by the space constant and its rise time by the time constant of the axon, both parameters of the resistance and capacitance of the axon and its membrane (Hodgkin, 1938; Hodgkin and Rushton, 1946). The presence of this passive local outward current in front of the action potential can be observed when conduction is blocked (Hodgkin, 1937a, 1937b), and its role was further demonstrated by the fact that the conduction velocity can be drastically slowed by increasing the external resistance around the nerve fiber, through which current must flow according to the local circuit theory

20 ms

(Hodgkin, 1964). In myelinated nerve fibers, the forward local outward current is sufficient to depolarize the successive nodes of Ranvier (Huxley and Stämpfli, 1949). When the node is depolarized to threshold the current turns inward when the Na+-ion channels open, while the segment of the axonal membrane behind the active part of the membrane demonstrates a slower and smaller outward current during repolarization. Hence, the membrane during the action potential constitutes a series of current sinks (where the current enters the axon) and current sources (where the current leaves the axon) that form the basis for the extracellular voltage changes. This voltage drop is an approximation of the timedependent membrane depolarization associated with the action potential, but “nerve in situ does not have an action potential of constant shape, for the magnitude and the shape of the recorded deflections depend upon the distance of the [micro]electrode from the nerve” (Figs. 2.8 and 2.9) (p. 403, Lorente de Nó, 1947). The shape and amplitude of the action potential recorded from nerve or muscle are determined by the placement of the generator in interstitial tissue that constitutes a volume conductor. This constraint on the understanding of the derivation of the cellular and compound action potential from excitable tissue has been dealt with in various publications

PHYSIOLOGY AND FUNCTION

0.5 mv

A2 15 0 mm

S

3 mm

10 mm 1

2 3 4 5

1 ms

Fig. 2.8 Compound nerve action potentials recorded from myelinated bullfrog nerve placed in two-dimensional medium. The recording electrodes were placed at 0, 3, and 10 mm from the nerve (Modified from Lorente de Nó 1947).

(Buchthal and Rosenfalck, 1966; Woodbury, 1966; Andreassen and Rosenfalck, 1981; Kimura, 1989; Dumitru and DeLisa, 1991). Thus, the intracellular spike in a volume conductor is recorded as an approximate measure of the action current associated with the local membrane current sources and sinks, and highly dependent on the distance from the nerve at which electrodes are placed (Lorente de Nó, 1947; Buchthal and Rosenfalck, 1966). Since these currents occur in connection with charges on the axonal membrane, this can be viewed as a layer of charges of opposite sign (dipoles) and since the nerve fiber is small in comparison with the volume conductor in which it is placed, the change in distribution of dipoles during the action potential can be simplified to equivalent dipoles (Lorente de Nó, 1947). To explain the action potential, a double dipole was postulated, one with a positive pole in the direction of propagation followed by one with inverted polarity (Soto and Vega, 1990). Recording of the field potential set up by dipoles is described by solid angle measurements, and in the resting cell, the charges of the membrane are opposite and equal with no net

33

electrical field, and the solid angle therefore zero (Woodbury, 1966) consistent with the fact that no current is flowing. During an action potential, unequal dipoles associated with depolarization are associated with a current flow that is proportional to the size of the solid angle and diminishes with the square of the distance to the recording point. The action potential propagated toward the recording electrode, therefore, presents a positive charge associated with the local current source (Fig. 2.10), a negative dipole associated with the sink of the inward current of the active region, and finally the positive dipole associated with the trailing repolarization and current source of the leaving action potential. The nerve action potential recorded by a monopolar electrode, is therefore, triphasic with a steep positive phase associated with the passive outward current, followed by a negative phase due to the active depolarization and finally a slower and smaller positive phase due to a slower repolarization. During bipolar recording, the shape is determined by the interaction of the triphasic potentials of opposite polarity at the two electrodes. The CMAP recorded from the end-plate region has a steep negative onset since the end-plate provides a current sink and there is no advancing current source. Recorded outside the end-plate the CMAP has a positive onset due to the strong negative potential at the end-plate seen as a positive potential at the recording site. Hence, the latency is measured to the takeoff of the response from base line (Fig. 2.11). 2.3.3. Motor nerve conduction studies Motor nerve fibers are studied by applying electrical stimuli to the nerve trunk and recording the evoked responses from one or more muscles supplied by the nerve. The particular nerves of the upper or lower extremities or cranial nerves that are studied depend on the clinical context (for details, see Chapters 23–26). In mononeuropathies or asymmetrical polyneuropathies, the clinically affected nerve(s) are targeted in the study, whereas in symmetrical polyneuropathies, the strategy is aimed at demonstration of widely distributed abnormalities. 2.3.3.1. Recording of the compound muscle action potential (CMAP) The smallest response that can be obtained by electrical or voluntary activation is the compound response from the muscle fibers, which are innervated by a single α-motor neuron, the motor unit (MU). At

34

CHRISTIAN KRARUP

B

A

C 100 μv

Electrode

Surface at wrist

50 Measured

of skin

2

30

4 20 6

1mm

Calculated (II)

8 Median nerve

10

10 50μv

12

2

4

6

D Amplitude (100 = electrode at the surface of nerve)

14 I II III IV V mm from 16 center of nerve 18 20 mm below skin

0

22 24 26

8 10 12 14 16 18 20 22 24 26 mm below skin I

100 70

II

50

III 30 IV V

20

10 7

0 1 2 3 4 5 msec

0

2

4

6

8 10 12 14 16 18 20 22 24 26 mm below skin

Fig. 2.9 Recording of the orthodromic compound sensory action potential from the median nerve at the wrist to show the influence of distance from the nerve on the amplitude and shape of the response. The curves in C and D were calculated from data of Lorente de Nó (From Buchthal and Rosenfalck, 1966).

increasing stimulus strength, the recorded response is the sum of the recruited motor unit potentials (MUP), the compound muscle action potential (CMAP) (Fig. 2.9). The CMAP amplitude is dependent on the Fig. 2.10 Orthodromic compound sensory (A) and muscle (B) action potentials recorded from the median nerve and the abductor pollicis brevis muscle, respectively.

summation and cancellation of phases of the individual MUPs, and MUs placed deeper in large muscles contribute less to the CMAP than superficial MUs (Parry et al., 1977). The MU force measured at the muscle

Median motor and sensory nerve conduction study

A

Digit I - wrist 37.2 μV, 3.0 ms, 43 m/s

CSAP

− 10 μV +

Wrist - APB

B

9.8 mV, 3.1 msec 3 mV

CMAP

0

20 ms

PHYSIOLOGY AND FUNCTION

35

CMAP from the APB at the end-plate and outside the end-plate Distance: 72 mm Wrist-APB (end-plate)

– 3 mV + 3.1 ms

Fig. 2.11 Compound muscle action potentials recorded over (top) and outside (below) the end-plate zone. The latency of the CMAP to the first deviation from the base line was the same at the two recording sites independent of distance.

Distance: 96 mm Wrist-APB (outside-end-plate)

3 mV

0

20 ms

tendon may be less subject to the location of the MU within the muscle, and recording of muscle force as well as the CMAP may provide additional information (Kerns et al., 1994). The muscle action potential is a prerequisite for the development of force associated with the excitation-contraction coupling (Fig. 2.12) (Krarup and Horowitz, 1979; Krarup, 1981). In order to study this latter part of muscle physiology, tension, shortening, or work may provide important information regarding contractile properties and subject to fatigue in addition to the size of the MU (Chan et al., 1998; Brown et al., 2000; Chan et al., 2001). The latency of the CMAP is measured to the take off of the signal from baseline and is the result of conduction along the fastest fibers in the nerve. Due to the delay of conduction along terminal fibers and at the neuromuscular transmission, the latency from the most distal site of stimulation to the muscle is not translated

CMAP brachial biceps

10 mV

200 ms

1 kg Twitch tension elbow flexors

Fig. 2.12 Compound muscle action potential from the brachial biceps muscle (top) and the isometric twitch tension from the elbow flexors (bottom) evoked by single supramaximal stimulation of the musculocutaneous nerve.

into the corresponding conduction velocity but is designated the “distal motor latency” (DML, ms). The motor conduction velocity (MCV, m/s) is calculated between stimulation sites along the nerve. In certain pathological states the relationship between the conduction velocities along distal and more proximal parts of the motor nerve fibers are relevant since the abnormalities may be unevenly expressed, and the ratio between the DML and MCV is calculated as the “terminal latency index” (TLI): distal distance/DML/MCV. In demyelinating polyneuropathy associated with anti-MAG gammopathy, the TLI is characteristically decreased (Kaku et al., 1994; Attarian et al., 2001) in accordance with the predominantly distal affection (Lunn et al., 2002). 2.3.4. Sensory nerve conduction studies Differential investigation of sensory nerve fibers is obtained by the study of purely sensory nerves such as the sural or the saphenous nerves or sensory branches of mixed sensory-motor nerves (for details, see Chapter 7). Electrophysiological studies of sensory fibers differ from motor fibers in that the compound sensory action potential (SNAP) is recorded directly from the nerve. The small signal is more difficult to distinguish from noise and, therefore, the SNAP often requires digital summation or averaging to ensure reproducibility in particular in diseased nerve, where the SNAP is reduced in size due to loss of fibers or poor summation due to dispersion. A major difference between the diagnostic yield of sensory and motor responses is related to the small amplitude of the SNAP (Dawson, 1956; Gilliatt et al., 1965; Buchthal and Rosenfalck, 1966). Thus, though modern equipment has improved signal-tonoise ratio, there is less confidence in the pathological

36

significance ascribed to the absence of a sensory than a motor response, in particular in the elderly. 2.3.4.1. Recording of the compound sensory action potential (SNAP) The SNAP may be recorded through surface or nearnerve electrodes. Surface electrodes have a diameter of 0.5–1 cm and are placed longitudinally on the skin above the nerve, for example the median nerve at the wrist or as ring electrodes at the digits, and have amplitudes of 5–50 μV. These electrodes allow resolutions of potentials of about 1 μV from noise, and the amplitude of the responses is dependent on the distance between the nerve and the electrode (Lim et al., 1995). In contrast, near-nerve electrodes require placement of an insulated needle with a 3-mm bared tip close to the nerve. Placement is optimized by stimulating through the needle while adjusting the threshold of an evoked motor (in mixed nerve) or sensory response (in sensory nerves) to <1 mA. At this low stimulation strength, the needle is placed <1 mm from the responding nerve fibers (Fig. 2.9) (Buchthal and Rosenfalck, 1966). The sensitivity is high, allowing recording of responses at ≤0.1 μV (Rosenfalck, 1978), and the rise-time of the SNAP is steep and may reveal individual components when the response is dispersed due to long conduction distance or to pathological variability in conduction velocities. Single stimulus recordings or averaging of few SNAPs are sufficient in normal nerve, whereas averaging of up to 1000 responses may be necessary to detect small responses of 0.1–0.2 μV. The SNAP may be recorded using uni- or bipolar electrode montages. The latter arrangement is often preferred as the stimulus artifact is smaller; however, the amplitude of the SNAP is influenced by the interelectrode distance due to phase cancellation (Gilliatt et al., 1965; Varghese and Rogoff, 1983; Andersen, 1985; Eduardo and Burke, 1988). A distance of 3–4 cm between surface recording electrodes is associated with the smallest distortion of latency and amplitude (Eduardo and Burke, 1988). Unipolar recording is obtained at near-nerve recording by placing the remote electrode at a transverse distance of 3–4 cm from the source, where the signal amplitude is less than 10% of that recorded close to the source (Buchthal and Rosenfalck, 1966). The SNAP may be recorded either antidromically or orthodromically, and both have their own advantages and disadvantages. Antidromic recordings of the SNAP evoked at several sites along the arm may be obtained with surface electrodes (Kimura et al.,

CHRISTIAN KRARUP

1988), but this is only possible to a limited extent in the leg. It is often impossible to record the orthodromic SNAP with surface electrodes at proximal sites (Olney, 1999), whereas this can be carried out in normal as well as pathological nerves in both the arms and the legs using near-nerve needle electrodes (Buchthal and Rosenfalck, 1966; Behse and Buchthal, 1971; Krarup et al., 1990a; Liguori et al., 1992; Krarup and Trojaborg, 1996). The SNAP is triphasic in shape with an initial positive phase (Fig. 2.13). The parameters of the SNAP that provide meaningful information in clinical evaluation include the amplitude and latency: The amplitude of the SNAP is measured peak-to-peak and is a compound of triphasic single fiber potentials (SFAP) each having an amplitude of ≤0.1 μV and a total duration of 1–2 ms when recorded with a needle electrode. Thus, the summation is less complete compared with the MUPs and the response more prone to cancellation of opposite phases. The fibers that contribute to the SNAP range above 7–8 μm and the peak-to-peak amplitude is determined mainly by fibers greater than 9 μm. The amplitudes of SFAPs from fibers of different diameters indicate that small fibers in the nerve have little influence on the SNAP. The amplitude of the extracellular SFAP is dependent on the area of the axon, and assuming a square relationship between the amplitude of the SFAP and fiber diameter (Rushton, 1951; Marks and Loeb, 1976; Loeb et al., 1977; Rindos et al., 1984), it may be estimated that the SFAP of a small myelinated fiber of 4 μm has an amplitude of only 10–15% of that of a fiber of 11 μm, and that it requires 8–10 times as many fibers to generate a potential that can be distinguished from noise (Krarup, 2004). Assuming the SFAP to have a triphasic shape with a duration of 2 ms, an amplitude of 0.105 μV for a fiber of 11 μm (with a similar total area of the positive and the negative phases) (Fig. 2.13), an exponential increase in duration with decreasing fiber diameter, and a conversion factor of 4.5 m/s/μm, (Buchthal and Rosenfalck, 1966; Paintal, 1966; Behse and Buchthal, 1978; Kovacs et al., 1979; Contento et al., 1983), we reconstructed the sural nerve SNAP (Fig. 2.13) based on the average morphometrical data from the sural nerve (Fig. 2.2) (Behse, 1990). Simulations (Fig. 2.14) indicated that the loss of small fibers of less than 6–7 μm has only negligible effect on the SNAP amplitude. The small contribution of small fibers in spite of their large number illustrates the effect of phase cancellation, which may have other paradoxical effects on the reconstructed

PHYSIOLOGY AND FUNCTION

37

A

B

Mean fiber diameter distribution from normal

Single fiber potential (SFAP)

sural nerve (n = 10) (Behse, 1990)

0.08 mean number of fibers = 6940 Amplitude (μV)

Number of fibers

0.10 2000 1800 1600 1400 1200 1000 800 600 400 200 0

(range 5075 - 9460)

0.06 0.04 0.02 0.00 −0.02 −0.04 −0.06 0.0

1

2

3

4

5

6

7

8

9

10

11

12

0.5

1.0

1.5

2.0

2.5

13 Duration (ms)

Fiber diameter (μm)

C

D Simulated sural SNAP at midcalf

Left sural nerve recording 20 LM-MC

Peak-to-peak = 30

μV – 10 μV +

Amplitude (μV)

15 10

0 −5

2.7 ms = (49 m/s)

μV

Peak-to-peak = 26

5

2.4 ms = (54 m/s)

−10 0 0

10 ms

1

2

3

4 5 6 7 SNAP latency (ms)

8

9

10

Fig. 2.13 (A) Fiber diameter histogram as in Fig. 2. (B) model SFAP from myelinated fiber of 11 mm. The total duration of the SFAP was 2 ms, with a faster rate of rise than of decline of the triphasic potential. The area of the depolarizing phase of the SFAP (positive values) was the same as the sum of the areas of the hyperpolarizing phases (negative values). (C) SNAP recorded from the sural nerve of a normal subject. (D) reconstructed SNAP conducted over the same distance as the recorded potential (From Krarup, 2004).

SNAP amplitude. Thus, loss of the largest fibers in the nerve was associated with an increase in amplitude, and even though such selective loss probably is extremely rare, it suggests that reduction in conduction velocity with slight or absent reduction in amplitude may occur through loss of very large fibers. In contrast to the CMAP, the effect of fiber loss on the SNAP is strongly influenced by the diameters of the conducting fibers. The latency of the SNAP is recorded to the first positive phase (in triphasic responses) or to the start of the negative phase (in biphasic responses that occur in bipolar recording montages due to cancellation of the first positive phase). This latency reflects conduction in large fibers of the nerve that summate to generate the first positive phase (Fig. 2.15) (Buchthal et al., 1984). In this respect the latency of the SNAP differs

from that of the CMAP as the latter is generated by the fastest conducting fiber(s) of the nerve. 2.3.5. Conduction in normal nerve 2.3.5.1. Motor and sensory conduction velocities The conduction velocity along normal nerve is a parameter, which reflects the diameter, the myelin thickness in relation to the diameter (g-ratio), the axoplasmic conductance, myelin capacitance, the area of the node of Ranvier, and the Na+ ion-channel conductance (Rushton, 1951; Moore et al., 1978; Ritchie, 1995). In cat the conversion factor of 6 m/s/μm in large fibers adequately describes the ratio between conduction velocity and the fiber diameter (Hursh, 1939; Berry et al., 1944; Boyd and Kalu, 1979), whereas in humans the conversion factor has been found to be

38

CHRISTIAN KRARUP

A

Number of fibers

Loss large fibers

+ 62

21 m/sec

33

− 1

2

3

4

5

6

7

8

9

0.5 μv

Popl. fossa

10 11 12 13

Loss small fibers

B

10 μv

Sura

2000 1800 1600 1400 1200 1000 800 600 400 200 0

+

Fibers 600

Amplitude fiber diameter distribution

Amplitude (μV)

45 40

Loss large fibers (μV)

35

Loss small fibers (μV)

400

30

200

25 20

n = 2600

15

n = 4000

10

0

5

14

0 0

C

2

4

6

8

10

12

14

16

Latency (ms)

Loss large fibers (ms) Loss small fibers (ms)

12

10

8

6

4

2

0

Diameter, μm

Fig. 2.15 Correlation between fiber distribution and the latency and conduction velocity of the compound sensory response. A conversion factor of 4.5 m/s/μm corresponds to the maximal conduction velocity of the largest fibers in the nerve. Modified from Krarup and Buchthal (1985).

Latency fiber diameter distribution

14

12

10 8 6 4 2 0 0

2

4

6

8

10

12

14

Fiber diameter (μm)

Fig. 2.14 (A) Fiber diameter distribution as shown in Fig. 2, with arrows indicating the direction of fiber loss in the model: Small fibers were gradually lost in one bin at a time (loss small fibers), or large fibers were gradually lost (loss large fibers). (B) shows the amplitude after loss of a particular bin of small or large fibers. (C) the effect of fiber loss on the latency of the SNAP in the model (From Krarup, 2004).

close to 4.5 m/s/μm (Behse and Buchthal, 1978). In normal fibers the g-ratio is 0.5–0.7 among large fibers, which is associated with the longest space constant λ and therefore “will cause the action current at one node to stimulate the next most strongly” (Rushton, 1951), whereas it deviates from this optimal value in smaller fibers suggesting that the conversion factor may also

change in different fibers (Boyd and Davey, 1968; Boyd and Kalu, 1979). The internodal length has only little effect on the conduction velocity, and this is in accordance with the finding that the conduction velocity in regenerated fibers is directly related to the fiber diameter (Berry et al., 1944; Cragg and Thomas, 1964; Fugleholm et al., 2000), whereas the internodal length remains shortened at 300–400 μm (Hiscoe, 1947; Vizoso and Young, 1948), in spite of persistent changes in the membrane potential of regenerated fibers (Moldovan and Krarup, 2004a, b). These considerations are of importance since a number of factors in pathological situations may cause a reduction in conduction velocity. Such factors include loss of large diameter fibers and abnormalities in myelin, but in addition less commonly considered changes such as atrophy of axons may cause considerable changes in conduction velocity (Baba et al., 1982; Gillespie and Stein, 1983). Moreover, the conduction velocity in clinical practice is an expression of the

PHYSIOLOGY AND FUNCTION

function of only the fastest conducting fibers, and if just one large α-motor fiber is left in the nerve the MCV may remain normal (Fullerton, 1969). Considerable effort has therefore been invested in assessing the conduction velocities also in slower conducting fibers (for details see Chapter 18). The distribution of conduction velocities (DCV) of individual fibers has been assessed (Cummins and Dorfman, 1981; Dorfman et al., 1982; Dorfman, 1984) using different methods such as the collision technique (for details see Chapter 16), where two stimuli are spaced with intervals to allow escape of fibers conducting with different rates from occlusion (Gilliatt et al., 1976; Nix et al., 1989) or calculations based on the change in shapes and amplitudes of the compound nerve or muscle response recorded at different conduction distances (Cummins et al., 1979a). The DCV of motor fibers ranged about 15 m/s between the fastest (60 m/s) and the slowest (45 m/s) conducting α-motor fibers in the median and ulnar nerves (Lee et al., 1975; Dorfman, 1984), whereas the DCV of the nerve action potential has a wider range due to the contribution of thinner fibers. The DCV indicates that the MCV or SCV may decrease resulting from fiber loss alone. Estimation of the DCV presupposes that the shapes and amplitudes of the SFAPs and the MUPs are known. However, the parameters depend on several factors such as the recording methods (Cummins et al., 1979b; Kovacs et al., 1979; Contento et al., 1983). Fast and slowly conducting potentials have been included in the models used in normal as well as diseased nerves by Stegeman and coworkers (Stegeman et al., 1979; Schoonhoven et al., 1987; Schoonhoven and Stegeman, 1991). In the normal sural nerve the minimum conduction velocity recorded with nerve electrodes is in the range of 10–20 m/s, corresponding to fibers diameters of 3–5 μm (Rosenfalck, 1978; Buchthal et al., 1984; Krarup and Buchthal, 1985). The amplitudes of these components are about 0.2–0.3 μV and averaging of 500–1000 traces is required for distinction from noise. In diseased nerve the minimum conduction velocity may be reduced as the only abnormality (Shefner et al., 1991a, 1991b, 1992). This may be due to conduction along regenerating fibers (Krarup et al., 1990b) or to effects of demyelination (Krarup and Trojaborg, 1996). 2.3.5.2. Effect of conduction distance on the CMAP and the SNAP The differences in duration, shape and amplitudes of SFAPs compared with MUPs and the broader distribution of conduction velocities in sensory than motor

39

nerve fibers have a crucial influence on the effects of temporal dispersion on the SNAP and the CMAP (Kimura et al., 1986, 1988; Krarup and Trojaborg, 1996; Liguori et al., 1992). The duration of the negative spike of the SNAP is 1–2 ms as compared to 5–10 ms of the negative phase of the CMAP. The SNAP amplitude is strongly dependent on conduction distance, which has much less influence on the CMAP amplitude (Kimura et al., 1986; Olney et al., 1987; Krarup and Trojaborg, 1996; Krarup, 2002; Van Aken and Van Dijk, 2003). Thus, the reduction in amplitudes of the median and ulnar SNAPs is 5–10 times larger than the corresponding CMAPs (Fig. 2.16A), and their increase in duration is correspondingly more pronounced (Fig. 2.16B). The greater effect of temporal dispersion on the SNAP than the CMAP (Fig. 2.16D) is due to the short duration of the SFAP compared with the MUP (Kimura et al., 1988) and the triphasic shape of the SFAP presumably has a strong phase canceling effect (Kincaid et al., 1988). Simulation of the CMAP in animal studies has also shown that the distal/proximal amplitude ratio in normal nerve is strongly dependent on the model parameters (Rhee et al., 1990). These differences in duration and amplitude of the SNAP and CMAP have a profound influence on the detection of conduction block due to focal demyelination of sensory and motor fibers. Whereas conduction block of motor fibers (MCB) can be demonstrated at distal-to-proximal amplitude reduction of more than 50% (Fig. 2.17A), the amplitude reduction of the SNAP over comparable distances due to normal temporal dispersion is up to ten times larger than that of the CMAP (Krarup, 2002). Thus sensory conduction block may be difficult to detect (Fig. 2.17B). The interaction between SFAPs and MUPs that determine the effect of temporal dispersion is dependent on both the synchronization and the number of single unit responses, which contribute to the compound responses. Thus the reduction of the proximal compared with the distal CMAP required to demonstrate MCB is difficult to state explicitly in cases of pronounced axonal loss. The SNAP tends to become polyphasic at axonal loss (see for example Stegeman for discussion (Stegeman et al., 1979)), and the temporal dispersion caused by conduction distance has little or no effect on the peakto-peak amplitude due to poor synchronization. In 1991 a consensus report agreed that MCB should be confirmed at short conduction distances, that it is unreliable when the CMAP is markedly depressed, and that it should only be accepted in connection with

40

CHRISTIAN KRARUP

A

C SNAP and CMAP amplitudes as function of

Sensory and motor response durations

conduction distance 350 120

300 Duration (%)

Amplitude (%)

100 80 60 40 20 0

250 200 150 100 50

0

100

200

300

400

500

600

700

0

Conduction distance (mm)

B

100

D

300

400

500

600

700

Amplitude area relationship

Sensory and motor latencies 140

10

63 m/s

120

8 6

Amplitude (%)

Latency (ms)

200

Conduction distance (mm)

69 m/s

4 2

100 80 60 40 20

0

y=341.37e −0.0138x R2=06341

0 0

100

200

300

400

500

600

50

700

Conduction distance (mm)

CMAP SNAP

100

150

200

250

300

350

Duration (%)

Fig. 2.16 The influence of conduction distance on the amplitudes and durations of compound motor (CMAP) and sensory (SNAP) action potentials from the median and ulnar nerves of normal subjects. The distances were measured from the wrist (distance = 0). The CMAP (filled circles, full lines) was recorded through surface electrodes over the abductor pollicis brevis or the abductor digiti minimi muscles, and the SNAP (open circles, dashed lines) through near-nerve needle electrodes at wrist, elbow, axilla, and Erb’s point. (A) the percentage amplitudes of the CMAPs and SNAPs were fitted with exponential functions and the 95% lower limits (short dashed lines) indicate the lower limits of normal reduction, which were markedly different for motor and sensory responses. (B) sensory conduction velocities were about 10% greater than motor conduction velocities. (C) the durations of the CSAPs increased markedly with conduction distance as compared with CMAPs. (D) the amplitude reductions were related to increases in duration. From Krarup (2002).

weakness (Cornblath et al., 1991). The American Association of Electrodiagnostic Medicine recommended in 1999 (Olney, 1999) a limit of 50% reduction in amplitude in the median or ulnar nerves and that short segment stimulation should be carried out. Comparing different criteria of MCB Pfeiffer et al. (Pfeiffer et al., 2000) found reductions of the CMAPs in amyotrophic lateral sclerosis and Charcot-MarieTooth type 1, which did allow distinction of these disorders from acquired demyelination. These findings emphasize that marked fiber loss and reduced excitability (Meulstee et al., 1997; Cappelen-Smith et al., 2001) interfere with investigation of MCB. The adoption of conservative criteria to identify MCB is a methodological compromise, since it is likely that mild MCB occurs in clinical situations. Thus, single fiber EMG electrodes have been used to detect drop-

out of components of the CMAP (Padua et al., 2001); however, phase cancellation of individual short spikes using these electrodes, as well as concentric needle electrodes, reduces their reliability to definitely identify MCB. Comparison of the maximal motor and sensory conduction velocities over the same nerve segment shows in most instances that the SCV is 5–10% greater than the MCV (Fig. 2.16C) (Dawson, 1956; Buchthal and Rosenfalck, 1966; Behse and Buchthal, 1971; Macefield et al., 1989; Krarup and Trojaborg, 1996; Krarup, 1999), whereas the threshold of sensory and motor fibers do not differ in all studies (Dawson, 1956; Buchthal and Rosenfalck, 1966). The greater SCV than MCV is in accordance with the larger diameters of type I afferent fibers in mixed nerve and Aα fibers in cutaneous nerves (Macefield et al., 1989) than the

PHYSIOLOGY AND FUNCTION

41

A

B

Left ulnar−motor

–

−

3

Left ulnar−sensory

2 mV +

1 μV

3

0.5 mV, 41 m/s

+

0.4 μV, 33 m/s

2 0.7 mV, 38 m/s

1 μV

2 0.2 μV, 44 m/s

1 2 μV

1 4.6 mV, 53 m/s

4.1 μV, 53 m/s

Wrist-APB

3 μV

Dig5-wrist 5.4 mV, 2.5 ms 20 ms

20 ms

CMAP amplitude

120 100 80 60 40 20 0 0

CSAP amplitude

50

100

150

120 100 80 60 40 20 0 0

200

CMAP latency

5 4 3 2 1 0 0

50

100

50

100

150

200

CSAP latency (ms)

(ms)

0

(% )

(% )

0

9.6 μV, 47 m/s

150

5 4 3 2 1 0

200

0

50

100

150

200

Conduction distance (mm)

Fig. 2.17 Motor and sensory conduction block in chronic inflammatory demyelinating polyneuropathy (CIDP): 28-year-old woman with progressive weakness and numbness of the hands for 4 years. There was marked weakness of median- and ulnarinnervated muscles and reduced touch and vibratory sensation in all fingers. Tendon reflexes were weak or absent. Spinal protein normal. Conduction studies of motor fibers (A) of the ulnar nerve showed conduction block localized to a 5–7 cm segment between elbow and wrist. The sensory responses decreased similarly across the same nerve segment (B) indicating sensory conduction block. The CMAP and the CSAP amplitude graphs (below) showed the normal average and lower 95% confidence limit of amplitude reduction of sensory (CSAP) and motor (CMAP) responses; the patient data were seen to deviate markedly from the normal limit. The CMAP and CSAP latency graphs (bottom) showed that the patient responses had longer latencies than control indicating reduced conduction velocities. From Krarup (2004).

diameters of the largest α-motor fibers (McLeod and Wray, 1967). The conduction velocities at different proximo-distal segments of the limbs are often considered to decrease according to tapering of nerve fibers; however, the evidence for such tapering is not convincing. Thus fiber diameters remain constant between branching points and have smaller diameters distal to branching, which is most pronounced distally close to innervation of target organs (Quilliam, 1956; Berthold and Rydmark, 1995), and comparisons of fiber diameter spectrums in normal human median and ulnar nerves show constant diameters between axilla and

wrist, whereas the diameters were smaller and internodes shorter distal to the wrist and at digital nerves (Buchthal and Rosenfalck, 1966; Caruso et al., 1992). Accordingly the conduction velocities were lower distal to than proximal to the wrist. In cat single fiber potentials had constant conduction velocities along the length of the hindlimb without nerve fiber branching (Rindos et al., 1984). Notwithstanding a constant fiber diameter distribution, the SCV is about 10% higher along the proximal than the distal arm. This discrepancy between anatomical and physiological findings can be explained by the effect of summation of positive

42

phases from large fibers that determine the latency of the SNAP. Thus, at short conduction distances the latency to the first positive peak is determined by fiber groups with smaller diameters (Horowitz and Krarup, 1992). At longer conduction distances the latency is determined by larger fibers due to temporal dispersion, and hence there is an apparent increase in conduction velocity (Krarup, 2004). 2.4. Motor unit number estimation (MUNE) The amplitude of the CMAP and SNAP reflects the number of large axons in the nerve. However, the CMAP in chronic axonal lesions is a relatively poor indicator of the number of α-motor axons due to the compensatory collateral sprouting of surviving motor fibers. The CMAP may therefore remain normal for extended periods in chronic progressive lesions until the capacity for sprouting is outweighed by the degenerative process (Brown and Feasby, 1974). This capacity for collateral sprouting does not exist in sensory axons and, therefore in chronic lesions the amplitude of the SNAP is a more sensitive indicator of axonal loss than the CMAP (Krarup, 1999). To obtain better insight into the presence of motor axon loss specialized studies have been developed to provide an estimate of the number of MU (McComas et al., 1971b), and such methods have demonstrated a correlation between electrophysiological findings and clinical deficits for example in amyotrophic lateral sclerosis (LomenHoerth and Olney, 2000; Arasaki et al., 2002). A number of methods (for details see Chapter 12) have been developed to provide an estimate of the number of motor units that are innervated by a functional α-motor axons (McComas et al., 1971a; Sica et al., 1974; McComas, 1995). They all have as their main goal the detection and delineation of an “average” MUP in the muscle and are based on the theory that the CMAP is an algebraic sum of the total number of this average MUP, i.e. MUNE can be obtained as a ratio of the CMAP/“average” MUP. To ascertain the amplitudes of MUPs, most studies have used threshold stimulation to activate individual α-motor fibers as indicated by discrete increments of the motor response when additional MUPs are recruited. The limitations and sources of error associated with the techniques have been discussed in several reviews (Shefner, 2001), and include “alternation” (Aoyagi et al., 2000), bias in the activation of fibers of particular diameters (Fang et al., 1997). To avoid some of these errors, microstimulation has been used to activate motor fibers (Arasaki et al., 2002). In

CHRISTIAN KRARUP

addition, MUNE has been carried out using MUPs recorded during voluntary activity as templates since smaller calibre α-motor fibers are recruited at weak effort (Shefner, 2001). To avoid the bias associated with activation of particular fibre groups, Daube and colleagues have developed the use of statistical methods to carry out MUNE at higher levels of stimulation (Daube, 1995; Henderson et al., 2003), and the use of this method has been further tested in different conditions (Daube, 1995; Olney et al., 2000; Henderson et al., 2003; Lomen-Hoerth and Slawnych, 2003). MUNE, while showing considerable variability (Simmons et al., 2001), has been used to follow the progressive loss of MUs in a number of disorders such as spinal muscular atrophy, amyotrophic lateral sclerosis, Charcot-MarieTooth disease, poliomyelitis, carpal tunnel syndrome and polyneuropathy (Ballantyne and Hansen, 1982; McComas et al., 1997; Cuturic and Palliyath, 2000; Aggarwal and Nicholson, 2002; Lewis et al., 2003). In regenerated nerve, MUNE has been found to provide better insight into the degree of motor reinnervation than the amplitude of the CMAP (Madison et al., 1999; Krarup et al., 2002). In partial conduction block, MUNE might be expected to provide better measure of the number of fibers blocked than the decline in amplitude of the CMAP; however, due to the preferential involvement of large fibers in compression the results were not useful (Jillapalli et al., 2003). It might be of interest to test the use of MUNE in demyelinating neuropathy associated with MCB. 2.5. Studies of nerve roots Studies of proximal nerve segments and spinal roots are needed both when the suspected pathological process is localized to these parts of the PNS and when involvement of the CNS is a differential diagnostic consideration. These areas of the PNS are assessed by the study of F-waves (Magladery and Mcdougal, 1950) or H-reflexes or from motor or somatosensory evoked potentials. 2.5.1. F-waves and reflex studies F-waves (for details see Chapter 9) are evoked by maximal stimulation of motor fibers and represent backfiring from the motor neuron. They are of particular use in situations, in which the distal motor nerve conduction studies show little evidence of fiber loss or slowed conduction (Kimura, 1997), and is of high sensitivity in lesions with only mild affection due to the inclusion of

PHYSIOLOGY AND FUNCTION

long nerve segments (Kohara et al., 2000). Thus, the absence of F-waves in the face of normal motor conduction studies or in patients with multifocal lesions (Chaudhry et al., 1994) provides evidence of proximal involvement. Particular attention is paid to the inherent variability of the F-wave latency and amplitude and should be distinguished from the late responses with constant discharge and latencies, the so-called A-waves that are evidence of axonal reflexes indicating branching probably on the basis of collateral sprouting or indicate ephaptic transmission between demyelinated nerve fibers (Fullerton and Gilliatt, 1965; Bischoff et al., 1996; Roth and Magistris, 1999). Late responses have not been found of particular help to diagnose patients with monoradiculopathy (Wilbourn and Aminoff, 1998). H-reflexes are monosynaptic reflexes evoked by stimulating the sensory fibers of a mixed nerve with a submaximal response that does not excite motor fibers (Veale et al., 1973). In newborns, H-reflexes can be recorded from a number of muscles but in adults they are present regularly only in the triceps surae muscle group (Fisher, 2002), and abnormalities may be present early in patients with radiculitis in Guillain–Barré syndrome (Gordon and Wilbourn, 2001). It should, however, be realized that central abnormalities in the reflex arch may interfere with the ability to elicit the H-reflex. The evaluation of conduction in cranial nerves presents particular problems. Only the part of the facial nerve distal to stylomastoid foramen is accessible to direct stimulation and the latencies and amplitudes of the CMAP in various muscles can be measured in different disorders and lesions associated with facial palsy. To access intracranial portions of the facial nerve, the use of the blink reflex (for details see Chapter 10) provides the possibility of measuring conduction of the facial nerve in series with the trigeminal nerve (Aramideh and Ongerboer de Visser, 2002). The blink reflex consists of early ipsilateral components (the R1 response) that are recorded from the orbicularis oculi muscle and later bilateral components (the R2 responses) and the distribution of abnormalities may be of help in the localization of trigeminal or facial nerve abnormalities. In addition, the masseter reflex may be of use in the testing of sensory as well as the motor branch of the trigeminal nerve (Auger, 1998). 2.5.2. Motor and sensory evoked potentials Motor-evoked potentials (MEP, for details see Chapter 19) are primarily used to assess conduction along the corticospinal pathways, but special techniques are used

43

to investigate the excitability of cortical neurons. In clinical settings MEPs are evoked by magnetic stimulation of the brain. Stimulation of the spinal cord or roots is carried out using magnetic or electrical pulses to measure central conduction time (MCCT) and to assess peripheral nerve conduction. Peripheral versus central abnormalities may also be evaluated by measuring F-wave latencies rather than using root stimulation to asses the peripheral part of the conduction path. However, at root stimulation activation of nerve fibers occurs at the level of the spinal nerve outside the intervertebral foramen and the central conduction time obtained as the difference between the cortical and the peripheral latencies includes conduction along roots, and the MCCT may therefore indicate apparent central abnormalities in for example GBS with conduction delay due to demyelination of roots (Ravnborg and Dahl, 1991; Takada and Ravnborg, 2000). In certain disorders affecting the CNS as well as the PNS, such as amyotrophic lateral sclerosis or inflammatory neuropathies, MEP studies are of use to localize conduction delays to both systems (Uncini et al., 1988; Boulesteix et al., 1995; Inaba et al., 2001; Oshima et al., 2001). Conduction deficits along central pathways may now be quantified using the triple stimulation technique that serve to resynchronize the dispersed conduction along the corticospinal path (Magistris et al., 1998, 1999; Bühler et al., 2001) and hence quantify loss of central fibers (Rösler et al., 2000). The triple stimulation technique may also be of use to quantify central conduction failure during fatigue (Andersen et al., 2003). Somatosensory evoked potentials (SEP) represent conduction along distal and proximal parts of the PNS, spinal roots, and central pathways localized to the dorsal column fibers (Chiappa, 1997). In patients with suspected radiculitis or proximal nerve demyelination in inflammatory neuropathies, SEP studies alone, or combined with late response recordings or MEP, may be useful to localize proximal slowing or conduction block (Fig. 2.18) (Parry and Clarke, 1988; Gilmore and Nelson, 1989; Ropper et al., 1990; Vajsar et al., 1992; Brown et al., 1993; Wong, 1997). 2.6. Studies of autonomic and small unmyelinated fibers In PNS the autonomic fibers are small unmyelinated axons that cannot be studied using direct stimulation and recording of nerve action potentials. Using microneurography, three types of afferent C-fibers have

44

CHRISTIAN KRARUP

Fig. 2.18 Motor evoked potentials (MEP) and sensory evoked potentials (SEP) from 29-year-old woman with slowly progressive paraparesis and sensory disturbances in the legs without sphincter symptoms since 8 years of age. MEP obtained by transcranial (A) and lumbar magnetic stimulation (B) with recording from the right abductor hallucis muscle, showed normal distal latencies (B) and prolonged latencies at cortical stimulation (A). The central conduction time (CCT) was markedly prolonged. The SEP was obtained by stimulation of the right tibial nerve at ankle and recording of the compound nerve action potential at the popliteal fossa (A), the electrospinogram at the Th12 spine (B), and the cortical response at C3 (C), averaging of 1000 responses in two separate bins for reproducibility. The nerve conduction velocity and latency to the electrospinogram were normal; no cortical response could be recorded.

been identified in normal human skin, type 1 and 2 were nociceptive and cold afferent respectively, whereas the function of type 3 was not identified (Serra et al., 1999). Clinical studies rely on the assessment of the galvanic skin response associated with reflex activation of sympathetic fibers and the sweat glands that they innervate, the so-called sympathetic skin response (SSR), cardiovascular reflex activity, evaluation of sweating or the measurements of psychophysic responses to heat or cold (Fowler et al., 1987; Le Quesne et al., 1990; Aminoff et al., 1996; Dutsch et al., 2002; Lacomis, 2002). Laser-stimulation of skin has been developed to study pain and autonomic pathways (Bromm and Chen, 1995; Cervera et al., 2002; Arendt-Nielsen and Chen, 2003). Skin biopsies have been instrumental in diagnosing small fiber neuropathy in parallel with or when large fibers were not affected (Kennedy et al., 1994; Lauria et al., 1999; Griffin et al., 2001; Novak et al., 2001; Polydefkis et al., 2001, 2002).

2.7. Studies of muscle Electromyography (EMG) is an essential part of the investigation of abnormalities of the PNS as it provides evidence of denervation and reinnervation in acute and chronic partial and complete nerve lesions. The aim of the EMG examination is to determine whether weakness is due to a neurogenic disorder or to myopathy. Since EMG findings often are non-specific, it is necessary to study: (1) the activity at rest (denervation activity—fibrillation potentials, positive sharp waves; fasciculations; complex repetitive discharges; myotonic activity; neuromyotonia); (2) motor units potentials (MUPs) during weak effort; and (3) the electrical activity at maximum effort (Buchthal and Kamieniecka, 1982). In some instances it may be necessary to supplement the examination with specialized studies such as analysis of turns and amplitude ratios at higher levels of effort (Fuglsang-Frederiksen et al., 1976, 1977).

PHYSIOLOGY AND FUNCTION

Due to the large variability of motor unit potential (MUP) duration and amplitude, it is necessary for quantitative evaluation to measure 20–30 MUPs from about 10 different insertion sites in the muscle to obtain a representative sample (Fig. 2.19), and findings should be compared to age-matched controls for the examined muscle (Rosenfalck and Rosenfalck, 1975). The specific abnormalities in neurogenic weakness include signs of chronic partial denervation with prolongation of the duration and increase of the amplitude of the average MUP during weak effort and increased amplitude and loss of motor units during maximal effort (Simonetti et al., 1999). Denervation activity as an abnormality is an unspecific finding that is seen in both neurogenic lesions and myopathy. However, when present together with specific changes of MUPs, fibrillation potentials and positive sharp waves indicate the presence of ongoing denervation. 2.8. Effects of biological and physical variables on PNS function A number of factors should be taken into consideration when interpreting physiological findings in peripheral nerves. The most important are age and temperature. 2.8.1. Age Increasing age affects nerve conduction as well as EMG. In newborn, the conduction velocities of both motor and sensory fibers are low and gradually increase to normal range at the age of 3–4 years (Wagner and Buchthal, 1972; Rabben, 1995; Devries, 1996; Cai and Zhang, 1997), and may be used to assess the development of preterm infants (Pierrat et al., 1996). Developmental changes of myelinated fibers occur in parallel (Ouvrier et al., 1987). A large number of studies have confirmed that the amplitudes of the SNAP, and to a lesser extent, the CMAP decrease with age in the adult (Buchthal and Rosenfalck, 1966; Behse and Buchthal, 1971; Rosenfalck and Rosenfalck, 1975; Horowitz and Krarup, 1992; Trojaborg et al., 1992; Trojaborg, 1992), and in about 10% of elderly subjects without neuromuscular disease it is not possible to record a sural nerve action potential with surface electrodes (Rivner et al., 2001). Though the quantitative data on the fiber distribution and number in nerves at different ages is sparse the evidence indicates a general decline in the density of fibers of the sural nerve with age (Jacobs and Love,

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1985). These authors found an increase in degeneration and regeneration of fibers but it is not known whether there was a general loss of the total number fibers. Other biological variables that have been found to influence nerve conduction parameters include gender and height. The amplitudes of the sural nerve SNAP are greater in females than in males when recorded with a near-nerve needle, which may reflect differences in subcutaneous tissue composition (Horowitz and Krarup, 1992), whereas surface recordings of the orthodromic SNAP in median and ulnar nerves did not differ in men and women (Shehab et al., 2001). Height has been found to be negatively correlated with conduction velocities in motor as well as sensory nerves in large population based studies (Solders et al., 1993; Letz and Gerr, 1994; Rivner et al., 2001), whereas other studies with fewer subjects but better control of environmental factors such as temperature have not confirmed this relationship (Trojaborg et al., 1992). Body mass index has been found to be correlated with conduction velocities in some studies but the relationship is weak. 2.8.2. Temperature By far the most important environmental factor that influences the results of conduction studies is temperature. Low temperature causes depolarization of the membrane potential and prolong refractoriness (Burke et al., 1999; Kiernan et al., 2001a). The exact correction factor for the effect of temperature is unknown, so the convention of adding 2 m/sec/˚C in clinical studies is at best an approximation (Cummins and Dorfman, 1981; Stegeman and De Weerd, 1982; Trojaborg et al., 1992; Franssen and Wieneke, 1994). Low temperature has profound effects on the SNAP and single-fiber action potential rise times and duration (Paintal, 1965, 1966). The distance between electrodes may also be a determining factor in how the amplitude of the SNAP is influenced by temperature (Alfonsi et al., 1987; Letz and Gerr, 1994; Pitt, 1996; Ashworth et al., 1998; Rutkove, 2001), since cooling is associated with slowing of conduction and less interaction and cancellation of phases with different polarity. At bipolar recording the amplitude increases at low temperature due to the prolonged duration and reduced phase cancellation at low conduction velocity. At monopolar recording the temperature had little and variable effect on the SNAP amplitude (Buchthal and Rosenfalck, 1966; Stegeman and De Weerd, 1982). In pathological nerve, in particular when

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Fig. 2.19 EMG at weak effort with recording of motor unit potentials (MUAP) from the brachial biceps muscle in normal muscle (A), myopathy (B), and neurogenic disorder (C). In the left column are shown examples of MUAPs, in the middle column is shown two motor units (A), degeneration and atrophy of muscle fibers in myopathy, and degeneration of one motor neuron with collateral sprouting and enlargement of the remaining motor unit (C). In the right column is shown the distribution of the duration of simple (open columns) and polyphasic (filled columns) MUAPs, and the mean findings and deviation from normal are shown above each distribution (normal: ±20%). Modified from Krarup (2003).

neurogenic

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46 CHRISTIAN KRARUP

PHYSIOLOGY AND FUNCTION

subject to demyelination, low temperature may mask abnormalities since the prolongation of the SFAP may allow conduction across segments of nerve fibers with reduced safety factor (Bostock et al., 1978, 1981; Franssen et al., 1999; Kaji, 2003). 2.9. Pathophysiological considerations The main purpose of the electrodiagnostic evaluation of peripheral nerve function is to determine the degree and type of pathophysiological changes to infer pathological abnormalities and to arrive at a specific clinical diagnosis. Achievement of this goal requires that PNS functional changes are assessed and interpreted in the appropriate clinical context, and usually that ancillary laboratory studies are carried out. The main pathological changes of PNS fibers include axonal loss and demyelination, and the most important function of electrophysiological examination is to distinguish these basic changes. However, demyelination rarely occurs in isolation, as for example, in hypertrophic hereditary neuropathy, where the degree of axonal loss is as severe as in the neuronal form (Behse and Buchthal, 1977; Buchthal and Behse, 1977). The distinction of primary axonal loss from demyelination relies on slowing of motor and sensory nerve conduction disproportionate to that which may be explained by loss of nerve fibers (Behse and Buchthal, 1978; Buchthal et al., 1984). Loss of fast conducting fibers is associated with reduction in conduction velocity, and slowing disproportionate to this fiber loss indicates the presence of other mechanisms, which include abnormalities in myelin, atrophy of axons, or ion-channel dysfunction. Degrees of conduction slowing that are consistent with demyelination have been established (Albers and Kelly, 1989; Ho et al., 1997). Paranodal or segmental demyelination may be distributed diffusely along the nerve fiber, as seen in most hereditary hypertrophic neuropathies, or focally or multifocally along the nerve fibers in acquired demyelinating poly- and mononeuropathies, including entrapment neuropathies. Accordingly, the pathophysiological changes in hereditary hypertrophic polyneuropathies are distributed equally along proximal and distal parts of the PNS, and conduction block or increased temporal dispersion are not characteristic features probably due to the remodeling of the ionchannel distribution in these chronic disorders. In contrast, conduction block or increased temporal dispersion or both are characteristic features of motor conduction studies in acquired demyelinating mono-

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and polyneuropathies. The changes in motor and sensory conduction differ in acquired demyelinating neuropathies. Sensory conduction studies in inflammatory demyelinating neuropathies are characterized by dispersion of the SNAP, whereas the conduction velocity is less reduced and conduction block less often apparent than in motor fibers (Krarup and Trojaborg, 1996). These differences in pathophysiological changes in sensory compared with motor fibers (Krarup et al., 1990a) may occur on the basis of antigenic differences or that sensory fibers due to their axonal characteristics (Burke et al., 2001; Lin et al., 2002) may be less prone to conduction block. Finally, it should be considered that detection of sensory conduction block requires other methodological considerations than in motor fibers due to the differences in MUPs compared with SFAPs. Slowing of conduction is also out of proportion to the diameters of the largest fibers in diabetes mellitus, and since demyelination is not a prominent feature in diabetic neuropathy it was suggested that ion-channel dysfunction occurs in diabetic nerve (Behse et al., 1977; Brismar, 1979; Brismar and Sima, 1981; Brismar, 1985). Changes in ion-channel function have been found during electrotonus in both humans (Horn et al., 1996) and experimental models (Yang et al., 2001). Axonal atrophy is associated with slowing of conduction but is difficult to distinguish from loss of large diameter axons. An increased g-ratio (Baba et al., 1982) and a reduced index of circularity (Arbuthnott et al., 1980a) supports the suggestion of axonal atrophy (Dyck et al., 1981; Dyck, 1993). Circularity was normal in diabetic nerve indicating that atrophy is not a prominent feature (Engelstad et al., 1997). Reduction in filament spacing directly indicate that the small fiber diameter in anti-MAG neuropathy is due to axonal atrophy (Lunn et al., 2002). Axonal loss is by far the most frequent consequence of metabolic, toxic, autoimmune, hereditary and physical lesions of the PNS. The hallmark electrophysiological feature is reduced amplitudes of motor or sensory responses or both according to the underlying disease. Chronic partial motor fiber loss may, however, escape detection if the possibility of collateral sprouting and reinnervation is not considered specifically, and the amplitude of the SNAP is more sensitive to chronic axonal loss than that of the CMAP. The electrodiagnostic examination should, therefore, include nerve conduction studies of nerves in both upper and lower limbs to assess motor and sensory fibers and EMG of distal and proximal muscles to

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determine the presence of chronic partial denervation, and MUNE may be useful to support the presence of axonal loss. Electrodiagnostic studies are particularly important in the diagnosis of sensory neuropathies (Rosenberg et al., 2001) and to assess the distribution and degree of abnormalities of patients with mononeuropathy multiplex (Krarup, 2002). Acknowledgements The work was supported by the Danish Research Council, the Lundbeck Foundation, the Novo Nordic Foundation and the Ludvig and Sarah Elsass Foundation. References Aggarwal, A and Nicholson, G (2002) Detection of preclinical motor neurone loss in SOD1 mutation carriers using motor unit number estimation. J. Neurol. Neurosurg. Psychiatry., 73: 199–201. Aidley, DJ (2001) The Physiology of Excitable Cells. 4th ed., Cambridge University Press, Cambridge. Albers, JW and Kelly, JJ (1989) Acquired inflammatory demyelinating polyneuropathies: clinical and electrodiagnostic features. Muscle Nerve, 12: 435–451. Alfonsi, E, Merlini, GP, Giorgetti, A, Ceroni, M, Piccolo, G, Agostinis, C and Savoldi, F (1987) Temperature-related changes in sensory nerve conduction: studies in normal subjects and in patients with paraproteinaemia. Electromyogr. Clin. Neurophysiol., 27: 277–282. Aminoff, M, Chelimsky, T, Fealey, R, Freeman, R, Gutrecht, J, Harati, Y, Hiner, B, Hoeldtke, R, Hubbard, J, Kaufmann, H, Kennedy, W, Khurana, R, Pfeifer, M, Polinsky, R, Russell, J, Stewart, J, Ferguson, JH, Altrocchi, PH, Brin, M, Goldstein, ML, Gorelick, PB, Hanley, DF, Lange, DJ, Nuwer, MR and vandenNoort, S (1996) Assessment: clinical autonomic testing report of the therapeutics and technology assessment subcommittee of the American Academy of Neurology. Neurology, 46: 873–880. Aminoff, MJ (1998) Electromyography in clinical practice. Clinical and Electrodiagnostic Aspects of Neuromuscular Disease. 3rd Ed. Churchill Livingstone, New York, Edinburgh. Andersen, B, Westlund, B and Krarup, C (2003) Failure of activation of spinal motoneurons after muscle fatigue in healthy subjects studied by

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Peripheral Nerve Diseases Handbook of Clinical Neurophysiology, Vol. 7 J. Kimura (Ed.) © 2006 Elsevier B.V. All rights reserved

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CHAPTER 3

Nerve biopsy Gyl Midronia,b, * and Juan M. Bilbaob,c a

Clinical Neurophysiology, St. Michael’s Hospital, Toronto, Canada b University of Toronto, Canada c Sunnybrook Health Sciences Center, Canada

3.1. Introduction This chapter reviews the application of nerve biopsy in management of patients with peripheral neuropathy, with emphasis on issues of interest to clinical neurophysiologists. The discussion will include a review of the process of selecting and sending patients for nerve biopsy, and a review of relevant findings. We will focus on a clinically useful approach. The chapter is not intended to provide a photomicrographic atlas of peripheral nerve pathology, and the interested reader should consult monographs dedicated to this broad topic (Midroni and Bilbao, 1995; King, 1999). 3.2. The contrasting nature of nerve biopsy and electrodiagnostic testing The information obtained from histological examination of the peripheral nerve tissue and the indications for nerve biopsy differ significantly from the information provided by and indications for electrodiagnostic testing. Electrodiagnostic examination (EDX) can assess function and identify axon and myelin dysfunction along numerous motor and sensory nerves. Clinically relevant peripheral nerves can be assessed, and a direct clinical—electrophysiological correlation can be drawn. Electrodiagnostic examination can assess severity of denervation, assess relative contributions of axonal loss or demyelination to motor deficits, and provide information about prognosis and progression. However, EDX assesses function, not structure. One cannot directly

* Correspondence to: Dr Gyl Midroni, MD FRCP(C). Director-Clinical Neurophysiology, St Michael’s Hospital, 30 Bond St, Toronto, Ontario, Canada M5B 1W8. E-mail address: [email protected] Tel.: +416-864-5193; fax: +416-864-6094.

infer the underlying pathology from EDX. In addition, the EDX assessment for demyelinating neuropathy focuses exclusively on motor involvement, with minimal information gathered from sensory fibers. Nerve biopsy directly contrasts with EDX. Only nerve biopsy can reveal the underlying pathology, usually interstitial and not neural, that identifies the etiology of a peripheral neuropathy. Nearly all of the most common indications for nerve biopsy relate to the search for interstitial pathology such as vasculitis, amyloid deposits, sarcoid granulomata, or various stored materials. Moreover, assessment of sensory fiber demyelination is a particular strength of nerve biopsy analysis—since a sensory nerve is biopsied. Techniques such as electron microscopy or fiber teasing may reveal subtle pathological findings that are not significant enough to cause a noticeable change in EDX, yet are highly significant for making an etiological diagnosis. The frequent failure of nerve biopsy as a diagnostic tool arises because this method can assess only the anatomy of a several centimeter segment of an exclusively sensory peripheral nerve. Any inference made from this to what may be occurring in the entire peripheral nervous system is uncertain. This sampling error significantly reduces the correlation between findings of demyelination and axonal loss on nerve biopsy, and the determination on EDX that a diffuse neuropathy is “axonal” or “demyelinating” (Logigian et al., 1994; Molennar et al., 1998; Haq et al., 2000; Bosboom et al., 2001). Because of the sampling issue, nerve biopsy generally cannot be used to meaningfully assess the severity of a neuropathy, nor predict prognosis, especially in the setting of non-axonal or non-length dependent disease. Hence, clinicians must use EDX and nerve biopsy in a complementary fashion. While EDX is the most important test in workup of peripheral neuropathy, necessary in all patients where this condition is suspected, nerve biopsy is reserved for those instances where no probable or definite diagnosis can be made

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using clinical judgement and all other laboratory means. When all else has failed, nerve biopsy may reveal the cause of a neuropathy and alter management. Still, nerve biopsy is not carried out with every undiagnosed neuropathy—in practice only a tiny minority of patients with peripheral neuropathy require biopsy—patient selection is paramount. 3.3. Selection of patients for nerve biopsy 3.3.1. Nerve biopsy as a diagnostic tool A diagnostic procedure should only be used if it provides information that impacts on patient management or treatment outcome. This is particularly true if that procedure has significant morbidity. With rare exceptions, nerve biopsy is not useful for prognostic purposes or for assessment of treatment efficacy (discussed below). Hence, almost the only indication for nerve biopsy would be the attempt to make an etiological diagnosis for an undiagnosed and sufficiently severe neuropathy. This is discussed in detail below. As nerve biopsy is more sensitive for detection of mild neuropathy than nerve conduction studies (Section 3.7.1), sometimes biopsy is performed to confirm a diagnosis of peripheral neuropathy (Logigian et al., 1994; Deprez et al., 2000). However, the clinical importance of making this distinction in patients whose peripheral neuropathy is so mild that it does not produce nerve conduction testing abnormalities is uncertain, and we would not routinely endorse this indication. Similarly, performing a biopsy to assess disease severity may have limited usefulness in CIDP (discussed in Section 3.6.3.2.), but this is a marginal benefit at best, and does not alter management and generally does not justify the morbidity of the procedure. 3.3.2. Potential of nerve biopsy to alter management Table 3.1 lists the ways in which nerve biopsy may be helpful. By far the most common indication for biopsy is the search for a dysimmune process that causes nerve damage, in the hope that treatment with immune modulating agents will provide benefit. The usual immune modulating treatments are intravenous immune globulin (IVIg), corticosteroids, cytotoxic agents, and other immune suppressive agents. In every instance the treatment regimen involves considerable costs and/or risk to the patient. In nearly every case, the treatments do not “cure” the condition, and treatment has to be sustained

MIDRONI AND BILBAO Table 3.1 Impact of nerve biopsy on patient management (1) Diagnosis of a potentially treatable neuropathy ● Immune-mediated neuropathy ● Vasculitis (primary PNS or secondary to systemic disease) ● Other unspecified Inflammatory neuropathy ● CIDP ● Sarcoidosis, perineuritis ● Paraprotein associated neuropathies ■ anti-MAG syndrome ■ POEMS syndrome ● Other treatable cause of neuropathy ● Leprosy (biopsy may also be useful for assessing treatment efficacy) ● amyloidosis ● CMV neuritis ● Neoplastic infiltration ● HIV diffuse infiltrative lymphocytosis syndrome (DILS) ● certain toxic exposures ■ Hexacarbon toxicity (neurofilament accumulation) ■ amphililic cationic drugs (Amiodarone, Perhexilene, Chloroquin) (2) Diagnosis of a genetically determined peripheral neuropathy (untreatable, but patient avoids unnecessary treatments and provides information about prognosis, for genetic counseling, etc) ● HNPP ● CMT-1 ● Congenital dysmyelinating neuropathies ● Storage diseases ● Others (giant axonal neuropathy, polyglucosan body disease) (3) Confirmation of diagnosis of neuropathy (limited value) (4) Assessment of severity of neuropathy (limited value)

for months or years. Undertaking such treatment would not be considered appropriate unless the neuropathy was sufficiently disabling and progressive that a reasonable patient and physician would be willing to accept the risks and burdens of such a regime. A patient must fulfil this criterion to be considered for nerve biopsy. The issue of progression of neuropathy should also be considered prior to decision about nerve biopsy. No matter how disabling, a neuropathic process that has not progressed in 1–2 years is not likely to be responsive to any treatment (the rare exception being multifocal motor neuropathy—a diagnosis where biopsy is not useful). Long-established axonal loss cannot be reversed, and treatment would at best stop further progression. Hence,

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if there is no progression, there is no role for potentially dangerous treatment. If there is no role for treatment, why potentially cause further morbidity with a biopsy? A unique indication for nerve biopsy occurs in the case of known neuritic leprosy, where the nerve specimen may provide a more accurate reflection of residual disease than skin biopsy specimens may indicate, and thus impact on selection of the antibacterial regime (Chimelli et al., 1977; Nilsen et al., 1989).

65 Table 3.3 Alternative tissues to be considered for biopsy ●

● ●

● ●

3.3.3. Other, less invasive, diagnostic means should be exhausted An increasing number of genetic diseases can be diagnosed by non-invasive means. Literature on nerve biopsy from 1 to 2 decades ago is dominated by reports of biopsy pathology in Charcot–Marie–Tooth (CMT) disease and Hereditary Neuropathy with Pressure Palsies (HNPP). Other neuropathies that previously required biopsy confirmation, genetic and even acquired, can be diagnosed serologically (Table 3.2). These indications for nerve biopsy have been abolished except under rare instances (when the non-invasive testing can be falsely negative or is not available). Moreover, many diseases that cause neuropathy may be more readily diagnosed by biopsy of other tissues (Table 3.3). Skin biopsy with quantitative assessment of dermal unmyelinated nerve fibers has recently emerged as a superior means for confirming diagnosis of small fiber neuropathy (Holland et al., 1998; Periquet et al., 1999). Table 3.2 Past indications for nerve biopsy, now diagnosed noninvasively (not including diseases in which biopsy pathology is nonspecific) Genetically Determined Disorders Charcot–Marie–Tooth disease type I (multiple genetic defects) ● known genetic causes of Dejerine Sottas Syndrome ● Hereditary Neuropathy with Pressure Palsies ● Familial Amyloid Neuropathy ● Storage diseases ● leukodystrophies ● gangliosidoses ● neuronal ceroid lipofuscinosis Acquired Neuropathies ● anti-Myelin Associated Glycoprotein (anti-MAG) Paraproteinemic neuropathy ●

Muscle (vasculitis, sarcoidosis, amyloidosis, storage disorders) Skin (amyloidosis, neuronal storage diseases) Salivary Gland (Sjogren’s syndrome, amyloidosis, sarcoidosis) Rectal mucosa (neuronal storage disorders, amyloidosis) Kidney (vasculitis, amyloidosis)

3.3.4. Combined nerve and muscle biopsies If a search for vasculitis, amyloidosis, or sarcoidosis is the primary indication for the biopsy then a combined nerve and muscle procedure should be carried out, as in many instances the muscle is diagnostic when the nerve is not (Said et al., 1988, 2002b; Collins et al., 2000; Deprez et al., 2000; Prayson, 2002). The potential increase in yield is substantial (see discussion of vasculitis below). Moreover, the morbidity of muscle biopsy is less than that of nerve biopsy, and often the muscle can be taken through the same incision as the nerve. Even if two incisions have to be made, the two separate wounds would heal simultaneously, limiting the amount of recovery time for the patient. This applies both to sural nerve biopsy with simultaneous biopsy of the gastrocnemius muscle, or superficial peroneal nerve biopsy, with simultaneous biopsy of the peroneus brevis muscle (Collins et al., 2000). Routinely collecting small specimens of skin and fat for histological examination will also increase the yield slightly at no extra cost in morbidity to the patient (amyloid, sarcoid, some storage diseases). 3.3.5. Obtaining consent for nerve biopsy The last step prior to arranging the nerve biopsy should be a fully informed patient’s consent to proceed with this procedure. This includes not only complete discussion of what the biopsy experience involves and the risk of short- and long-term morbidity (Table 3.4), but also a realistic review of the potential for impact upon management. The option to treat on suspicion of a specific diagnosis is always open, especially when CIDP is considered. In addition, it has been our experience on several occasions that patients agree to a biopsy, a “treatable” diagnosis is made, but then the patient refuses treatment with immune suppressive agents once the side-effects and risks of these modalities are discussed.

66 Table 3.4

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●

Talking points for nerve biopsy consent (1) Review Complications ● Pain of procedure itself (patient feels nerve being cut) ● Immobility for several days, local pain at incision for several days/weeks ● Poor healing or wound infection (2–22%, increased risk if on steroids) (Rappaport et al., 1993; Collins et al., 2000, Gabriel et al., 2000) ● Traumatic neuroma (1%) ● Spontaneous neuropathic pain/allodynia from cut nerve segment ● short and medium term (10–40%), (Dyck et al., 1984; Solders,1988) ● long term (1 year or longer) 10–33%, usually not severe (Dyck et al., 1984; Neundorfer et al., 1990; Perry and Bril, 1994; Gabriel et al., 2000) ● Numbness due to removal of nerve. This tends to markedly improve/resolve over months to years (Pollock et al., 1983; Theriault et al., 1998) (2) Explain Yield of biopsy ● biopsy alters management in about 20% of carefully selected cases, helps in about 20–40% of others, not helpful in about 50% (3) Discuss Impact on management ● Full review of risks and benefits of immune suppressive treatment if biopsy shows a “treatable” condition. Patient should agree in advance to be treated if biopsy shows something “treatable” (4) Offer the option of watchful waiting ● Continue to follow clinically—the process may stabilize or improve spontaneously ● Can reconsider biopsy if process is definitely progressing and disabling

We have found that when biopsy fails to yield a clear diagnosis patients may become frustrated, feeling that they have been subjected to this procedure for no reason, and that their hope for an answer was dashed. Others have also reported this (Gabriel et al., 2000). In fact, the majority of biopsies are not very helpful, no matter how carefully selected (Section 3.4.)—so patients should be prepared for this possibility in advance. Table 3.4 summarizes the points to review with patients as part of the consent. 3.3.6. Summary of indication for nerve biopsy With these considerations, we suggest that the following requirements are necessary to proceed with most nerve biopsies:

●

●

●

progressive and sufficiently disabling peripheral neuropathy of unknown etiology with objective deficits on exam and electrodiagnostic testing. The pattern of involvement is irrelevant. Although mononeuropathy multiplex has the highest diagnostic yield (Section 3.4.4.1), specific and treatable etiologies that can be diagnosed with biopsy may present with any distribution of deficits and any electrophysiological findings. no diagnosis has been made so far employing routine and specialized non-invasive tests, preferably including assessment at a specialized neuromuscular center. alternative sites of biopsy are either exhausted, or unlikely to yield an answer. fully informed patient’s consent has been obtained. (Table 3.4)

3.4. Yield of nerve biopsy It is helpful to have some idea of the diagnostic yield of nerve biopsy, and whether certain clinical or laboratory parameters can help select patients for this procedure. 3.4.1. Personal experience In our personal series of 234 consecutive biopsies over a 7-year period ending in 1993, with detailed clinical information available for review, biopsy was judged to be “essential” for patient management in 21% (49) cases (Midroni and Bilbao, 1995). Most often, the biopsy was essential because it revealed necrotizing vasculitis (19/49 cases). Other settings where a specific finding permitted correct diagnosis and optimal management were amyloidosis, sarcoidosis, leprosy, and infiltrative nerve lymphoma. In three cases, an unexpected findings of primary macrophage mediated demyelination was suggestive of CIDP even when this diagnosis had not been seriously considered because no findings of demyelination were seen on the nerve conduction studies done prior to the biopsy. Nerve biopsy was judged to be “helpful” in 22% of cases, in the sense that it supported a clinical diagnosis of CIDP, or it confirmed a suspected genetically determined neuropathy such as CMT-1, or Hereditary Neuropathy with Pressure Palsies (HNPP). However, in such instances the biopsy did not alter management either because no management was available in the case of a genetic disease, or because the management plans were the same before and after biopsy—specifically in

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the case of CIDP—hence the classification as “helpful” rather than “essential.” In addition, a decade later, the need for nerve biopsy for diagnosis of CMT-1 and HNPP has been almost eliminated. 3.4.2. Literature data In the only prospective work on the issue of how nerve biopsy impacts on management Gabriel et al. (2000) studied 50 consecutive nerve biopsies with the treating physician required to declare in advance of biopsy a treatment plan, and this plan then reconsidered in light of the biopsy results. Nerve biopsy altered the diagnosis and management in only 7 of 50 cases, 3 being unexpected vasculitis, the others being lymphomatous infiltration, CIDP, IgM anti-MAG neuropathy, and Guillain–Barré syndrome. It is arguable whether the last case should be included. However, the investigators also concluded that nerve biopsy was “helpful” in 60% of this carefully selected series of nerve biopsies by an expert neuromuscular group. The spectrum of pathology seen in this study is very similar to what we have seen in our experience. In an analysis of about 300 biopsies from two academic institutions Deprez et al. (2000) found that about 45% of biopsies were essential or helpful, with the essential biopsies representing about 1/3 of this 45%. Argov et al. (1989) found nerve biopsy to alter management in 19% of 53 consecutive biopsies. Neundorfer et al. (1990) felt that 27% of 56 sural nerve biopsies they reviewed were crucial for diagnosis. Logigian et al. (1994) found a 20% frequency of diagnostic interstitial pathology in 107 biopsies. In addition 5% of biopsies showed significant demyelination even though the electrophysiology was axonal (2/5) or indeterminate (3/5) (Logigian et al., 1994). Thus, 25% of the biopsies in this series were considered highly informative. In this last work the authors also note that in an additional 12% mild abnormalities on nerve biopsy were crucial in proving that the patient had a peripheral neuropathy rather than radiculopathy, but whether this altered patient management significantly is not stated. Chia et al. (1996) found that 33% of biopsies performed in 100 consecutive patients over the age of 65 with “disabling neuropathy” showed vasculitis. Oh found that 24% of 385 biopsies showed specific histology (Oh, 1990). Overall, then, a diverse series of reports have consistently found that even with careful selection only about 15–30% of biopsies provide information essential to patient management, and that an additional 20–40% are

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helpful. The two most important findings are evidence for necrotizing vasculitis, or unexpected evidence of primary demyelination in an apparently axonal polyneuropathy (Haq et al., 2000, Vallat et al., 2003). The proportion of “helpful” biopsies is likely to be on the low end of the 20–40% range, as in most of the studies quoted above biopsy was considered to be helpful if it confirmed a genetic neuropathy or confirmed a diagnosis of CIDP, which in current practice are largely obsolete indications for biopsy. Viewed from another perspective it appears that about half of biopsies provide no useful management modifying information other than confirming the presence of a neuropathy. 3.4.3 Predicting usefulness of biopsy with electrodiagnosis Normal nerve conduction studies, or a normal sural sensory response, predict a lower chance that nerve biopsy of the sural nerve will reveal important findings, but do not rule out this possibility altogether. A useful biopsy alters management, and in our personal series 94% (31 of 33) patients with normal EDX had nerve biopsies which had no impact on clinical management (Midroni and Bilbao, 1995). Still, in 2 of 33 such biopsies unexpected and “essential” diagnoses of Fabry’s disease and sarcoidosis were made. Deprez et al. (2000) defined biopsy usefulness as we do, and found no useful biopsies in eight patients with normal nerve conduction testing. While a nerve biopsy may show that a neuropathy is present where electrodiagnosis fails to confirm neuropathy (Logigian et al., 1994) this could only occur in the setting of very mild neuropathy, and hence the clinical impact of such a finding is minimal. In our personal retrospective review series, there was no significant difference in the frequency of an “essential” biopsy in axonal vs demyelinating neuropathies (22%), but a demyelinating neuropathy was more likely to show a “helpful” finding (46 versus 19%) (Midroni and Bilbao, 1995). Our experience mirrors that of other workers in the field. Gabriel et al. (2000) had smaller numbers, but this study had the advantage of being rigorous and prospective. These investigators found no clear predominance of axonal or demyelinating electrophysiological features in the seven nerve biopsies that altered management, and a slight but not significant predominance of demyelinating neuropathies in the “helpful” biopsy group. Deprez et al. (2000) found no difference

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in essential or helpful biopsy results between axonal, demyelinating, or mixed neuropathies (41, 46, and 37%, respectively), nor did Neundorfer et al. (1990) (N = 56). Only Argov et al. (1989) found that biopsy usefulness was higher in demyelinating processes (65%, 17/26) than in axonal processes (11%, 3/27). However the definition of a “useful” biopsy included many patients in whom Charcot–Marie–Tooth or CIDP was confirmed, indications that would not be appropriate for nerve biopsy in current practice. 3.4.4. Predicting usefulness of biopsy based on clinical features 3.4.4.1. Mononeuropathy multiplex Logic, personal experience, and literature data support the notion that nerve biopsy is more likely to be useful in a multifocal peripheral neuropathy than in a diffuse distally predominant and symmetrical process. The causes of mononeuropathy multiplex are usually inflammatory conditions such as vasculitis in which biopsy is often diagnostic, while the causes of diffuse distally predominant neuropathies are generally axonal metabolic conditions in which there is no diagnostic pathology to be found. This impression is confirmed by Deprez et al. (2000) with biopsy results being essential in 7/16 cases and helpful in 5/16 cases who had a mononeuropathy multiplex picture. This is contrasted with 3/25 essential and 12/25 helpful in patients with an asymmetrical polyneuropathy, and 17/197 essential and 60/197 helpful biopsies in patients with a distal symmetrical polyneuropathy. Similarly, in a prospective study of 50 biopsies, where management was modified in seven cases, five of the seven essential biopsies came from 9 patients with mononeuropathy multiplex (Gabriel et al., 2000). Chia et al. (1996) in a study of 100 consecutive patients with disabling neuropathy over 65 years of age found vasculitis in 33 patients, 21 of whom had a mononeuropathy multiplex picture. Patients with other etiologies in this group (paraproteinemia, drug-induced neuropathy, alcohol, diabetes, etc.) in which biopsy was not diagnostic generally had a symmetrically distally predominant neuropathy. The exceptions were 14 patients with CIDP, most of whom (66%) had a symmetrical picture. Such data strongly suggest that mononeuropathy multiplex is a better indication for biopsy than distally predominant symmetrical diffuse polyneuropathies. Asymmetrical polyneuropathy patients are an intermediate group, which presumably have an intermediate biopsy yield.

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3.4.4.2. Small fiber neuropathy There is no literature data on the topic, but experience indicates that small fiber neuropathy in general has a low biopsy diagnostic yield. There are some rare conditions that might be unexpectedly diagnosed by nerve biopsy—amyloidosis in particular, very rarely Fabry’s disease, Tangier disease, sarcoid neuropathy or vasculitis (Lacomis et al., 1997). However, the practical impact on management is limited and a higher yield/less invasive means to try to confirm the diagnosis is usually present (Section 3.3.3). We have found that patients with neuropathic pain, especially those with relatively little objective deficit, seem to have a higher incidence of sustained post biopsy pain, sometimes indefinitely. 3.4.4.3. Motor dominant neuropathy A pure or predominantly motor syndrome is less likely to show useful pathological findings, primarily because the biopsy is taken from a sensory nerve. However, there are sufficient exceptions to this rule that one should still consider biopsy—especially in the context of CIDP, where biopsy alone may provide the evidence for primary demyelination (see Section 3.6.3). 3.4.4.4. Late onset sensory predominant polyneuropathy Within the larger group of patients with undiagnosed neuropathy, all clinicians who deal frequently with peripheral neuropathy are familiar with the subgroup of older patients who present with an indolent distally predominant, often sensory dominant, peripheral neuropathy. Pain can be a prominent feature, while motor deficits are minimal or absent and deficits confined below the knees even after several years of symptoms. Initial diagnostic testing reveals no answer, and the electrophysiology is axonal. With very few exceptions these patients should not undergo biopsy. The commonest identifiable etiologies in this group are thought to be mild geneticallydetermined axonal neuropathies, paraproteinemia, alcohol abuse, and impaired glucose tolerance. None of these have pathognomonic findings on biopsy, explaining why the biopsy yield in this group is very low (Dyck et al., 1981b; McLeod et al., 1984; Grahmann et al., 1991; Notermans et al., 1994, 1996; Wolfe et al., 1999; Sumner et al., 2003). The two most common biopsy-diagnosable conditions, vasculitis and CIDP, rarely present with this clinical picture. In addition, even if a biopsy were to reveal an inflammatory process the treatment in this group of

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elderly patients (corticosteroids or other immune modulating agents) may be worse than the problem. Most often, no treatment would be undertaken unless the process had evolved into something significantly more disabling. This is not to say that older patients with peripheral neuropathy should not undergo biopsy—clearly there is a high diagnostic yield in patients selected properly—the 100 consecutive patients over 65 described by Chia et al. (1996) had a “disabling” neuropathy, and would not fit into this group of patients. 3.4.5. Nerve biopsy specificity and sensitivity The ability of nerve biopsy to detect a given process depends on the particular condition and on the amount of tissue taken. This is discussed below in Section 3.6, where the conditions for which biopsy is performed are reviewed. Specificity is less often considered in regard to biopsy interpretation, but the clinician must be wary of assuming that the biopsy is the “final answer.” There are a number of instances where the biopsy report may mislead. Age related changes are often misinterpreted as indicators of mild pathology, both axonal and demyelinating. The changes with age that electromyographers are accustomed to accepting with regard to normal sural sensory amplitude and conduction velocity are mirrored in the histological changes seen in this nerve with age—up to a 50% drop in nerve fiber density, shift in the fiber diameter histogram towards small fibers with thinner myelin, increase in numbers of regenerating clusters, and a marked increase in the findings of demyelination/remyelination on teased fiber studies. (Tohgi et al., 1977; Jacobs and Love, 1985; Midroni and Bilbao, 1995). A few polyglucosan bodies may be a normal age-related finding, and not indicative of the extremely rare condition Adult Polyglucosan Body Disease (Midroni and Bilbao, 1995). “Pseudo-onion bulbs” that are simply indicative of axonal regeneration can be hard to distinguish from genuine “onion bulbs” (indicative of demyelination/ remyelination) and may lead a pathologist astray (Midroni and Bilbao, 1995). The emergence of techniques to detect inflammatory cells (lymphocyte markers) facilitates the identification of inflammation in the nerve. However, lymphocytes are found normally in peripheral nerve, often in a perivascular pattern, and there is the risk of over-interpreting the cells as an indicator of inflamma-

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tory neuropathy (Bosboom et al., 1999). Even an abnormal increase in inflammatory cells is nonspecific. A collection of perivascular inflammatory cells, however prominent, does not constitute vasculitis unless there is evidence of vessel destruction (Section 3.6.2). Amyloidosis must be diagnosed with caution. The diagnosis is obvious when there are large intraneural pools of congophilic material. However, a nerve specimen may reveal focal Congo Red staining due to fibrin, elastin fibers, or mast cell granules. These materials do not exhibit birefringence under polarized light, but other non-amyloid materials can—some stored lipids, some fungi, plant cell walls, and cotton have this property. Oxytalan filaments, a normal component of the elastic fiber system in the endoneurium, also have been misinterpreted as amyloid fibrils at the ultrastructural level. One must always question the significance of small amyloid deposits in epineurial connective tissue; it may be “senile” amyloid, or secondary amyloid and, hence, not relevant to the neuropathic process. All these and other issues are best addressed by close communication between a pathologist familiar with clinical aspects of neuropathy and a neurologist who understands the strengths and weaknesses of nerve biopsy as a diagnostic tool. When a biopsy finding does not seem plausible, it should be reconsidered in light of the known limitations of the method. 3.4.6. Diagnostic yield of nerve biopsy in children There is little information available on the yield of nerve biopsy in children—it probably is substantially lower than in adults given the different disease spectrum in this population. In particular, CIDP is less common than genetic conditions as the cause of early onset demyelinating neuropathy (Plante-Bordeneuve et al., 2001), and vasculitis and paraproteinemia vanishingly rare (Ouvrier et al., 1999). However, treatable neuropathic etiologies do occur in childhood, and CIDP can be particularly difficult to distinguish from a genetic condition in very young patients (PlanteBordeneuve et al., 2001). Genetic and biochemical testing has supplanted biopsy as the primary means of diagnosis of various genetically determined neuropathies and storage diseases. However, atypical presentations and the remaining conditions for which genetic tests do not yet exist (predominantly, a variety of congenital dysmyelinating neuropathies, discussed in Section 3.6.8.2) may still require invasive testing. Nerve biopsy remains an option in the workup of

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pediatric neuropathies if all other means have been exhausted and the patient disabled—as is the case in adult-onset peripheral neuropathy. 3.5. Technical aspects of nerve biopsy 3.5.1. Surgical removal The sural nerve is most commonly biopsied because of several favorable attributes including easy access for surgical removal and correlation with electrodiagnostic testing, relative protection from local trauma, distal location (most neuropathies are distally predominant), and functionally insignificant deficit resulting from removal (Dyck and Lofgren, 1968; McLeod et al., 1969). In addition, there is a large literature on the normal findings in this nerve. Another commonly biopsied nerve is the superficial peroneal nerve at the ankle, with similar favorable features (Collins et al., 2000; Mendell et al., 2001). Other sensory nerves have been studied to a much lesser extent (superficial radial, dorsal ulnar cutaneous nerve, femoral sensory branch). Motor nerves are almost never biopsied— although there are anecdotal reports of this technique (Stevens et al., 1973; Corbo et al., 1997). Fascicular nerve biopsy had been advocated for a time as a less morbid alternative to full thickness nerve biopsy (Dyck and Lofgren, 1966). In this procedure, the nerve is carefully dissected intraperatively and a few fascicles removed, the nerve remaining in continuity. However, this technique has generally been abandoned as it compromises the principal reason for doing the biopsy, making a diagnosis. A search for vasculitis is the commonest indication for nerve biopsy, and as discussed below (Section 3.6.2) there is no clinical or electrodiagnostic means of ruling out this possibility. The diagnostic pathology is almost invariably found in the epineurium (Collins et al., 2000), and a partial biopsy will reduce the amount of epineurial tissue available for examination and seriously compromise the procedure. Moreover, the intimate neural handling involved in a fascicular biopsy is likely to produce artifacts that will cloud interpretation. Finally, it is not clear that fascicular nerve biopsy has a lower morbidity than full-thickness nerve biopsy (Pollock et al., 1983; Solders, 1988; Gabriel et al., 2000). Having a pathology technologist present at the time of the surgical procedure is essential. Removing the nerve and putting it into a formalin filled jar for transport is unacceptable. The technologist should receive the specimen fresh, stretch it slightly on a flat wooden

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stick, remove a segment for snap freezing in isopentane-liquid nitrogen, and immerse the remainder in cooled 2.5% glutaraldehyde in 0.025–0.05 M cacodylate buffer at pH 7.4 and osmolarity of 340 mOsm for transport to the laboratory. In the lab portions of the nerve are placed in B5 fixative (or formalin) for paraffin embedding and fiber teasing, and the rest returned to gluteraldehyde for further preparation including plastic resin embedding and teasing. Tissue preparation takes several days, and if there is a need for urgent diagnosis of an interstitial change, especially vasculitis, this can be done on the spot with a quick section of frozen nerve tissue. 3.5.2. Histological techniques An experienced pathologist with special interest in nerve biopsy interpretation should be involved in review of a biopsy specimen. Access to the full range of specialized techniques involved in nerve biopsy interpretation is mandatory (see below). There are two key types of information to be collected. Most important is whether a specific etiologic diagnosis can be made, such as necrotizing vasculitis, granuloma detection, amyloidosis, leprosy, etc. Almost always, this is a finding made on the basis of abnormalities in the interstitial compartment, i.e., not the axons or myelin, but the supporting tissues of the epineurium, and less often the endoneurium or perineurium. This assessment is mostly done with standard histochemical and immunohistochemical techniques on paraffin embedded 5–10 μm “thick” sections. A secondary aim is to characterize the process as axonal or demyelinating. This is done with more specialized techniques that highlight neural tissue, such as plastic resin embedded tissues which permit cutting of 1 μm sections “semi-thin” sections for light microscopy, and 0.1 μm “thin” sections for electron microscopy, as well as nerve fiber teasing and quantitative morphometric analysis where appropriate. Assessment of interstitial change is relatively straightforward and mostly along the lines of traditional histology. However, the identification of axonal and demyelinating changes is more challenging, very specialized to the field of peripheral nerve pathology, and subject to a high degree of inter-observer variability (Logigian et al., 1994; Bosboom et al., 2001). Assessment of the neural tissues is less likely to provide diagnostic information than assessment of the interstitial compartment, but in some cases biopsy provides evidence of a demyelinating neuropathy in a

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process previously thought to be axonal in nature based on electrophysiological grounds—raising the possibility of CIDP (Section 3.6.3.2.). A particular point of difficulty is clarifying which findings are artifacts of preparation, and which are key indicators of nerve disease—this requires considerable experience and understanding of technical issues involved in biopsy (Midroni and Bilbao, 1995). Another difficult issue is interpretation of age-related change, which are easily misinterpreted as indicators of mild or even moderate neuropathy (Tohgi et al., 1977; Jacobs and Love, 1985; Midroni and Bilbao, 1995). The elements of a nerve biopsy examination are listed in Table 3.5. In our practice, step 1 is performed in all cases, step 2 in about half of cases, fiber teasing in a tiny minority of cases (but this depends on the resources available to the laboratory), and morphometry predominantly used as a research tool. The reader interested in a detailed discussion of these issues is referred to a monograph on the subject (Midroni and Bilbao, 1995). 3.5.2.1. Light microscopy After fixing and embedding in paraffin, full crosssections as well as longitudinally cut “thick” sections are stained with Hematoxylin-eosin-Luxol fast-blue, PAS, fibrin stains and Congo Red. Serial sections stained with these, as well as with Perl’s Prussian blue for iron deposition, facilitate a search for vascular injury. Staining for acid-fast organisms is used when leprosy is suspected. Immunostains for lymphocytes, and smooth muscle actin, and endothelium are also routinely employed. Other immunostains can be used in specific settings: transthyretin and immunoglobulin light chains to characterize amyloid deposits, IgM heavy chain for anti-MAG syndrome, endothelial immunolabels for study of possible vascular injury, Table 3.5 Steps in assessment of a nerve biopsy specimen (1) Light microscopy ● assessment of fascicular anatomy and specimen quality ● standard histochemistry on paraffin sections ● immunohistochemistry on paraffin or frozen sections ● assessment of semi-thin sections (2) Electron microscopy (3) Fiber teasing (4) Morphometry

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epithelial membrane antigen for identification of perineurial cells, and neurofilament immunostain for quick quantification of unmyelinated fibers. Although it is possible to highlight axons or myelin on paraffin embedded tissue using special techniques, this is unnecessary given the superior results obtained from plastic embedded semi-thin sections—the primary goal of reviewing paraffin embedded material is to assess the interstitium. Storage material is sometimes not identified in paraffin embedded material, as the preparation process removes some lipids. Frozen sections histostained with Hirsch-Peiffer stain are needed to look for metachromatic leukodystrophy, and Oil-Red-O for other lipid accumulations (other leukodystrophies, sphingolipidoses, Tangier disease). After fixing in gluteraldehyde and embedding in plastic (Epon/Araldite), semi-thin cross- and longitudinal-sections are counterstained with Toluidine Blue (or equivalent) stain. This tissue is optimal for light microscopic examination of axon and myelin changes (Section 3.6.1.). 3.5.2.2. Electron microscopy Plastic embedded material may be cut “thin” (0.1 μm sections) and examined under the electron microscope. Previously osmicated resin-embedded material stained with lead citrate highlights myelin and other lipid membranes, producing an appearance similar to that obtained with Toluidine blue stained semi-thin sections, but of a much higher resolution. This method will show unmyelinated fibers in a more detailed manner than other techniques, and permit their quantification. Electron microscopy may also clarify the nature of ongoing demyelination—primary macrophage-mediated demyelination, which is strongly suggestive of CIDP or Guillain–Barré Syndrome, or less often widely spaced myelin (indicative of anti-MAG syndrome) or uncompacted myelin (suggestive of POEMS syndrome). Conversely, what may appear to be demyelination on light microscopy will sometimes be revealed as an early stage of axonal degeneration. Electron microscopy has the potential to significantly add to diagnostic yield. Deprez et al. (2000) found added information in 14% of biopsies taken for EM study, while Haq et al. (2000) found that EM findings of macrophagemediated demyelination were the most sensitive and specific findings on nerve biopsy for CIDP, as well as the best predictors of treatment responsiveness. Electron microscopy is also the best method for detection of some storage material (metachromatic

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leukodystrophy, globoid cell leukodystrophy, adrenomyeloneuropathy, Fabry’s disease, Farber’s disease (Midroni and Bilbao, 1995) as well as detection of tubuloreticular inclusions, which point towards possible diagnoses of SLE or HIV-associated neuropathy (Midroni et al., 2000). Electron microscopy should be performed on any biopsy specimen in which the initial light microscopic biopsy review has not revealed a diagnosis. 3.5.2.3. Fiber teasing Nerve fiber teasing is generally regarded as the gold standard for clarifying the axonal versus demyelinating nature of a peripheral nerve process, subject of course to the limitation that the process in the biopsied segment be accurately reflective of the more diffuse process (Section 3.7). This method requires considerable technical expertise in both collection and preparation of the specimen. The tissue has to be removed for teasing soon after of initial arrival at the pathology laboratory, placed in osmium tetroxide in Millonig’s buffer at pH 7.4, then dehydrated and macerated in glycerin from 30 to 100%. The resulting nerve tissue can be manually “teased” under a dissecting microscope such that single axons and their surrounding myelin are separated for individual examination. Fiber teasing permits an excellent assessment of the relation between axons and their myelin, and may be the most sensitive means of detecting subtle demyelination. However, the degree to which fiber teasing adds management-modifying value to the biopsy is probably minimal. Logigian et al. (1994) found a 79% (41/52) concordance between interpretation of fiber teasing and cross sections. Deprez et al. (2000) found a similar concordance rate of 83% in 102 biopsies, but noted that teasing added useful information in only 4% of the 102 nerve biopsies. Teasing confirmed a diagnosis of HNPP in three cases and revealed primary demyelination to support a diagnosis of CIDP in one instance. In the latter study electron microscopy was more useful as an adjunctive technique than fiber teasing (Deprez et al., 2000). Bosboom et al. (2001) found that the usefulness of teased fiber preparations in making a diagnosis of CIDP, and distinguishing CIDP from idiopathic axonal polyneuropathy, was very limited, essentially noncontributory, based on the inability of two pathologists to consistently distinguish between CIDP and idiopathic axonal polyneuropathy. In another study of CIDP patients, there was no correlation between elec-

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trodiagnostic and teased fiber criteria for demyelination, and electron microscopic criteria appeared to be both more sensitive and more specific for a diagnosis of CIDP than any teased fiber findings (Haq et al., 2000). Uncini et al. (1999) found that teasing did not help separate between diabetic neuropathy and CIDP. 3.5.2.4. Peripheral nerve morphometry Quantitative analysis of peripheral nerve material provides insight into patterns of anatomical change in various peripheral nerve disorders. It also injects some objectivity into what remains a very subjective interpretation of tissue, error prone and highly dependent on the observer’s experience and bias (Pamphlett and Sjarif, 2003). Fiber density and diameter histograms provide information about the numbers and size distribution of myelinated and unmyelinated fiber diameters. This can provide objective evidence of a small fiber neuropathy and point towards some of the specific etiologies of this syndrome—we have had the experience of amyloid neuropathy with no amyloid deposits on nerve biopsy, yet a small fiber neuropathy documented on biopsy and clinically (Fig. 3.1C). The G-ratio, which measures the ratio of a diameter of an axon without its myelin to the diameter with myelin, approaches 1 in the congenital dysmyelinating ratios where diffusely very thin myelin is seen. Quantification of such elements as regenerating clusters, denervated Schwann cell subunits, and collagen pockets, may also provide some insight into axonal loss. Statistical analysis of demyelinating changes on individual teased fibers may provide evidence that demyelination is “secondary” rather than primary. Readers interested in these and related techniques can refer elsewhere (Midroni and Bilbao, 1995). In current clinical practice of peripheral nerve pathology, these methods are rarely employed due to cost and more importantly due to the fact that they almost never lead to specific diagnoses that alter management. These techniques are mostly a research tool. 3.6. Nerve biopsy findings in specific disorders The pathologist reviews the biopsy with two goals: first, to characterize the neural changes and time course of disease activity (axonal, demyelinating, acute, chronic); and second, and more importantly, to attempt to identify the specific cause of the process. These issues are reviewed below.

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Fig. 3.1 Sural nerve biopsy specimen in cross-section, resin embedded tissue, stained with Toluidine Blue. Low power view (A) shows 7 fascicles. The epineurial compartment contains collagen, vessels (including arterioles), and adipose cells. In higher magnification (B) of the endoneurium, a normal population of large and small myelinated fibers is readily seen, with unmyelinated fibers also barely visible. In (C) there is severe loss of nerve fibers, but the remaining few are large myelinated axons. This relative sparing of large myelinated fibers is unusual and suggested the diagnosis of amyloid neuropathy in this patient, subsequently confirmed by other techniques.

3.6.1. Axon and myelin changes The two cardinal neural findings on nerve biopsy are those of myelin change and axon change. These are generally best assessed on the plastic-embedded semithin (1 μm) sections, with added input from special techniques such as EM, fiber teasing, or quantitative morphometry.

3.6.1.1. Assessment of axonal changes The pathologist estimates overall axonal loss subjectively, based on the expected density of myelinated axon profiles in the cross-section, allowing for age and for the particular nerve (Fig. 3.1). A disproportion between loss of small myelinated and large myelinated fibers is important if noted (Fig. 3.1C). Selective

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loss of small fibers is distinctly unusual and raises a limited differential diagnosis, with amyloidosis being the main concern if diabetes is ruled out. This assessment can be done more rigorously with morphometry, but is not necessary in most cases. Although the element of human error in quantifying nerve fiber loss is substantial (Pamphlett and Sjarif, 2003), the diagnostic value of nerve biopsy comes not from subtle differentiation of varying degrees of axon loss, but from detection of pathognomonic interstitial findings. Unmyelinated fiber density can be assessed on light microscopy using special methods, but is best done with electron microscopy. The examiner may identify cross-sections of nerve fibers with ongoing Wallerian degeneration, appearing as balls of myelin debris where a myelinated axon previously resided (Fig. 3.2), indicating that this segment of the axon has degenerated in the preceding weeks, i.e., subacute degeneration. Widespread Wallerian degeneration, especially if in a multifocal pattern, is suggestive of vasculitic neuropathy (Fig. 3.4). If no such profiles are seen on a background of reduced axon fiber density, the process is presumed to be more indolent and chronic, most typical of a genetic process. However, any sufficiently chronic process may take this appearance. Another finding indicative of chronicity is regenerative clustering-3 or more tightly apposed small

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myelinated fibers, usually in a circular formation, usually accompanied by some unmyelinated axons (Fig. 3.3A). This finding is indicative of an axon that had degenerated previously and has now undergone proximal sprouting with the regenerating fibers travelling together down the residual basal lamina of the original axon. Rarely, giant axonal swellings can be seen that may be the “growth cone” at the tip of a regenerating axon. All of these findings are nonspecific, but provide useful information as to the severity and tempo of a neuropathic process. 3.6.1.2. Assessment of myelin changes Demyelination is most dramatically seen when an axonal profile is seen to be entirely denuded of its myelin, with myelin debris typically present in adjacent macrophage processes (Fig. 3.3A). This appearance is highly suggestive of primary macrophage mediated demyelination, but it is sometimes hard to be sure that one is not seeing demyelination secondary to axonal disease, with scavenger macrophages just removing debris, not “stripping” myelin. Seeing more than a rare denuded axon is unusual in any setting except Guillain–Barré syndrome or CIDP. Much more often the examiner finds evidence of remyelination, with myelin inappropriately thin for axonal diameter (Fig. 3.3B). This occurs because remyelination

Fig. 3.2 Sural nerve biopsy specimen in longitudinal section, resin embedded tissue, stained with Toluidine Blue. Rows of myelin ovoids indicate subacute Wallerian degeneration.

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Fig. 3.3 Sural nerve biopsy specimen in cross-section, resin embedded tissue, stained with Toluidine Blue. Patients with CIDP. In figure A, one “naked” axon is surrounded by a debris-filed macrophage, and sits next to a regenerating cluster of axons. A thinly myelinated nearby indicates previous demyelination with remyelination. The overall population of axons in the cross-section is markedly reduced. In figure B, the population of axons is relatively preserved, but nearly all are thinly myelinated and onion bulbs are prominent (*).

produces a myelin ring which is not as thick as the original. Onion bulbs (Fig. 3.3B), wherein concentric layers of Schwann cell processes surround a central axon, suggest recurrent demyelination and remyelination events and are a strong indicator of a chronic demyelinating process. Caution should be taken to distinguish these from pseudo-onion bulbs (Midroni and Bilbao, 1995). 3.6.1.3. Pathological distinction between axonal vs demyelinating neuropathy The pathologist examines the features noted above and attempts to characterize the neuropathy as axonal, demyelinating, or mixed in nature, as well as quantify severity and chronicity. The assessment can be challenging, since chronic as well as hyperacute axonal pathology may produce secondary myelin change. Conversely, some degree of axonal degeneration is routine in most “demyelinating” neuropathies. The distinction comes down to the pathologist’s personal judgement, and this is fraught with potential for error, both because of human factors and because of the inherent limitations of the nerve biopsy (Section 3.7). Frequently no certain determination of whether

the process is axonal or demyelinating in nature can be made. 3.6.2. Vasculitis A search for vasculitis is the most common single indication for nerve biopsy, and necrotizing vasculitis is the commonest management altering finding on nerve biopsy (Oh, 1990; Logigian et al., 1994, Midroni and Bilbao, 1995; Chia et al., 1996; Deprez et al., 2000). Although vasculitic neuropathy is typically painful, sensorimotor, and multifocal, with concurrent multi-organ involvement, the process can be purely sensory, painless, symmetrical and distally predominant, and strictly confined to the peripheral nervous system (Kissel et al., 1985; Said et al., 1988; Midroni and Bilbao, 1995; Chia et al., 1996; Davies et al., 1996; Collins et al., 2000). Regardless of clinical features, it is always possible that nerve biopsy will show vasculitis as the unsuspected cause of a neuropathic process, and hence this is the single strongest indication for proceeding with nerve biopsy. The sensitivity of nerve biopsy for detecting vasculitis is uncertain because of the lack of a gold

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standard for diagnosis, and may vary with the tempo and distribution of the disease. A diagnostic yield of 50–70% may be a reasonable estimate if only nerve biopsy is carried out (Dyck et al., 1987; Said et al., 1988; Hattori et al., 1999). Deprez et al. (2000) found a higher sensitivity in patients with subacute/acute progression than in those with chronic progression (>6 months from onset to biopsy). Adding a muscle biopsy will increase the yield by 15–45% (Said et al., 1988; Collins et al., 2000; Deprez et al., 2000). The unequivocal histological diagnosis of vasculitis requires two elements—transmural infiltration of the vessel with inflammatory cells, and evidence of vessel destruction (fibrinoid necrosis, thrombosis, vascular proliferation or hemorrhage) (Fig. 3.4). This usually occurs on a background of massive Wallerian degeneration, often patchy rather than diffuse (Fig. 3.5). Mere inflammatory infiltration of the nerve or perivascular inflammatory cuffing is not sufficient for a diagnosis of vasculitis. However, perivascular inflammation accompanied by prominent Wallerian degeneration, or vascular thrombosis/hemorrhage, is suspicious. Karrhyorhexis (fragmentation of nuclear material of invading leukocytes) is another suggestive finding. In isolation, however, perivascular inflamma-

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tory cuffing is quite nonspecific, and should not be used as evidence for a diagnosis of vasculitis. The specimen should be exhaustively examined employing multiple stains and step-sections. The relevant vascular pathology is almost invariably confined to or dominant in the epineurial compartment (Davies et al., 1996; Ohkoshi et al., 1996; Collins et al., 2000), affecting arterioles and small arteries that have a clearly defined elastic lamina and often a muscular layer. Although there is considerable overlap, the larger epineurial vessels, 100–400 μm in diameter, are more likely to be affected in polyarteritis nodosa, rheumatoid vasculitis, Wegener’s or ChurgStrauss Angiitis. Smaller vessels in the 40–100 μm range are more likely to be affected in nonsystemic vasculitic neuropathy, SLE, or microscopic PAN (Ohkoshi et al., 1996). True endoneurial vasculitis is very uncommon, but when seen strongly suggests HIV-associated or SLE associated disease—a possibility that can sometimes be confirmed by the electron microscopic finding of tubuloreticular inclusions (Midroni et al., 2000). False positive “suspicious” findings (vascular thrombosis, capillary proliferation, focal Wallerian degeneration, hemosiderin deposits) may occur (Collins et al.,

Fig. 3.4 Peripheral nerve vasculitis, paraffin embedded tissue. Figure A (H&E stained) shows an obliterated epineurial arteriole, densely infiltrated and surrounded by inflammatory cells, with prominent fibrinoid necrosis. In figure B, the epineurial arteriole shows recanalization after previous injury, and staining with Prussian Blue identifies hemosiderin-filled macrophages indicating prior perivascular hemorrhage.

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Fig. 3.5 Sural nerve biopsy specimen in cross-section, resin embedded tissue, stained with Toluidine Blue. In low power (A), the entire nerve cross-section is seen, with two fascicles showing severe recent injury, while several fascicles below show only loss of axons. Higher power examination of the severely affected fascicle (B) shows endoneurial hemorrhage. All visible axons are undergoing Wallerian degeneration.

2000). Some of these seem especially common in diabetics and elderly individuals in our experience, probably due to a combination of nonspecific perivascular cuffing and age- or diabetes-associated vessel degeneration. 3.6.3. CIDP and Guillain–Barré Syndrome 3.6.3.1. Histological findings The major site of pathological activity in CIDP may be at the nerve root, motor > sensory (Dyck et al., 1975), and hence cannot be fully assessed by a distal sensory nerve biopsy. Nevertheless, early literature in CIDP emphasized the utility of peripheral nerve biopsy in diagnosis, with cardinal features of ongoing demyelination with remyelination on teased fibers, and especially macrophage mediated-myelin stripping on electron microscopy (Dyck et al., 1975; Prineas and McLeod, 1976). The latter finding, where macrophage process are visualized in the process of penetrating and destroying myelin around an intact axon, is considered very suggestive of CIDP or Guillain–Barré syndrome, and rarely seen in other demyelinating conditions. In typical nerve biopsies the pattern of involvement is multifocal rather than diffuse—in an extreme case

an entirely normal nerve fascicle may be adjacent to one showing severe focal demyelination and/or axonal loss and/or onion bulb formation. Sometimes the endoneurial compartment appears “edematous,” with an amorphous substance accumulating predominantly in the subperineurial area. Inflammatory infiltration in peripheral nerves has been overemphasized as a feature in CIDP, occurring in only 10–50% of biopsies, and rarely impressive (Midroni and Bilbao, 1995; Bosboom et al., 1999; Bouchard et al., 1999). The cells tend to be found perivascularly, both in the epineurium and endoneurium. The infiltrating cells are predominantly T lymphocytes which immunostain positive for the pan-T Cell CD3 antigen. Plastic sections typically show combined features of axonal loss and active axonal degeneration, along with demyelinated axons, as well as thinly remyelinated axons (Fig. 3.3). Onion bulbs that consist of concentric lamellae of Schwann cell processes around a central axon, are seen in a significant minority of patients, variably prominent (Fig. 3.3B). If doubt remains as to the nature of a demyelinating process, electron microscopy is often useful in revealing whether the demyelination is due to primary macrophage-mediated myelin stripping (favoring

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CIDP or GBS) or another cause. Rarely, electron microscopy will show this feature when light microscopy does not show convincing demyelination, and such a finding becomes the only clue that the patient might have a treatable inflammatory neuropathy (Haq et al., 2000). Similarly, teased fibers are more sensitive for detection of demyelination than examination of cross-sections, and may show demyelination or remyelination that is overlooked by other methods. The nerve biopsy pathology of GBS is essentially the same as that of CIDP, except that the changes are more acute, on average showing more prominent demyelination and less chronic axonal loss. Macrophage-mediated demyelination is the hallmark, and an EM search will usually show this finding (Brechenmacher et al., 1987). In the “axonal” form of Guillain–Barré syndrome macrophage-mediated demyelination is not a feature, and lymphocyte infiltration also seems minor. Instead the process appears to involve a penetration of the peripheral nerve by macrophages via the node of Ranvier, axonal degeneration rather than demyelination being the primary process (Griffin et al., 1996). Such findings are not likely to be seen in a sural nerve specimen taken far from the site of most active disease (root and nerve terminal). We have, however, observed adaxonal macrophages in nerve biopsy material on rare occasions, and our impression is that this finding is associated with more severe axonal loss. This was reported by other workers long prior to the identification of “axonal” GBS (Brechenmacher et al., 1981). GBS is a clinical diagnosis, and biopsy does not impact on management unless it reveals an alternative diagnosis—vasculitis most commonly. Hence, there is no role for nerve biopsy in suspected GBS unless highly atypical clinical or laboratory findings are present. 3.6.3.2. Utility of nerve biopsy in CIDP Despite the ability of nerve biopsy to reveal suggestive or pathognomonic findings for this relatively common and treatable cause of neuropathy, in recent years the role of this procedure in CIDP has diminished substantially. Workers generally agree that nerve biopsy has little added value in in patients with a clinical and electrophysiological picture highly suggestive of CIDP (Molenaar et al., 1998; Bosboom et al., 2001). In a significant minority of cases with CIDP, the typical biopsy findings are not seen, so a negative biopsy does not rule out the diagnosis, even if axonal degeneration predominates (Dyck et al., 1975; Prineas and McLeod, 1976; Barohn et al., 1989). Inflammatory

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infiltration, even if present, is nonspecific (Bosboom et al., 1999). Particularly telling is the study of Bosboom et al. (2001) who studied 21 consecutive biopsies in patients meeting criteria for CIDP in comparison to 13 patients with a chronic idiopathic axonal neuropathy (CIAP). Techniques included the full spectrum of pathological investigations, including light microscopy of thick and semi-thin sections, teased fibers, and morphometry. There was very substantial overlap in the teased fiber findings of demyelination/remyelination, inflammatory infiltration, and onion bulb formation between the two patient groups. Two experienced neuropathologists were unable to make a consistent distinction between the CIDP and CIAP group by reviewing teased fiber material from the two groups. The real value of nerve biopsy in CIDP emerges when the diagnosis is not suspected on clinical or electrophysiological grounds. Several studies have documented and emphasized the ability of pathological assessment to suggest CIDP when electrodiagnostic testing fails to do so, or the clinical picture is atypical (Chia et al., 1996; Haq et al., 2000; Logigian et al., 1994; Vallat et al., 2003) and this has been our experience also. Fiber teasing and electron microscopy are two means of pursuing the search for primary demyelination, and it is not clear if one method is superior to the other—very likely the yield of either method depends on the quality of preparation and intensity of examination of the tissue. Haq et al. (2000) found that electron microscopy was somewhat more helpful in detecting primary demyelination and predicting response to treatment. Some studies have suggested that pathological features of very active demyelination in CIDP may predict a more severe course (Bouchard et al., 1999; Haq et al., 2000), but other have failed to confirm this (Bosboom et al., 2001). In our experience, biopsy features do not predict disease course nor correlate with severity. One would expect that motor axon loss would be the principal predictor of prognosis, a matter best assessed electrophysiologically and clinically, not with a few millimeters of sensory nerve from the distal lower limb. 3.6.4. Amyloidosis A search for amyloid remains one of the major indications for nerve biopsy. However, amyloid usually deposits diffusely throughout the body, and in many instances it may be possible to document this using a

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biopsy site with less morbidity than nerve biopsy (skin and fat, salivary gland, muscle, bone marrow, or even kidney). The sensitivity of nerve biopsy in amyloid neuropathy is not known, but may be surprisingly low. Simmons et al. (1993) found only two of eight biopsies positive in patients ultimately shown to have amyloid neuropathy and we certainly have had negative nerve biopsies in patients subsequently documented to have amyloid neuropathy. This underscores the need to look for other tissues to sample. If nerve is taken, a combined procedure with muscle is well advised (Rajani et al., 2000). In addition, readily available sequencing of the transthyretin gene will identify nearly all patients with familial amyloidosis. Amyloid is an extracellular proteinaceous substance, which appears amorphous by light microscopy and fibrillar under the electron microscope. It is composed of beta-pleated sheets forming matted linear non-branching fibrils 7–10 nm in diameter. All amyloid types have a common constituent—amyloid P protein. Amyloid types are distinguished by their “major protein,” quantitatively the dominant component of the deposit. In primary amyloidosis, the major protein is immunoglobulin light chain; in familial neuropathic amyloidosis the major protein is usually a mutant transthyretin protein, and much less often

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Gelsolin or ApoLipoprotein A1 (Midroni and Bilbao, 1995). Secondary amyloid, in which the major protein is serum Amyloid A protein, is associated with chronic inflammatory disease, and neuropathy is NOT a clinical feature, even though the protein may be found in nerve or muscle specimens. Senile amyloid contains transthyretin as the major protein, but is nonpathogenic, tending to deposit in small quantities in a perivascular pattern in the very old (>80). We have seen this falsely interpreted as an indicator of amyloid neuropathy on occasion. Neuropathy is often the presenting manifestation of systemic amyloidosis (Rajani et al., 2000). A paraprotein is not initially known to be present or may remain undetectable in a significant minority of patients with primary amyloid neuropathy, and a family history of neuropathy is not present in all patients with familial amyloid neuropathy. Hence, nerve biopsy may provide the only means of making the diagnosis when all tests have been exhausted. The cardinal finding is detection of amyloid deposits (Midroni and Bilbao, 1995). These are seen best on Congo Red or Thioflavin-stained specimens, and small deposits are seen more dramatically under polarized light, where they demonstrate apple-green yellow birefringence (Fig. 3.6). Some non-amyloid

Fig. 3.6 Amyloid neuropathy, paraffin-embedded material. With Congo Red staining (A) endoneurial amyloid deposits are readily seen, and these stand out even more when the specimen is examined under polarized light (B), showing apple-green yellow birefringence.

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materials (fibrin, elastin, mast cell granules) may stain with Congo Red, but do not display birefringence. The amyloid can be seen in any compartment— endoneurium, perineurium, and epineurium, as well as the epineurial fat. Vascular accumulation is common (Rajani et al., 2000). Occasional inflammatory cells may be seen, but this is never a prominently inflammatory process. Axonal loss is always present, with myelin changes minor or absent (Rajani et al., 2000). Clinically, amyloid neuropathy can be a small fiber neuropathy at onset, and this is sometimes reflected in the nerve biopsy, where small myelinated and unmyelinated fibers can be lost disproportionately to large myelinated fibers—the opposite of what happens in most neuropathic processes. This can be recognized subjectively through visual inspection by an experienced pathologist, and can be demonstrated more rigorously using a fiber diameter histogram (Section 3.5.2.4). If such a pattern is seen, even in the absence of amyloid, an extended search for amyloid should be carried out, and if this is unrevealing additional nonneural tissues can be considered for biopsy. If amyloid is detected immunostaining is very helpful, as commonly available immunostains can identify whether the amyloid major protein is transthyretin or immunoglobulin light chains. These immunostains, however, can be both falsely negative and falsely positive, and should be performed in conjunction with control tissue staining, and in step-sections with Congo Red staining. Immunostaining is generally more reliable for frozen tissue than for paraffin embedded material. Although electron microscopy in theory can show amyloid fibrils where light-microscopic techniques do not, when the deposits are very small or predominantly found in the subperineurial region, this should be interpreted with great caution, as any non-congophilic fibrillar material seen in the endoneurial and subperineurial region is more likely to be oxytalan, a part of the elastic fiber system (Midroni and Bilbao, 1995). 3.6.5. Leprosy While in parts of the undeveloped world leprous neuropathy is a continuing management problem, even in highly-developed countries one may encounter the occasional unexpected finding of leprous neuropathy (personal experience, Gabriel et al., 2000; Said, 2002a). Origin from an endemic area is the most significant clue to a possible diagnosis of leprous neuropathy. In a majority of instances symptom onset is within a year of infection. However, the neuropathy

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can be very indolent and patients may present up to several years after departure from a high-risk area, so a high index of suspicion is required. If skin lesions are present, these rather than nerve should be biopsied to confirm a diagnosis. However, in “pure neuritic leprosy,” there will be no clue to the diagnosis other than possibly the epidemiological risk factors of the patient (Jacob and Mathai, 1988). The pathological findings in leprous neuropathy depend on the immune response of the patient to the bacillus (Midroni and Bilbao, 1995). Regardless of the subtype, the peripheral nerve is usually obviously enlarged. When little or no immune response to the bacilli is mounted, the disease is termed pluribacillary (lepromatous) leprosy with large numbers of organisms throughout many tissues, but especially the peripheral nerves that may provide a relatively protected site from the immune response. The typical finding is accumulation of M. leprae organisms in macrophages and Schwann cells, forming “foamy cells,” staining positively with special stains such as Fite or Auramine Rhodamine (Fig. 3.7A). Lymphocytic and epithelioid cell infiltration is a relatively minor feature, and the nerve architecture is generally preserved. In early onset of disease, there is relatively minor nerve fiber loss, with mild degrees of demyelination/remyelination noted. As the disease progresses axons are lost and an increasing degree of endoneurial fibrosis emerges. Electron microscopy can provide very dramatic evidence of M. leprae accumulation in a variety of cell types, but most prominently in non-myelinating Schwann cells (Fig. 3.7B), for which they have a special tropism (Rambukkana et al., 2002). At the other end of the immunological spectrum is paucibacillary (tuberculoid) leprosy—here the immune response is intense, and organisms are rarely or never seen even with special stains. Epithelioid histiocytes and giant cell granuloma formation is the hallmark, often with central caseation, and the process can be very destructive to the neural architecture. Perineurial infiltration with histiocytes is prominent, which brings perineuritis and sarcoidosis into the differential diagnosis. In the intermediate (“borderline”) histological range, one sees an intense inflammatory response dominated by lymphocytes, with some organisms still detectable. There is a spectrum of pathology, with more prominent histiocytes and less prominent organisms earning the designation “borderline tuberculoid,” and more prominent organisms with few or no histiocytes earning the designation “borderline leproma-

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Fig. 3.7 Leprous neuropathy. Resin-embedded Toluidine Blue stained cross-section of the endoneurium shows several “globi”-macrophages filled with M. leprae organisms. Thinly myelinated axons are seen. Electron microscopy (B) reveals intact-appearing M. leprae organisms (dark circle within clear halo) predominantly within non-myelinating Schwann cells, as well as M. leprae debris in a macrophage.

tous.” Perineurial inflammation is also common in borderline leprosy. Vasculitis can be seen in leprosy, probably in response to large quantities of M. leprae antigen released into the nerve and the resultant immune response. This is often accompanied by a multisystem diffuse inflammatory process (Erythema nodosum leprosum). Even in known neuritic leprosy, there may be a role for nerve biopsy, as the nerve specimen may provide a more accurate reflection of disease activity than skin biopsy specimens and thus impact on selection of the antibacterial regime (Chimelli et al., 1977; Nilsen et al., 1989). If suspicion for leprosy is strong, but the biopsy diagnosis cannot be confirmed by conventional methods, the paraffin embedded tissue may be used for PCR. 3.6.6. Sarcoidosis Although often suspected and rarely proven, this condition remains an indication for nerve biopsy. A muscle specimen should be taken at the same time, or even in preference to a nerve specimen, since the yield of

muscle biopsy is high even if the dominant process is neuropathic (Said et al., 2002b). Clinical findings are nonspecific: multifocal or diffuse, non-length dependent or distally predominant and symmetrical, usually with axonal electrophysiology, but rarely with demyelinating neuropathy—even conduction block (Midroni and Bilbao, 1995; Said et al., 2002b). The histological hallmark is the presence of non-caseating granulomata, with multinucleate giant cells variably present. The granulomata may occur in the epineurial or endoneurial compartments. Necrotizing angiitis can be part of the histological picture (Said et al., 2002b). Inflammatory infiltration of the perineurium is a somewhat special feature of sarcoid neuropathy, not seen in the more common inflammatory neuropathies such as CIDP or vasculitis. If seen this finding should prompt a further search for sarcoidosis even if no granulomas are present. There may be a clinical and histological overlap with a less common syndrome called Sensory Perineuritis, which may constitute a distinct clinicopathological entity or may represent a focal form of sarcoidosis in some cases (Krendel and Costigan, 1992; Midroni and Bilbao, 1995). Neuritic

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leprosy should also always be considered in the differential diagnosis of prominent perineurial inflammation and intraneural granulomata. 3.6.7. Paraprotein-associated neuropathies A variety of peripheral neuropathies may be seen in patients who have a circulating paraprotein. The accepted associations are listed in Table 3.6. In general, with the exception of immunoglobulin deposition disease, which is exceedingly rare, and amyloidosis, the diagnosis is generally made by non-invasive means. However, a brief review of the expected pathology is provided. It should be noted that in most patients who have a peripheral neuropathy and a circulating paraprotein the association may be co-incidental (two conditions common in elderly patients), and no specific pathology is expected or found. Much attention has been focused on IgM anti-Myelin Associated Glycoprotein (anti-MAG) neuropathy, because the causal connection is most convincing, and clinical, electrophysiological, and biopsy findings most unique (Midroni and Bilbao, 1995). Histologically, this is a mixed axonal and demyelinating neuropathy, with prominent but indolent demyelination/remyelination seen on light microscopy. Onion bulb formations are seen routinely, although they are not as dramatic as those seen in CMT-1 or some cases of CIDP. Mild inflammatory infiltration is often present. In this neuropathy, there is an increased prominence of occasional hypermyelinated fibers with redundant myelin loops, sometimes reminiscent of what is seen in Hereditary Neuropathy with Pressure Palsies, but to a much lesser degree (Sander et al., 2000). The most pathognomonic finding in anti-MAG neuropathy is “widely spaced myelin” seen on elecTable 3.6 Spectrum of paraprotein-associated neuropathies ●

● ● ● ● ●

●

IgM anti-Myelin Associated Glycoprotein (antiMAG) paraproteinemic neuropathy Amyloid neuropathy Immunoglobulin deposition disease CIDP associated with an IgG or IgA paraprotein Cryoglobulinemic neuropathy POEMS syndrome (polyneuropathy associated with organomegaly, endocrinopathy, monoclonal gammopathy, and skin changes) Neuropathy in osteosclerotic myeloma (overlap with POEMS)

tron microscopy, which is due to penetration of IgM paraprotein into the myelin sheath, separating this tightly wound lipid bilayer at its extracelluar interface, the intraperiod line (Figs. 3.8A, 3.9A). IgM immunostaining of myelin is readily identified on frozen tissues (Fig. 3.9B), but is less specific than the EM change. Macrophage mediated demyelination can also be seen in this neuropathy (Vital et al., 2000). The POEMS syndrome features a nonspecific light microscopic appearance with variable elements of indolent axonal degeneration and demyelination/ remyelination. A somewhat unique ultrastructural finding is uncompacted myelin (Fig. 3.8B), whereby the intracellular apposition of the myelin sheath layers is disrupted, separating the major dense line into two lines. Although a very careful search may be required, some report that the sensitivity of this finding for the POEMS syndrome approaches 100% (Vital et al., 2003). However, uncompacted myelin is not diagnostic, as it can be seen in an occasional patients with CIDP with or without a paraprotein, rare patients with Hereditary Neuropathy with Pressure Palsies, and more consistently in genetically determined neuropathy due to a mutation in the P0 protein (Gabreels-Festen et al., 1996; Vital et al., 2003). A picture clinically indistinguishable from CIDP without a circulating paraprotein also occurs in patients with a paraprotein. The histology appears to be that of typical CIDP (Vital et al., 2000) with no special axonal or myelin findings, although immunolabeling sometimes shows that the paraprotein deposits in the endoneurium (Midroni and Bilbao, 1995). Immunoglobulin deposition disease is a rare condition where monoclonal immunoglobulin light and/or heavy chains deposit diffusely in tissue, without forming amyloid. Neuropathy can occur in this condition, due to massive accumulation of immunoglobulin in the nerve (Midroni and Bilbao, 1995). Cryoglobulinemic neuropathy can reveal a wide spectrum of findings (Midroni and Bilbao, 1995). This condition may be more common than suspected due to its association with Hepatitis C, and skin lesions may not always be present (Gemignami et al., 2002). In some cases, necrotizing vasculitis is predominant, and the process is usually predominant or exclusively axonal in nature. More often nonspecific inflammatory changes are detected, or a bland loss of axons noted with no suggestion of inflammation. There have been rare reports of cryglobulin precipitate obstructing endoneurial microvessels (Prior et al., 1992).

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Fig. 3.8 Paraproteinemic neuropathy, Electron microscopy. Widely spaced myelin (A) is seen in this patient with the antiMAG syndrome. The separation of myelin layers occurs at the Intraperiod Line (extracellularly). This finding is specific for the anti-MAG syndrome. In Figure B, uncompacted myelin is seen, with separation occurring at the Major Dense Line (intracellularly). This finding is relatively nonspecific, and occurred in a patient with a paraproteinemic neuropathy but no evidence of the POEMS syndrome.

3.6.8. Genetically determined neuropathies The greatest change in the role of nerve biopsy over the past decade has been the emergence of non-invasive genetic testing for diagnosis of a genetically determined neuropathy. Early nerve biopsy literature abounded with descriptions of genetic neuropathies such as Charcot–Marie–Tooth disease (CMT), Hereditary Neuropathy with Pressure Palsies (HNPP), or Friedrich’s Ataxia. In many patients nerve biopsy does reveal pathognomonic or highly suggestive findings (HNPP, CMT-1), and in others it is quite nonspecific (CMT-2, Friedreich’s Ataxia). However, since genetic testing will now identify the vast majority of CMT-1 and CMT-X patients, essentially all HNPP patients, as well as many spinocerebellar ataxias and many metabolic diseases, the role of nerve biopsy has become quite limited. As of this writing, blood tests are still not available for most cases of type II CMT, but the histology is quite nonspecific in any case. For this reason, the discussion below is quite abbreviated and for further details of nerve pathology the reader is referred to more comprehensive monographs (Midroni and Bilbao, 1995). 3.6.8.1. Charcot–Marie–Tooth disease Histological features of CMT-1 are mostly similar regardless of the underlying mutation. There is a loss

of myelinated fibers to varying degrees, with prominent findings of demyelination/remyelination and diffuse onion bulb formation. The latter result from recurrent events of demyelination with remyelination occurring over long periods of time, with gradual accumulation of supernumerary Schwann cell processes in a concentric pattern around the axon. The process is quite indolent, and hence active demyelination (denuded axons) is almost never seen in adults. However, in young patients with CMT-1, the prominence of visible ongoing demyelination may lead to concern about CIDP and the pathological distinction between the two may be impossible— marked focality of pathology and inflammatory infiltration would favor CIDP (Gabreels-Festen et al., 1992). An uncommon ultrastructural finding that suggests the diagnosis of the P0 mutation (CMT1B) is the presence of uncompacted myelin (Gabreels-Festen et al., 1996) (see discussion of POEMS syndrome, Section 3.6.7), but this can rarely be seen in CIDP (Vital et al., 2003). In type II CMT, regardless of genetic basis, the pathology is nonspecific. Indolent axon loss dominates, with little debris noted on the biopsy—just the reduced number of axons. Some secondary demyelination occurs, resulting in myelin thickness often being reduced relative to axonal diameter. However,

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Fig. 3.9 Anti-MAG paraproteinemic neuropathy. Electron microscopy with anti-IgM immunogold staining tracks the entry of pathogenic IgM paraprotein into the myelin, entering from the extracellular compartment. Light microscopic immunofluorescence also reveals dense accumulation of IgM around the axons.

active demyelination is not seen. Efforts at axonal regeneration produce variably prominent regenerating clusters, but this can occur in any chronic neuropathy, genetic or acquired. X-linked CMT is due to a mutation either in the connexin32 gene, or in related nearby non-expressed genetic elements which control the expression of this protein. Again, the pathology is nonspecific, but just as

the condition is often considered “intermediate” between CMT-1 and CMT-2, so too is its pathology (Sander et al., 1998; Vital et al., 2001). Classic onion bulbs are not very frequent, but “pseudo onion bulbs” are quite typical. In these formations, a regenerating cluster is centered by a myelinated fiber and surrounded by several unmyelinated axons with their associated Schwann cells. Axon numbers are often

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relatively preserved in CMT-X, as large myelinated fibers degenerate or atrophy, but regenerative clustering provides some numeric compensation (Sander et al., 1998; Vital et al., 2001). Again, as is typical for genetic processes the pathology appears quite indolent, with no active axonal loss (i.e., Wallerian degeneration) or active demyelination visible on cross-sections, although teased fibers may show some ongoing axonal degeneration. 3.6.8.2. Congenital dysmyelinating neuropathies In pediatric practice, another group of genetically determined demyelinating neuropathies is important, presenting either with congenital neurological abnormalities or with symptom onset in early childhood, typically characterized by severe slowing of nerve conduction velocities to <10 m/s (where measurable). In cases where the inheritance can be shown to be autosomal recessive, these patients are classified as CMT-4, but many workers still employ the historical term “Dejerine–Sottas Syndrome” (Plante-Bordeneuve and Said, 2002). Genetically and clinically this group of patients is highly heterogeneous, including mutations in genes which have been implicated in other forms of CMT (Tyson et al., 1997; Berger et al., 2002; PlanteBordeneuve and Said, 2002) and, similarly, the pathology is highly heterogeneous. A number of findings may be seen in a child presenting with severe early onset or congenital demyelinating neuropathy, and for now no satisfactory comprehensive histological or genetic classification exists. The findings that provide a clue to this diagnosis on a nerve biopsy done in a pediatric setting include: ● ● ●

●

●

●

●

Diffusely amyelinated axons Severely hypomyelinated axons Demyelinating neuropathy with active myelin debris removal and some evidence of remyelination Diffusely extremely thinly myelinated fibers within classic onion bulb formation (classic finding of Dejerine–Sottas Syndrome, several genetic loci) Hypomyelination with basal lamina onion bulbs (CMT-4A, 4C) Prominence of hypomyelinated fibers with “focally folded myelin” but no onion bulbs (CMT-4B, several genetic loci) Prominence of “unstable” (apparently degenerating) myelin

85

In addition, infantile onset CIDP does occur, and is a treatable condition that should not be overlooked (Plante-Bordeneuve et al., 2001). Clues here would be macrophage mediated demyelination and significant inflammatory infiltrate, but the absence of these does not rule out this diagnosis. 3.6.8.3. Hereditary Neuropathy with Pressure Palsies In Hereditary Neuropathy with Pressure Palsies (HNPP), the pathognomonic finding is focal myelin swellings, also called tomaculae, which are usually accompanied by evidence of demyelination and remyelination, small onion bulb formations, and little or no axon loss. Although usually the findings are quite dramatic on routine semi-thin sections, teased fibers best reveal the tomaculae, and rarely may provide the only clue of this diagnosis. However, myelin tomaculae occur, though usually to a less impressive extent, in other conditions including anti Myelin Associated Glycoprotein paraproteinemic neuropathy, CMT-1, CMT-4, and even CIDP (Vital et al., 1989; Sander et al., 2000). Genetic testing has, therefore, superseded histological diagnosis for diagnosis of HNPP. If the common 17p deletion is not found, a point mutation in the PMP-22 gene should be sought (Lenssen et al., 1998). The pathology in patients with a point mutation differs quantitatively from that of patients with the common deletion in that the former group have more severe axonal loss, more prominent onion bulb formation, and less prominent myelin tomaculae. This parallels the clinical observation that in some patients with the point mutation the clinical picture has similarities to CMT-1 (Lenssen, 1998). 3.6.8.4. Other genetically determined Neuropathies In spinocerebellar generations, including Friedreich’s Ataxia, a neuropathy is often present, and the histological picture is usually nonspecific, showing predominance of large fiber axonal loss, variable prominence of regenerating clusters, and secondary demyelination. Giant Axonal Neuropathy is a disease of pediatric interest in which nerve biopsy may reveal pathognomonic findings. There is severe axonal loss with dramatic axonal swelling due to massive neurofilament accumulations. The condition is stereotyped clinically and the gene defect known to be in the Gigaxonin gene (Bomont et al., 2003), so likely genetic testing will eliminate the need for nerve biopsy in this setting. Nerve biopsy should usually not be the pathway to diagnosis of any of the lipid storage disorders, as biochemical or genetic testing is available for all of these

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diseases. However, occasionally, the diagnosis is unsuspected due to an atypical presentation, especially late onset storage disorders with clinically isolated neuropathy. In particular, metachromatic leukodystrophy (Felice et al., 2000), Fabry’s disease (Fig. 3.10), Globoid cell leukodystrophy (Sabatelli et al., 2002), Tangier disease, and adrenomyeloneuropathy may be unsuspected diagnoses that nerve biopsy reveals. The interested reader is referred to a more detailed exposition (Midroni and Bilbao, 1995). 3.6.9. Diabetes The biopsy findings of typical small fiber and/or sensorimotor distally predominant symmetrical diabetic neuropathy are generally nonspecific: axonal loss, variable evidence of demyelination/remyelination, hyalinization of blood vessels, no or slight inflammatory infiltration, and slight vascular or perineurial injury. Demyelinating features do not predominate in this setting, but a rare demyelinated axon and even small onion bulb may be seen. Disproportionate small fiber involvement can also occur, and parallels the occurrence of small fiber neuropathy in diabetics,

Fig. 3.10 Fabry’s disease. Resin-embedded Toluidine Blue stained cross-section shows accumulation of stored material in perineurial cells. The axon population appears intact, as normally there is little or no myelinated axon loss in this small fiber neuropathy.

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especially in the early stages of the condition (Midroni and Bilbao, 1995; Sumner et al., 2003). Diabetic patients can also present with a multifocal neuropathy, proximally predominant in the lower extremities, with elements of plexopathy or polyradiculopathy. This condition typically evolves subacutely and is initially dominated by pain. In this setting, one may find biopsy evidence of ischemic injury to nerve, including focal axonal loss, perineurial injury, inflammatory vasculopathy, and even frank necrotizing vasculitis. This may be seen either on the biopsy of proximal nerve segments or less often the sural nerve (Dyck et al., 1999; Said et al., 2003). Nerve biopsy should not be performed to separate between diabetes and CIDP—the histological distinction is poor, and clinical and electrophysiological criteria more useful (Uncini et al., 1999). 3.6.10. Nerve biopsy in other conditions 3.6.10.1. Peripheral neuropathy in patients with systemic malignancy Nerve biopsy is rarely useful in a patient presenting with neuropathy in the context of malignancy. Histological features of paraneoplastic neuropathy, whether subacute sensory neuropathy or the more common length-dependent varieties, are nonspecific and cannot be distinguished from features of druginduced neuropathy. In some instances inflammatory infiltration can be seen, but does not reach the degree sufficient to diagnose necrotizing vasculitis. Nerve biopsy is very helpful in those extremely rare instances of malignancy which infiltrates the nerves directly. Lymphoma is the most common cause, myeloma and leukemia being extremely rare alternatives. Only about half of such patients are known to have systemic malignancy at the time of biopsy diagnosis, and occasionally the peripheral nerve lymphoma may represent the initial manifestation of disease recurrence (Midroni and Bilbao, 1995). Non-Hodgkin’s lymphoma is far more likely to manifest in this manner, but no particular subtype seems more likely to cause neural involvement. A diffuse massive infiltration is the most common finding, and epineurium, perineurium, or endoneurial may be most affected. Subtle infiltration can occur, and this mandates a thorough examination of as many sections as reasonably possible. At times the neoplastic infiltrate is strikingly angiocentric. The diagnosis of neoplastic neuropathy is suggested by detection of atypical cells, marked cellular pleomorphism, and rarely mitotic

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figures. However, when a well-differentiated lymphoma infiltrates nerves it can be very difficult to distinguish from benign perivascular inflammation. In an HIV-positive patient, a distinction from diffuse infiltrative lymphocytosis syndrome needs to be made (Section 3.6.10.2). In endovascular lymphoma (angiotropic T cell lymphoma), the malignant cells disseminate widely throughout the body affecting multiple organs through microvascular obstruction—rarely they may cause peripheral neuropathy and nerve biopsy may disclose the neoplastic cells obstructing endoneurial and epineurial microvessels (Fig. 3.11). 3.6.10.2. Peripheral neuropathy in HIV-positive patients The commonest neuropathies seen in HIV-positive patients are length-dependent sensory predominant neuropathies, either due to HIV infection directly, or due to side-effects of medications required to combat the virus. In either case, the neuropathy has a nonspecific nature, with relatively mild axonal loss, and no significant inflammatory infiltration. Rarely, a more aggressive neuropathy can occur due to vasculitis in HIV-positive patients. It can affect epineurial, and more uniquely endoneurial (Midroni

Fig. 3.11 Intravascular lymphoma. Paraffin embedded H&E stained section shows atypical mononuclear cells occluding endoneurial microvessels.

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et al., 2000), vessels. CIDP and GBS occurring in the context of HIV infection do not have any unusual histological features which separate them from non-HIV associated cases. Very rarely, and in the latest stages of HIV illness when the host is maximally immune suppressed, cytomegalovirus can cause a polyradiculoneuropathy. The pathological features are severe axonal neuropathy with prominent inflammatory infiltration, sometimes necrotizing vasculitis, and rarely cytomegalic inclusions detectable in endothelial cells (Midroni and Bilbao, 1995). Patients with diffuse infiltrative lymphocytosis syndrome (DILS) manifest with multiorgan lymphocytic infiltration, especially affecting the salivary glands, but commonly also with peripheral neuropathy that is painful, diffuse or multifocal. Biopsy in these patients reveals an intense angiocentric lymphocytic infiltrate, which might be mistaken for well-differentiated lymphoma, but actually consists of a polyclonal lymphoid population (Gherardi et al., 1998). Regardless of the mechanism of underlying neuropathy, a common pathological feature in peripheral nerves (and indeed any tissue) of HIV-positive patients is tubuloreticular aggregates. These are distinctive reticular aggregates of irregularly branched and anastamosing tubules 20–30 nm in diameter most often seen in endothelial cells, easiest to locate just outside the nuclear envelope. They are somewhat nonspecific, as they represent a cellular response to high levels of circulating interferons, and have also been reported in neuropathy associated with SLE, as well as in a wide variety of non-neural tissues (Midroni et al., 2000). 3.6.10.3. Toxic neuropathy Nearly all toxic exposures, whether pharmaceutical, environmental, or industrial that cause peripheral neuropathy, produce a nonspecific clinical picture of distal axonopathy, and a correspondingly non-specific histological picture showing axonal neuropathy, of varying severity. There are only rare exceptions to this. Among pharmaceutical agents Chloroquin, Amiodarone, Perhexilene, and Disulfiram are the only causes of a toxic neuropathy (often neuromyopathy) with characteristic histological features. The first three of these pharmaceutical agents interfere with lysosomal metabolism causing cellular accumulation of lipid substance. All organs are affected, and the peripheral nerves will reveal this finding readily, with crystalline, lamellar, or granuloreticular accumulations inside lysosomes of any cell, but most readily seen in endothelial cells. Axonal loss and varying degrees of

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demyelination can occur. Note that any patient taking these drugs may accumulate these lysosomal changes, even if asymptomatic, and hence a nerve biopsy revealing typical lysosomal inclusions does not prove that a patient’s neuropathy is due to the drug—if there are other plausible alternatives they must also be considered. Disulfiram is a known cause of peripheral neuropathy and in a significant minority of patients will cause a neuropathy characterized by accumulation of neurofilaments in axons. The more common picture is nonspecific axonal loss. Toxic exposure to hexacarbons (n-Hexane, methyl n-butyl ketone) also causes a neuropathy, which has been characterized variably as an axonal or demyelinating process. The clinical picture can even mimic CIDP. The pathognomonic ultrastructural finding is axonal swelling due to neurofilament accumulation.

et al., 1994), and this has also been our experience (Table 3.7). However, the significance and specificity of biopsy evidence of very mild neuropathy in a patient with normal nerve conduction studies is uncertain, given the wide range of normal and especially age related “abnormalities” in asymptomatic individuals (Tohgi et al., 1977; Jacobs and Love, 1985). It is also possible for nerve biopsy to be normal when conduction studies are not, presumably because the pathological process is not present in the distal sensory segment biopsied, but rather in the motor fibers, typically more proximally (Logigian et al., 1994). However, it has been our experience that most often a negative nerve biopsy in the setting of abnormal electrodiagnostic testing means that the process is not a peripheral neuropathy—the commonest alternative diagnoses being spinal stenosis and myelopathy.

3.7. Nerve biopsy—electrophysiological correlation

3.7.2. Nerve biopsy and EDX disagree in a significant minority of cases

A question of particular interest to neurophysiologists is how well nerve biopsy and electrophysiological findings of demyelination vs axonal degeneration correlate, and when the findings are discordant, who is “right”? In general the correlation is good (60–80%, discussed below), but in a significant minority of instances there is discordance. This is as likely due to a failure of electrophysiology to identify the relevant process as to a failure of biopsy to sample a segment of nerve in which the relevant pathological process is active.

In our personal series of 207 consecutive biopsies with electrophysiological correlation, concordance was seen in 71% (Table 3.7). Of the 65 biopsies with electrophysiological evidence of primary demyelination by standard EDX criteria biopsy was interpreted as showing an axonal neuropathy in 19 instances (29%). Of 109 patients interpreted as having an axonal neuropathy electrically, the evidence of primary demyelination was seen on biopsy in 20 (28%). These results parallel those of Logigian et al. (1994). Two recent studies of CIDP patients confirm the not-infrequent failure of clinical, electrophysiological, and histological correlation, yet come to opposing conclusions about which technique is more reliable. Bosboom et al. (2001) found that pathology frequently failed to support clinically and electrophysiologically clear-cut diagnoses of CIDP, and concluded that

3.7.1. Nerve biopsy is more sensitive for mild neuropathy than EDX Nerve biopsy has consistently been found to be a more sensitive means of detecting neuropathy than electrodiagnostic studies (Behse and Buchthal, 1978; Logigian Table 3.7 Biopsy/EDX correlation (N = 207)

Electrodiagnosis

Biopsy

Totals

Axonal Demyelinating/Mixed Normal

Axonal

Demyelinating/mixed

Normal

86 20 3 109

19 45 1 65

14 3 16 32

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biopsy was not particularly useful in this setting. In contrast, others (Haq et al., 2000; Vallat et al., 2003) emphasized the ability of nerve biopsy to detect demyelinating disease not apparent electrophysiologically, and predict treatment responsiveness, concluding that histology was the “gold standard” more than electrophysiology. 3.7.3. Reasons for electrodiagnostic—Histological discordance 3.7.3.1. Biopsy failure—Sampling limitations In processes tending to be active proximally, and/or favoring motor nerve fiber involvement, such as some cases of CIDP and all cases of multifocal motor neuropathy, distal sensory nerve biopsy is at an obvious disadvantage. Moreover, in processes that are multifocal by nature, a several mm segment of nerve may altogether miss the relevant pathology by random chance—even if the process is not confined proximally or to motor nerves. There is an inherent bias for nerve biopsy to detect axonal degeneration preferentially to demyelination. This occurs because when a sensory nerve has undergone axonal degeneration anywhere distal to the spinal ganglion fiber loss and/or Wallerian degeneration will be visible all along the segment of the nerve distal to the site of injury. In contrast, demyelination is identified only if it is active precisely at the several millimeter segment examined histologically. 3.7.3.2. Biopsy failure—Secondary demyelination misinterpreted as primary A number of studies have shown that chronic axonal dysfunction can induce secondary demyelination along the unhealthy atrophic axon (Dyck et al., 1981a). However, the axon remains alive and remyelinates, with resultant thin myelin, relatively short internodes, and even rare onion bulbs. This can create the false impression of primary demyelination. 3.7.3.3. EDX failure—Axonal neuropathy can cause conduction velocity slowing Electromyographers are familiar with the many ways in which slowing of conduction velocity may be seen in the absence of a diffuse demyelinating condition (Logigian et al., 1994). These include: (i) secondary demyelination; (ii) slowed conduction velocity along thinly myelinated regenerating axons; (iii) loss of rapidly conducting large myelinated fibers; (iv) conduction block in the fastest conducting fibers; and (v)

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retrograde axonal atrophy from sites of focal compression. Hence, a process that is fundamentally axonal in nature can produce nerve conduction studies that can suggest primary demyelination, as occurred historically in the early reports of uremic neuropathy or diabetic neuropathy (Dyck et al., 1981a). Modern efforts to create an electrodiagnostic definition of “demyelinating” neuropathy are clinically useful, but specificity and sensitivity remain uncertain (Bromberg, 1991; Haq et al., 2000). 3.7.3.4. EDX failure—EDX does not provide information about all nerve fibers The velocities and latencies reported in routine nerve conduction studies are those of the fastest-conducting fibers, and if a significant proportion of such fibers are spared in a demyelinating process, the reported conduction velocities will not fall into the demyelinating range. Similarly, the dominant contributor to any recorded sensory potential is the volley of activity arising from large diameter fast conducting Aα fibers. Little or no information is derivable about unmyelinated or small myelinated fibers (Dyck, 1990), which means that amplitude and conduction velocity provide no information about the majority of fibers in the sensory nerve. Near-nerve studies partially obviate this problem, but are not commonly performed. 3.7.3.5. EDX failure—EDX provides very limited information about sensory axons An effort to directly relate sural sensory nerve amplitude (recorded cutaneously) with sural fiber density measured histologically showed that a drop of 150 myelinated nerve fibers per mm2 correlated with a 1 μV reduction in the sural (Russell et al., 1996). Ironically, in this work, the poorest correlation between sural nerve fiber density and any clinical or electrophysiological measure was with the actual sural sensory amplitude. Significantly better correlation was seen for nerve fiber density against any of: (i) sum of muscle strength testing scores (NDS); (ii) sum of motor Compound Muscle Action Potentials; (iii) sum of Sensory Nerve Action Potentials (SNAP); (iv) vibration detection threshhold; and (v) current perception threshold. Electrodiagnostic examination fails even more dramatically when a demyelinating process affects sensory axons. Sensory responses quickly trend to “non-recordable” in the setting of sensory fiber demyelination because of the low amplitude and short duration of the individual responses, magnifying

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effects of phase cancellation. Histological studies may reveal largely intact nerve fiber density in patients with absent sensory responses because of this dispersion and phase cancellation effect (Bouchard et al., 1999). Hence, in a sensory dominant demyelinating neuropathy, electrodiagnosis of demyelination is nearly impossible, but biopsy is ideally suited to the task (Vallat, 2003). While near nerve methods can partially address this problem, they are time consuming, not routinely practiced, and all widely accepted criteria for electrophysiological identification of a demyelinating neuropathy focus upon motor nerve conduction results, not sensory results.

are invariably those in which the pathologist and the neurologist have collaborated, exchanging their knowledge about what is suspected and is reasonable clinically, and what the biopsy may or may not be able to reveal about the patient’s condition.

3.7.3.6. Biopsy and EDX failure—A wide range of normal Both clinical neurophysiologists and neuropathologists are aware of the wide range of normal inherent in their diagnostic method. In the EMG lab, a patient may have sensory and motor amplitudes drop by 50% or more, and still be within the “normal range.” Conduction velocity lower limits of normal are a bit more robust, but still patients can have a significant drop from their baseline and remain within the normal range. In the pathology lab, a typical normal range of sural nerve fiber density is 5000–10 000 myelinated fibers per mm2, and hence for a patient starting out in the high part of this range significant drop can occur before one is certain that there has been loss. With age, the range of normal only becomes broader, amplifying the problem. However, nerve biopsy has the advantage here, in the sense that seeing even a few actively degenerating or demyelinating axons is abnormal, and identifies a neuropathic process. In contrast, EDX assesses only the remaining healthy nerve, leaving no means for mild pathology to express itself. This, presumably, is the main reason why nerve biopsy is more sensitive than electrodiagnosis at detecting mild neuropathy (Section 3.7.1 above).

References

3.8. The complementary nature of nerve biopsy and electrodiagnostic testing We hope that the reader of this chapter has come to the conclusion that EDX and nerve biopsy are diagnostic procedures with unique strengths and weaknesses. The information provided to the clinician by these methods is complementary, and needs to be interpreted in a clinical context with an understanding of where each method is strongest, and where the pitfalls may lie for either technique. The most helpful biopsies

Acknowledgement For over two decades, Sandra Cohen has been providing outstanding technical support in the Pathology Laboratory at St. Michael’s Hospital. This chapter and much of the other work we have done over the years would not have been possible without her.

Argov, Z, Steiner, I and Soffer, D (1989) The yield of sural nerve biopsy in the evaluation of peripheral neuropathies. Acta Neurol. Scand., 79: 243–245. Barohn, RJ, Kissel, JT, Warmolts, JR and Mendel, JR (1989) Chronic inflammatory demyelinating polyradiculoneuropathy. Clinical characteristics, course, and recommendations for diagnostic criteria. Arch. Neurol., 46: 878–884. Behse, F and Buchthal, F (1978) Sensory action potentials and biopsy of the sural nerve in neuropathy. Brain, 101: 473–493. Berger, P, Young, P and Suter, U (2002) Molecular cell biology of Charcot–Marie–Tooth disease. Neurogenetics, 4: 1–15. Bomont, P, Ioos, C, Yalcinkaya, C, Korinthenberg, R, Vallat, JM, Assami, S, Munnich, A, Chabrol, B, Kurlemann, G, Tazir, M and Koenig, M (2003) Identification of seven novel mutations in the GAN gene. Human Mutation, 21: 446. Bosboom, WMJ, Van den Berg, LH, De Boer, L, Van Son, MJ, Veldman, H, Franssen, H, Logtenberg, T and Wokke, JHJ (1999) The diagnostic value of sural nerve T cells in chronic inflammatory demyelinating polyneuropathy. Neurology, 53: 837–845. Bosboom, WMJ, van den Berg, LH, Franssen, H, Giesbergen, PCLM, Flach, HZ, van Putten, AM, Veldman, H and Wokke, JHJ (2001) Diagnostic value of sural nerve biopsy in chronic inflammatory demyelinating polyneuropathy. Brain, 124: 2427–2438. Bouchard, C, Lacroix, C, Plante, V, Adams, D, Chedru, F, Guglielmi, JM and Said, G (1999) Clinicopathologic findings and prognosis of chronic inflammatory demyelinating polyneuropathy. Neurology, 52:498–503.

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Brechenmacher, C, Vital, C, Laurentjoye, L and Castaing, Y (1981) Ultrastructural study of peripheral nerve in Guillain–Barré sydnrome: presence of mononuclear cells in axons. Acta Neuropathol. Suppl, VII, 249–251. Brechenmacher, C, Vital, C, Deminiere, C, Castaing, Y and Gbikpi-Benissan, G (1987) Guillain–Barré syndrome: an ultrastructural study of peripheral nerves in 65 patients. Clin. Neuropathol., 6: 19–24. Bromberg, MB (1991) Comparison of electrodiagnostic criteria for primary demyelination in chronic polyneuropathy. Muscle Nerve, 14: 968–976. Chia, L, Fernandez, A, Lacroix, C, Adams, D, Plante, V and Said, G (1996) Contribution of nerve biopsy findings to the diagnosis of disabling neuropathy in the elderly. A retospective review of 100 consecutive patients. Brain, 119: 1091–1098. Chimelli, L, Freitas, M and Nascimento, O (1977) Value of nerve biopsy in the diagnosis and followup of leprosy: the role of vascular lesions and usefulness of nerve studies in the detection of persistent bacilli. J. Neurol., 244: 316–323. Collins, MP, Mendell, JR, Periquet, MI, Sahenk, Z, Amato, AA, Gronseth, GS, Barohn, RJ, Jackson, CE and Kissel, JT (2000) Superficial peroneal nerve/peroneus brevis muscle biopsy in vasculitic neuropathy. Neurology, 55: 636–643. Corbo, M, Kamel Abouzahr, MK and Latov, N, et al. (1997) Motor nerve biopsy studies in motor neuropathy and motor neuron disease. Muscle Nerve, 20: 15–21. Davies, L, Spies, JM, Pollard ,JD and McLeod, JG (1996) Vasculitis confined to peripheral nerves. Brain, 119: 1441–1448. Deprez, M, Ceutrick-de Groote, C, Gollogly, L, Reznik, M and Martin, JJ (2000) Clinical and neuropathological parameters affecting the diagnostic yield of nerve biopsy. Neuromuscul. Disord., 10: 92–98. Dyck, PJ and Lofgren EP (1966) Method of fascicular biopsy of human peripheral nerve for electrophyiologic and histologic study. Mayo. Clin. Proc., 41: 778–784. Dyck, PJ and Lofgren, EP (1968) Nerve biopsy. Choice of nerve, method, symptoms, and usefulness Med. Clin. North. Am., 52: 885–893. Dyck, PJ, Lais, AC, Ohta, M, Bastron, JA, Okazaki, H and Groover, RV (1975) Chronic Inflammatory Polyradiculoneuropathy. Mayo. Clin. Proc., 50: 621–637. Dyck, PJ, Lais, AC, Karnes, JL, Sparks, M, Hunder, H, Low, PA and Windebank, AJ (1981a) Permanent

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Peripheral Nerve Diseases Handbook of Clinical Neurophysiology, Vol. 7 J. Kimura (Ed.) © 2006 Elsevier B.V. All rights reserved

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CHAPTER 4

Pathophysiology of demyelination and axonal degeneration William F. Browna, Anh X. Nguyenb and Bradley V. Watsonc a McMaster University Medical Center, Hamilton, Ontario, Canada London Health Sciences Centre, EMG Lab, London, Ontario, Canada c CentraCare Clinic, Saint Cloud, MN, USA

b

4.1. Introduction The peripheral nervous system (PNS) serves as the two-way communication link between the central nervous system (CNS) and the periphery. The signals take the form of transient electrical all-or-nothing “action potentials,” which briefly reverse the axolemmal resting transmembrane potential by as much as 120–130 mV and are transmitted in humans, at speeds which range from little as 1 m/s or less to as much as 70 m/s. The speed of transmission depends on the diameter of the nerve fiber and whether the nerve fiber is myelinated or not. For most of the PNS, Schwann cells maintain the myelin sheath of myelinated nerve fibers. However for those components that lie within the CNS, including the intramedullary components of the motor axons, autonomic axons and the central processes of the dorsal root ganglion cells, the myelin sheath is maintained by oligodendroglial cells. Both the Schwann cells and oligodendroglial cells are responsible for the development and maintenance of their respective myelin sheaths. 4.2. Cable properties of nerve fibers Conduction in long cylindrical conductors such as nerve fibers is influenced by the “cable” properties of the fiber (Ruch et al., 1965; Nicholls et al., 2000; Brown, 2002). For example, a depolarizing stimulus current, applied at one point to the nerve fiber, enters the nerve fiber at that point, spreads longitudinally down the core of the axon and leaks outward through the membrane to return via

* Correspondence to: William F. Brown, McMaster University Medical Center, 1200 Main St West, Hamilton, Ontario, Canada L8S 2J9. E-mail address: [email protected] Tel.: +905-468-8949; fax: +905-468-8949.

the extracellular space to the point of stimulation. How far the internal longitudinal current (ILC) extends beyond the site of stimulation depends on: (1) the magnitude and duration of the applied stimulating current; (2) the capacitance and resistance of the membrane; and (3) the internal longitudinal resistance (ILR). The foregoing influences on the cable properties are especially important to understand the changes in conduction velocity and the safety factor for conduction observed in demyelinated nerve fibers. 4.3. Conduction in normal myelinated nerve fibers The evidence is excellent that conduction in normal myelinated nerve is saltatory. Active membrane ionic currents are limited to the nodal membrane. A.demonstration of the current pattern in myelinated nerve fibers by Tasaki and Takeuchi (1942), Tasaki (1953), and later Huxley and Stampfli (1949a, 1949b), along with the evidence reviewed by Rogart and Stampfli (1982). Huxley and Stampfli were able to demonstrate in single myelinated nerve fibers that: (1) The transverse (radial) membrane current in the internodal segments was outward and passive; and (2) The active inward transverse (radial) current was confined to the node of Ranvier. The magnitude of the outward transverse membrane current diminishes as the distance increased from the preceding active node of Ranvier; and (3) The latency of the external longitudinal current (ELC) abruptly increased at the beginning of successive internode segments. This latency “step” reflects: (1) the time required for the outward transverse membrane current driven by the action potential in preceding nodes, to depolarize the next node to threshold and generate a

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new action potential; and (2) the distributed capacitance of the internode that progressively delays the transverse membrane current as the impulse traverses the internode. The latency steps between inward currents at successive nodes of Ranvier vary from 70 μs in the cat to 25 μs in the rat. The original method pioneered by Huxley and Stampfli was later modified by Rasminsky and Sears (1972) and Bostock and Sears (1978) to investigate conduction in single myelinated nerve fibers within an intact multifiber spinal root preparation in the rat. Important additional points from these latter investigations, with respect to saltatory conduction in myelinated nerve fibers include: (1) Although conduction in myelinated nerve fibers is saltatory in the sense that the active inward Na+ currents are confined to the nodes of Ranvier, the accompanying action potential is continuously conducted across successive internodes as a steadily advancing wave of depolarization and repolarization. (2) The action potential occupies space as well as time. For example, at any one time, the action potential may extend over as many as 10–30 successive internodes. The wavelength of the action potential may be calculated by multiplying the duration of the spike in milliseconds by the conduction velocity in m/s. For example, the wavelength for a nerve fiber conducting at 60 m/s, would equal 60 × 0.5 − 1.0 or approximately 30–60 mm. Therefore, the evidence is strong that generation of the action potential is confined to successive nodes of Ranvier. In myelinated nerve fibers, the density of Na+ channels is lower in the internode (less than 25/μm2,) compared to the node of Ranvier (1000/μm2 see Ritchie, 1979, 1995; Arroyo and Scherer, 2000; Peles and Salzer, 2000). Clustering of the Na+ channels in the nodal region is probably greatly influenced by specific contacts with Schwann cells (Joe and Angelides, 1992, 1993; Novakovic et al., 1996). The tendency for Na+ channels to cluster together might also reflect fundamental properties of certain types of axons (Johnston et al., 1996; Vabnick and Shrager, 1998). Few, if any, K+ channels are found in the nodal region, while a variety of K+ channels can be found in the internodal segment, mostly in the paranodal region. In the absence of a fast-rectifying K+ current

in the nodal region, excitability is rapidly restored in the nodal region through a combination of rapid Na+ channel inactivation and the presence of a relatively large non-voltage-dependent leakage current. The two chief factors that influence conduction velocities in myelinated nerve fibers include the fiber diameter (thickness of the myelin sheath), and the internode length. Myelinated nerve fibers conduct at speeds roughly proportional to their diameters, whereas unmyelinated nerve fibers conduction velocity is proportional to the square root of the diameter of the nerve fiber. Increasing the thickness of the myelin sheath at the expense of the axon increases the ILR and slows the impulse. On the other hand, increasing the diameter of the axon at the expense of the myelin sheath increases the capacity of the internode, and delays conduction by increasing the capacitive load on the ILC. It turns out that the optimal ratio of axonal diameter to overall nerve fiber diameter is approximately 0.6 (Rushton, 1951). 4.4. Conduction in demyelinated nerve fibers Primary or secondary demyelination is one of the most common morphological features of many peripheral and central nervous system diseases. In the PNS, primary demyelination stems from direct injury to the Schwann cell or the myelin sheath. Secondary demyelination, on the other hand, usually results from a primary injury to the neuron or its axon and tends to begin at the distal extremities of affected nerve fibers. The distribution of primary demyelination is usually far more variable. In the primary and secondary varieties, myelin retraction and breakdown usually begins in the nodal region leading to paranodal demyelination. Later, the demyelination may extend throughout the entire internode segment producing segmental demyelination. In response to the breakdown of myelin, surviving Schwann cells proliferate. In primary demyelination, remyelinated internodes tend to be shorter than normal segment and vary widely in length. This stands in contrast to remyelination of regenerated nerve fibers where the reconstituted internodes tend to shorter but more uniform in length. In some chronic demyelinating disorders, repeated bouts of demyelination and remyelination may lead to onion bulb-like thickenings of the peripheral nerves with the result that the nerve becomes firmer and thickened to palpation.

PATHOPHYSIOLOGY OF DEMYELINATION AND AXONAL DEGENERATION

4.5. The physiological consequences of demyelination and remyelination The physiological consequences of demyelination depend on the extent and severity of the demyelination as well as the types of nerve fibers involved. Demyelination impairs the transmission of impulses by changing the biophysical properties of the paranodal and internodal membranes and has been studied in single nerve fibers by Bostock and Sears (1978). For example, loss of the myelin sheath increases the capacitance and diminishes the transverse resistance in the paranodal and internodal regions. These changes increase the load on the available and finite ILC generated by the preceding “active” node of Ranvier. The ILC generated by the latter must reach the next “inactive”, node of Ranvier in sufficient quantity to bring the region to threshold and generate an action potential if conduction is to continue along the nerve fiber. If the drain on the ILC imposed by the leakage current in the demyelinated regions is too large, insufficient current may be available to depolarize the next node of Ranvier to threshold with the result is that the impulse blocks and further transmission of the action potential comes to an end. Demyelination in the paranodal region is especially critical for three reasons. Firstly, retraction of the myelin sheath and Schwann cell processes increases the available nodal membrane area, thereby diluting the available depolarizing current over a larger membrane surface (Koles and Rasminsky, 1972; Rasminsky and Sears, 1972; Bostock and Sears, 1978). Secondly, potassium channels that are normally present in the nearby paranodal-axolemmal membrane become exposed and as a result the membrane becomes hyperpolarized, thereby reducing the safety factor for transmission in the region (Bostock et al., 1981; Chiu and Ritchie, 1981; Bostock and Rasminsky, 1983). Thirdly, the Na+, K+-ATPase electrogenic pump in the nodal region tends to drive the membrane toward the K+ equilibrium potential, further increasing the load on the ILC available for generating an action potential at the next nodal region in line. 4.5.1. Conduction block Conduction may be blocked in several ways. For example, reducing the ILC generated from the preceding active node of Ranvier to below that required for the next in-line node of Ranvier to reach threshold will block conduction. Alternatively, any increase in the load on the available ILC required for generating an

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action potential at the next node of Ranvier, will also block conduction. Both basic mechanisms are summarized in Table 4.1. Diseases, which destroy or functionally interfere with the sodium channels, may reduce the total inward sodium current and reduce the available ILC below what is necessary to generate an action potential at the next in-line node of Ranvier. Putative Na channel blocking factors (Brinkmeier et al., 1992a; Takigawa et al., 1995) and other neuroelectric blocking compounds such as interleukin-2 (Brinkmeier et al., 1992b) or nitric oxide (Redford et al., 1997; Shrager et al., 1998; Renganathan et al., 1999) may block conduction in myelinated nerve fibers even in the absence of demyelination. The security of transmission of the impulse may also be reduced at sites where the available ILC is faced with a greatly expanded surface membrane, which imposes an additional load on the available current to the point where the safety factor for transmission may be Table 4.1 Mechanisms for blocking conduction Reduce the available ILC required for activating the next node of Ranvier ●

● ●

● ● ● ●

Reduce the number of sodium channels in the nodal region Block the sodium channel with blocking-antibodies Bind the sodium channels by toxins such as saxitoxin or tetrotoxin Reduce the opening times of sodium channels Increase the temperature of the nerve Expose the potassium channels in the paranodal region Change the kinetics of the sodium channels*

Increase the ‘load’ on the ILC required to activate the next node of Ranvier ● ●

● ● ●

● ●

*

Increase the ILR of the axoplasm Hyperpolarization of the nodal region through the action of the Na+, K+-ATPase, electrogenic pump in the nodal region, especially in response to high frequency trains of impulses Increase the specific resistivity of the axoplasm* Reduce the diameter of the axon Increase the passive, transverse leakage currents through the membrane Paranodal demyelination Internodal demyelination

Denotes possibilities for which there is no direct experimental evidence.

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jeopardized. Examples of such impedance mismatches (Waxman, 1977, 1978, 1980, 1995) in normal nerve include: (1) the large unmyelinated initial segments of motor neurons; (2) branch points in myelinated nerve fibers where the branches impose extra loads on the available ILC originating at or proximal to the branch points; and (3) transition sites between the proximal myelinated and the terminal unmyelinated segments of nerve fibers. The normally lower safety factor for transmission of impulses at the above sites makes such transmission at these sites more susceptible to conduction block as compared to the adjoining regions, in the presence of demyelination.

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the block but not distal to the block have been observed in some cases (Fig. 4.1). Another method for detecting FDB is to look for rate-limited firing of motor units during maximal voluntary contractions. Normally subjects can drive their motor units at 30–50 Hz and sometimes much higher for short periods in maximum contractions. The inability to voluntarily activate motor units at rates much above 20 Hz, especially coupled with periodic dropping out of discharges, strongly points to ratelimited block involving the associated motor nerve

4.5.2. Frequency-dependent block One consequence of demyelination is the reduced ability to transmit trains of impulses (Rasminsky and Sears, 1972; Smith and Hall, 1980; McDonald, 1982). The onset firing frequencies of voluntarily recruited motor units vary from 8 to 13 Hz and firing rates as high as 30–50 Hz or more may be reached during maximal voluntary contractions. Moreover, normal single motor nerve fibers are quite capable of following frequencies of stimulation as high as 100 Hz without evidence of conduction block. However, conventional studies looking for conduction block seldom stimulate the motor nerve at high enough frequencies to reveal frequency-dependent block (FDB). However, there are several ways of detecting FDB and in the case of voluntary contractions, activity or rate-dependent block (ADB or RDB). One method for assessing ADB is to compare the size of the maximum M potential evoked before and after 30–60 s of a maximum voluntary contraction of the affected muscles. Normally, the maximum M potential changes little following maximum voluntary contraction. However, in some patients with demyelinating neuropathies, reductions in the size of the maximum M potential of the order of 20–50% or more, may be observed following the contraction. Possible reasons for such decrements include RDB within the distal preterminal and terminal branches of the affected motor nerve fibers or within the parent motor nerve fiber. Frequency-dependent block may be revealed by supramaximally stimulating the affected motor nerve at frequencies between 5 and 30 Hz, proximal and distal to the site of the putative block. In entrapment neuropathies such as the ulnar nerve in the region of the elbow or the median nerve in the carpal tunnel, obvious decrements in response to stimulation proximal to

Fig. 4.1 Frequency-dependent block in a patient with an ulnar neuropathy localized to the elbow. Routine nerve conduction studies demonstrate marked conduction slowing (9 m/s), mild conduction block and increased temporal dispersion across the elbow segment (top trace). Repetitive nerve stimulation (RNS) using 10 and 20 Hz frequency above the elbow (AE) revealed a further reduction greater than 50% in the hypothenar “M” potential. No decrement was observed in response to similar RNS procedures at the wrist. Moreover, the decrement seen at higher frequencies appeared quantal in size, suggesting the presence of block in single hypothenar motor nerve fibers. W, wrist; BE, below elbow; AE, above elbow; Ax, axilla.

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fibers, especially where no evidence exists of an alternative lesion affecting in the CNS that might otherwise account for the difficulties recruiting motor units.

Table 4.2

4.5.3 Conduction block in acquired and hereditary demyelinating neuropathies

Increase the ILC available for depolarizing the next node of Ranvier

Conduction block in human hereditary demyelinating neuropathies is much less common than is the case in acquired demyelinating neuropathies, despite the sometimes very slow conduction velocities of the former. Why this should be the case is unclear. Possibly, the development of new sodium channels in the paranodal and internodal axolemmal membranes together with remodeling of these regions might serve to increase the effectiveness of the available sodium current sufficient to sustain transmission and overcome any tendency to block between successive nodal regions.

●

Mechanisms for restoring conduction in demyelinated nerve fibers

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●

●

4.6. Restoration of conduction in demyelinated nerve fibers The preceding discussions outlined the major effects of demyelination. We now turn our attention to mechanisms which can restore conduction (Table 4.2). Remyelination of previously demyelinated regions is one obvious means for restoring conduction In demyelinating neuropathies. Based on computer simulation studies, Koles and Rasminsky (1972) showed that conduction might be blocked when the myelin thickness in the affected internode is reduced to less than 2.7% of normal. However, restoration of only a few additional wraps of myelin was all that was required to restore conduction to the affected internode (Koles and Rasminsky, 1972; Saida et al., 1979). In some demyelinated regions, conduction may become continuous, as illustrated by Fig. 4.6. Indirect evidence for continuous in humans has been shown in severe ulnar neuropathies in the region of the elbow and the median nerve in the region of the carpal tunnel and are associated with maximum motor conduction velocities in the affected region of less than 5 m/s (Brown and Watson, 2002). Remyelination in the paranodal region is especially important because of the presence of potassium channels in this region, exposure to which shortens and reduces the ILC available for depolarizing the nodal region. Indeed, Bostock et al. (1978) have shown that blocking the outward currents associated with the K+ channels in experimentally demyelinated nerve fibers by applying 4-aminopyridine (4-AP),

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●

Increase the number of sodium channels in the nodal region Neutralize or remove blocking-antibodies to sodium channels Prolong the opening times of sodium channels Cool the nerve fibers Delay inactivation of the sodium channels Establish new nodes of Ranvier at shorter intervals Block potassium channels with 4-aminopyridine (4-AP) or 3,4-diaminopyridine (3,4-DAP) Inhibit Na+, K+ -ATPase activity in the nodal region with ouabain or digitalis Remyelination and covering exposed potassium channels in the paranodal region Change the kinetics of the sodium channels*

Reduce the ‘load’ on the ILC available to activate the next node of Ranvier ●

● ●

Reduce the passive, transverse leakage currents through the membrane Remyelination in the paranodal region Remyelination in the internodal region

Reduce the ILR of the axoplasm ● ●

Reduce the specific resistivity of the axoplasm* Increase the diameter of the axon

*Denotes possibilities for which there is no direct experimental evidence.

3–4-diaminopyridine (3–4 AP) or tetra-ethyl-ammonium to the affected fibers restores conduction to previously blocked fibers. Moreover blocking the Na+, K+ -ATPase electrogenic pump in the nodal region, that normally hyperpolarizes the membrane, especially in response to high-frequency trains of impulses, might restore conduction as well. Experimentally, the pump has been blocked by ouabain (Kaji and Sumner, 1989a) or digitalis (Kaji and Sumner, 1989b). To the extent that conduction may be blocked by Na+ channel blocking factors (Brinkmeier et al., 1992; Takigawa et al., 1995), other neuroelectric blocking compounds such as interleukin-2 (Brinkmeier et al., 1992b) or nitric oxide (Redford et al., 1997; Shrager et al., 1998; Renganathan et al., 1999), removal or

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neutralization of such factors might restore conduction. Such a hypothesis might well explain the rapid reversal of conduction weakness in some cases of Guillain–Barré polyneuropathy in response to plasmaexchange or IVIg. Proliferation of Schwann cells and the formation of new shorter internodes accompany repair of demyelinated regions of nerve fibers. Each internode corresponds to the territory of a single Schwann cell. During this process, the development of clusters of new sodium channels forms the basis for the formation of new nodes of Ranvier. Such clusters of Na channels at putative nodes of Ranvier may depend on direct contacts and interactions between the axon and nearby myelinating cells (Tzoumaka et al., 1995; Dugandzija-Novakovic et al., 1995; Ching et al., 1999; Dupree et al., 1999; Rasband et al., 1999). The extent to which the densities of sodium channels in the newly formed nodal region correspond to those at the older established nodes is unknown. However by shortening the distance between adjacent nodes, the overall safety factor for transmission may be increased in the affected nerve fibers. Paradoxically, however, the conduction velocity may be slowed somewhat by the increased leakage of the transverse currents through the more numerous nodes of Ranvier created in the wake of regeneration. 4.7. Guillain–Barré’ polyneuropathy: A model for the clinical and pathophysiological features associated with demyelination in humans Guillain–Barré’ polyneuropathy is an excellent model for illustrating the relationships between the clinical features, electrophysiological findings, and the accompanying pathological changes including demyelination. Characteristic clinical hallmarks include paralysis, loss of tendon reflexes, paresthesia, and in some cases, impaired sensation and autonomic dysfunction. The most plausible basis for the weakness is conduction block. However, the pathophysiological basis for the conduction block is controversial. Obvious biophysical changes such as paranodal and internodal demyelination as described by Asbury et al. (1969) would be reason enough for the conduction block; but it is unlikely that these changes explain the sometimes very rapid evolution of the weakness (within hours) or sometimes rapid improvement in some cases following treatment with IVIg or plasma exchange. Such rapid functional changes suggest that circulating factors (possibly blocking antibodies) acting on the Na+ channels proba-

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bly play an important role. Indirect support for this theory is the fact that the maximum motor conduction velocities may be entirely normal early in the course of the disease when the patient is maximally paralyzed. Normal or nearly normal motor and sensory conduction velocities coupled with striking conduction block early in the disease is also consistent with the idea that only a few paranodal regions and internodes may be sufficiently demyelinated or otherwise affected to account for the block. Indeed demyelination affecting as few as 2–3 successive nodal regions and their associated internodes, may be all that is required to produce conduction block and may for example, represent less than 1% of the course of longest nerve fibers in the PNS. The distribution of the conduction block in Guillain–Barré polyneuropathy varies from patient to patient and throughout the course of the disease. In some cases, the chance of conduction block per unit length of nerve appears to be uniformly distributed between the roots and motor point. In other cases, especially within the first few days of the onset of the paralysis, the block appears to chiefly involve the distal preterminal and terminal branches of the motor nerve fibers and the proximal nerve roots (Fig. 4.2). Such a distribution of conduction block may reflect the fact that the blood-nerve barrier is “naturally” more open at the distal extremities of nerve fibers and within the nerve roots. The fact that the earliest observable changes in conduction may be restricted to the proximal and distal segments of the PNS is one reason why conventional motor and sensory conduction studies may be normal at a stage when the patient is paralyzed. Similar distributions for conduction slowing and probably block may be seen in sensory nerve fibers (Brown and Feasby, 1984). Early in the disease, somatosensory evoked potential studies conduction may be entirely normal between the wrist and Erb’s point but the cord dorsum potential may be delayed and temporally dispersed in response to median nerve stimulation at the wrist. Of course, all the above distributions reflect transmission in the larger myelinated nerve fibers and provide no information about pathophysiological changes in small myelinated or unmyelinated nerve fibers in Guillain–Barré polyneuropathy. Slowing of conduction velocities probably explains the loss of time-coded events such as tendon reflexes and vibration sense (Brown, 1984). For example, the unitary excitatory postsynaptic potentials generated by as many as 30–50 or more primary spindle afferents must sum more or less synchronously to bring

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Fig. 4.2 A case of Guillain–Barre’ polyneuropathy, 5 days following the onset of paralysis. This example demonstrates multiple regions of motor conduction block (distal to the wrist, across the elbow and more proximally between the spinal roots and axilla) along the ulnar nerve, recording from the hypothenar muscles. High voltage, low output impedance stimulation was used to supramaximally stimulate the spinal roots. W, wrist; BE, below elbow; AE, above elbow; Ax, axilla; SR, spinal roots (From Brown, WF (1993) Acute and chronic inflammatory demyelinating neuropathies. In: Clinical Electromyography, Boston, Butterworth-Heinemann, IInd ed., pp. 533–559. Reprinted by permission of Butterworth-Heinemann, an imprint of Elsevier).

motor neurons to threshold. In the case of vibration sense the impulses in the requisite rapidly adapting mechanoreceptors would have to arrive in more or less synchronously in bursts entrained at 128 or even 256 Hz for vibration sense to be perceived. Ectopic discharges similar to those observed in demyelinated peripheral nerves (Burchiel, 1980) and dystrophic roots devoid of myelin (Rasminsky, 1978, 1982, 1983) probably explain the paresthesia observed in the primary demyelinating peripheral neuropathies in man (review by Culp and Ochoa, 1982). This type of abnormal spontaneous activity could, through collisions between naturally and ectopically originating impulses in the same nerve fibers block transmission of some of the impulses. Ectopic impulses might also alter the excitability and threshold of the nerve fibers in which they originate and possibly reduce the safety factor for transmission of naturally occurring impulses in the same fibers as well as slowing and possibly blocking impulses in affected nerve fibers. Fasciculations and myokymia may be observed in GBS and other demyelinating diseases, presumably as a result of ectopic discharges of individual motor nerve fibers. In the motor system, slowing of the conduction velocities probably has little impact on the force output of a muscle. One reason for this lies with the fact that the force of a voluntary muscle contraction primarily depends on: (1) the numbers and types of motor units recruited; and (2) the firing patterns and firing rates of those motor units. However, the timing of the

discharges of the various motor units relative to one another is probably not an important factor governing the force generated by the contraction. Hence, motor conduction slowing, even when severe, probably has little or no impact on the strength of a voluntary contraction (Burke, 1980). The preceding point was brought home to the authors through a case of chronic inflammatory polyneuropathy where there was no demonstrable weakness despite conduction velocities never exceeding 5 m/s in any motor nerve, even during periods of remission. 4.7.1. Axonal degeneration in Guillain–Barré’ syndrome Axonal degeneration is common in experimental antiserum-mediated demyelination (Saida et al., 1981) and in Guillain–Barré’ polyneuropathy itself. For example, reactive changes and sometimes degeneration of motor neurons and Wallerian degeneration in peripheral nerve fibers have been observed in Guillain–Barré’ polyneuropathy (Haymaker and Kernohan, 1949; Asbury et al., 1969; Asbury and Johnson, 1978). In keeping with the latter is the finding of denervation in some patients with this disease (Eisen and Humphreys, 1974; McLeod et al., 1976; Brown and Feasby, 1984) and diphtheritic polyneuropathy, another primary demyelinating neuropathy (Kurdi and Abdul-Kader, 1979). Axonal degeneration, either as a secondary effect of primary demyelination or perhaps the primary attack on the axons themselves

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is probably one of the causes for persistent weakness in Guillain–Barré’ polyneuropathy. However, before any EMG evidence of denervation appears it may be impossible to distinguish between conduction block in motor axons undergoing axonal degeneration in which conduction has not yet ceased entirely (Erlanger and Schoepfle, 1946). In the axonal variant of the Guillain–Barré syndrome, axonal degeneration is the single most prominent feature. These cases often progress rapidly to nearly complete or complete somatic muscle paralysis and recover slowly, often leaving their victim with severe and lasting motor deficits. In some cases the axonal degeneration seems to be confined to the distal branches of the nerve fibers and such cases may recover relatively early and completely (Brown et al., 1993).

with localized conduction slowing, conduction block and physiological evidence of Wallerian degeneration distal to the site of the compression (see review by Gilliatt, 1980a). Models of acute compression injury, including the underlying pathology, are probably entirely analogous to the acute radial and common peroneal neuropathies seen in humans (Brown and Watson, 2002) one example of which is illustrated by Fig. 4.3. Invagination of the nodes of Ranvier into the adjacent paranodal region facing away from the site of compression is the most characteristic and early hallmark of acute compression injury followed by demyelination (Fig. 4.4). Wallerian degeneration may be found especially in lesions associated with a high degree of conduction block.

4.8. Conduction block in chronic inflammatory demyelinating polyneuropathy

Chronic experimental nerve compression on the other hand is characteristically associated with detachment of the myelin from the nodal regions facing the site of the compression; at the opposite ends of the internode the myelin buckles and heaps up (Fig. 4.5). These tadpole-like lesions are associated with demyelination and remyelination and regeneration clusters consisting of thinly myelinated immature regenerating nerve fibers. The foregoing changes have also been observed in human chronic entrapment neuropathies (Neary and Eames, 1975; Neary et al., 1975; Ochoa and Marotte, 1973; Gilliatt, 1980b). Chronic nerve compression may be associated with: (1) conduction slowing; (2) possible conduction block; (3) continuous conduction in the region of the compression; (4) Wallerian degeneration distal to the site of the compression; (5) slowed conduction proximal as well as distal to the site of the compression; and (6) ectopic impulse generation and cross talk.

In the chronic acquired and sometimes relapsing type of inflammatory polyneuropathy, demyelination and remyelination and various degrees of onion bulb formation may be found (see reviews by Asbury and Johnson, 1978; Schaumburg et al., 1983). In the relapsing variety, conduction block is characteristically most prominent during the relapse and improves as the degree of clinical weakness improves. Here too, as in Guillain–Barré’ polyneuropathy, conduction block may partially recover within a few hours or days. Maximum motor conduction velocities are often severely reduced between relapses (< 20 m/s) with striking abnormal temporal dispersion and apparently normal strength. The last observation gives weight to the contention that even very slow conduction velocities in motor axons are quite compatible with normal strength and despite the severity of the waveform morphology in terms of its dispersion, phase cancellation is likely occurring rather than actual pathological conduction block. 4.9. Demyelination and conduction block in compression-related neuropathies 4.9.1. Acute compression-related neuropathies Experimental compression of peripheral nerves, whether acute or chronic, is associated with characteristic structural changes in the nodal and paranodal regions, demyelination and in some instances Wallerian degeneration. These lesions are commonly associated

4.9.2. Chronic compression-related neuropathies

4.10. Duration of conduction block Many experimental attempts have been made, using a pneumatic tourniquet, nylon cord or other compressive devices, to better understand the pathophysiology of acute compressive neuropathies with respect to nerve function. The application of these devices in animals (Denny-Brown and Brenner, 1944; Fowler et al., 1972; Rudge et al., 1974) have demonstrated similar results, namely, normal conduction velocities distal to the tourniquet and either conduction block or reduced conduction velocities localized directly under the edges of the tourniquet where nerve damage is the greatest

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Fig. 4.3 Example of motor conduction block (arrows) in an acute compressive lesion of the peroneal nerve, localized to the fibular head. Inching studies were done in (B) and (D) to better localize the lesion across the fibular head. S1–S3, stimulation sites at the ankle, fibular head and popliteal fossa, respectively; PL, peroneus longus; AL, anterolateral muscles; TA, tibialis anterior; EDB, extensor digitorum brevis (Redrawn and adapted from Brown, WF, Watson, BV (1991) Quantitation of axon loss and conduction block in peroneal nerve palsies. Muscle Nerve, 14: 237–244. Reprinted by permission of John Wiley & Sons, Inc.).

(Ochoa et al., 1971, 1972). Conduction velocities under the tourniquet, adjacent to its edges, are usually normal. Tourniquet studies in man (Yates et al., 1981), again have demonstrated similar results with further observation of increased distal latencies with corresponding decreases in conduction velocity distal to the tourniquet implicating ischemia as the lone factor causing these changes (Fig. 4.6). Beneath the compression site, it has been argued whether or not functional abnormalities occur primarily as a result of ischemia or mechanical compression or a combination of both. Generally speaking, any conduction abnormalities dis-

tal to the tourniquet are due to ischemia alone, whereas, abnormalities directly under the tourniquet are likely a result of both ischemia and mechanical compression, particularly across the tourniquet’s edges or border zones. In most cases, when compression times are relatively short in duration, nerve conduction studies return to normal almost immediately following the release of pressure. When the duration of compression is prolonged or when pressures are excessive, nerve conduction abnormalities localized to the edges or border zones of the tourniquet may take longer to recover.

Fig. 4.4 Illustration of the intussusception of the nodes of Ranvier in relation to the pneumatic tourniquet. The direction of nodal displacement was always toward the uncompressed tissue in all fibers examined (Adapted from Ochoa, J, Fowler, TJ and Gilliatt, RW (1972) Anatomical changes in peripheral nerves compressed by pneumatic tourniquet. J. Anat., 113: 433–455. Reprinted by permission of Blackwell Publishing, Oxford).

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Fig. 4.5 Diagrammatic illustration of the effects of long-term or repeated compression of a peripheral nerve fiber. Immediately adjacent to the site of compression (A), many fibers are demyelinated or are thinly remyelinated. Internodes farther removed both proximal and distal to the site of compression show strikingly polarized morphological change (B). These changes consist of loss of the attachment of the myelin lamellae at the ends of internodes facing the site of compression and progressive slippage of the myelin sheath. The latter tended to heap up toward the opposite end of the internode. The slippage and subsequent breakdown of the myelin sheath in the paranodal region produce paranodal demyelination. Further removed from the site of compression, these changes become less pronounced, and the paranodal and internodal regions become correspondingly more normal (C). Perineural thickening and endoneural edema as well as clusters of regenerated and remyelinated fibers are also common findings (Neary and Eames, 1975) (From Brown, WF (1984) Conduction in abnormal nerve. In: The Physiological and Technical Basis of Electromyography. Boston, Butterworth-Heinemann, pp. 37–94. Reprinted by permission of Butterworth-Heinemann, an imprint of Elsevier).

Conduction block accompanying ischemia (< 1 h) may last only seconds up to a few hours following the release of the tourniquet (Denny-Brown and Brenner, 1944). Prolonged pressures (> 2 h) may result in variable, but, longer recovery times due to structural changes within the nerve, namely, nodal intussusceptions causing paranodal demyelination. Acute conduction block in humans also occurs following very short periods of compression (< 1 h), seen for example in some cases of “Saturday night palsy” or peroneal nerve palsy localized to the fibular head; recovery usually occurs within days or at the most several weeks following onset. Other examples of rapid recovery following other mechanisms of compression are illustrated in Figs. 4.7 and 4.8. Occasionally, conduction block may persist for many months thereafter, such has been observed by the authors in cases of acute peroneal palsies and in animal studies (Fowler et al., 1972). One explanation for the long delay in recovery may be the result of paranodal swelling, which has been observed to last for several months following compression (Ochoa et al., 1972). In acute immune-mediated diseases such as GBS, recovery of conduction block is variable and is usually nonuniform throughout the course of the nerve (Fig. 4.9). In the more chronic immune mediated diseases, for example, CIDP or multifocal motor conduction block neuropathy, electrophysiological evidence of conduction block may persist for months to years despite normalizing of strength testing following treatment. Care should be taken, however, when attempting to correlate recovery of conduction block with clinical recovery. For example, in some cases where strength may have returned to normal,

increased temporal dispersion and waveform cancellation may so reduce the maximum “M” potential across previously blocked regions of the nerve, that it may be impossible to assess conduction block using electrophysiological studies. Conversely, there are those cases with residual clinical weakness despite electrophysiological recovery of conduction block. This group of patients is of particular interest because the weakness may be due to frequency dependent block, described earlier in the chapter. Neuropathies associated with vasculitis may be accompanied by electrodiagnostic findings, which may be mistaken for conduction block associated with demyelination, especially in the first few days following the onset of the neuropathy (Fig. 4.10). However after several days the compound muscle action potential distal to the block characteristically falls dramatically in a manner consistent with infarction of the nerve and Wallerian degeneration of the nerve distal to the site of the infarction. 4.11. Axonal degeneration 4.11.1. Wallerian degeneration Wallerian degeneration is a term used to describe the structural alterations in normal nerves distal to a crush injury or transection of the nerve and has been the subject of several reviews (Lieberman, 1971, 1974; Joseph, 1973; Thomas, 1974; Dyck, 1975; Urich, 1976; Allt, 1976; Sunderland, 1978, 1979, 1980; Asbury and Johnson, 1978; Selzer, 1980; Zochodne, 2000). The emphasis here is on those physiological abnormalities that accompany the early degenerative

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proximally for several millimeters and further in more extensive injuries. Reactive changes such as central chromatolysis develop in the parent motor neuron or dorsal root ganglion cells, the intensities of which are related to the proximity of the transection to the neuron and the age of the subject (Sunderland, 1978). Conduction velocities may be slowed up to 70 mm proximal to experimental transection or crush injuries, (Cragg and Thomas, 1961; Kuno et al., 1974a, 1974b; Mendell et al., 1976). Reasons for this occurring may be as a result of axonal atrophy (Cragg and Thomas, 1961; Aitkin and Thomas, 1962) and widening of the nodal regions along with internodal demyelination. (Dyck et al., 1981). Similar reductions in conduction velocities proximal to compressive or transection injuries have been reported in humans (Gilliatt, 1973; Stöhr et al., 1977; McComas et al., 1978). 4.11.3. Distal to the transection

Fig. 4.6 Left: Schematic of the relative positions of the stimulating electrodes at the levels of the wrist (S1), just distal to the tourniquet (S2), and proximal to the tourniquet (S3). Right: Selected records to illustrate the changes in the hypothenar (HT)’ “M” response to stimulation at the S1, S2 and S3 levels at various times prior to and after tourniquet inflation and following release of the tourniquet. Note: (1) the earliest evidence of conduction block occurs across the tourniquet between S2 and S3; (2) persistence of the distal “M” response (S1); (3) the overall cascade beginning at the most proximal level of stimulation (S3) and moving towards S1 and (4) the relative degree of conduction block and delay (From Yates, SK, Hurst, LN, Brown, WF (1981) The pathogenesis of pneumatic tourniquet paralysis in man. J. Neurol. Neurosurg. Psychiatry, 44: 759–767).

and later regenerative processes, both proximal and distal to the level of the injury, and which are most relevant to electrodiagnostic studies in humans. The physiological changes distal to the injury site are identical for both crush and transection injuries. There are, however, important distinctions between the two types of injury in the course of the subsequent repair and regenerative processes. 4.11.2. Proximal to the transection Even when a nerve has been cleanly transected, retrograde degeneration in the axon commonly extends

Following transection of the phrenic nerve in the rat, the earliest degenerative changes begin in the motor axon terminals and the endplates (Miledi and Slater, 1970). These changes are associated with cessation of both endplate potentials (EPPs) and miniature endplate potentials (MEPPs). The latent period between the transection and failure of neuromuscular transmission is proportional to the length of the distal stump (Miledi and Slater, 1970). Neuromuscular transmission fails before conduction failure in the nerve fibers (Miledi and Slater, 1970; Gilliatt and Hjorth, 1972; Wilbourn, 1977). Conduction velocities in the degenerating axons generally remain normal until transmission completely blocks; although conduction velocities may be reduced somewhat just prior to complete loss of excitability (Gilliatt and Hjorth, 1972; Wilbourn, 1977). In degenerating axons, transmission may be blocked at one or more points distal to the site of injury (Erlanger and Schoepfle, 1946). In humans, the nerve becomes unexcitable distal to a transection, within three to seven days (Gilliatt, 1973; Wilbourn, 1977). Hence, it is important for assessing the prognosis in incomplete transection to carry out the electrophysiological studies at least 7 to 10 days postinjury. In the early postdenervation period, the muscle membrane becomes partially depolarized. This is accompanied by the appearance of tetrodotoxinresistant action potentials and changes in the kinetics of the sodium channels (Thesleff, 1974). Thereafter, the extrajunctional regions of the membrane become

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Fig. 4.7 Recording from the thenar “M” potential during supination in a 67-year-old male with “dynamic” conduction block across the forearm as a result of nerve ischemia. The first and second waveforms for each study (A), (B), (C), (D) and (E) were elicited following stimulation of the median nerve at the wrist and elbow, respectively. Note: there was little change in conduction velocity despite progressive conduction block, which was complete at approximately 35 minutes of forearm supination. The conduction block recovered completely within 2 minutes of forearm pronatation (From Watson, BV, Parkes, AW, Brown JD (2002) Transient forearm conduction block in the median nerve. Muscle Nerve, 25: 456–460. Reprinted by permission of John Wiley & Sons, Inc).

much more sensitive to acetylcholine, indicating the development of more extrajunctional acetylcholine receptors. Denervated muscle membranes become more irritable. The latter is manifest by increased insertional activity, provoked by insertion or movement of a needle electrode in the affected muscle and the appearance of fibrillation potentials. Fibrillation potentials represent spontaneously generated single muscle fiber action potentials originating in the region of the old neuromuscular junction or at various extrajunctional sites along the muscle fibers. The latent period between the injury and the appearance of fibrillation activity is proportional to the length of the motor axons distal to their sites of interruption. For example, it may take three to four weeks for abnormal spontaneous activity to develop in the hand muscles following injuries to the C8–T1 roots or adjacent lower brachial plexus in humans. On the other hand, only one to two weeks may supervene between interrupting the C5–6 roots or adjacent upper brachial plexus and the appearance of fibrillation potentials in the rhomboid, spinatii, or deltoid muscles. In a similar length-dependent manner only seven to ten days may be required following acute injuries to the lumbar or sacral roots before fibrillation activity appears in the lumbosacral paraspinal muscles. Provided some part of the motor innervation is spared, within a short time, the

remaining intact motor axons begin to sprout terminal and preterminal collateral branches. These proceed to reinnervate those muscle fibers that have lost their innervation. This process changes the innervation patterns in the muscle and leads to grouping together of like-innervated muscle fibers. Following acute interruption of the axonal continuity by either crush or transection, regeneration of the axons in the proximal stump begins within 24 h. Fig. 4.11 illustrates the temporal-spatial progression of regeneration in a previously severed cat nerve. Sprouts and growth cones develop at the proximal cut ends of axons, sometimes within 24 h. Some sprouts appear to spring from nodal regions proximal to the injury site. One important distinction between crush and transection injuries affecting peripheral nerves lies with how the two affect the continuity of the Schwann cell basement membrane (see Asbury and Johnson, 1978). In the case of all but the most severe crush injuries, continuity of the basement membranes is retained while in the case of transection of the nerve, continuity is lost. Basement membrane tubes are continuous between the roots and the distal terminal axonal branches, and provided they remain intact, may serve as guides for the regenerating axon sprouts to cross the site of the injury in the shortest time and reach their intended peripheral targets (muscle or sense organs).

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Fig. 4.8 Example of nerve ischemia in the superficial peroneal (SP), deep peroneal (DP) and sural sensory nerves during noninvasive ankle distraction in a control subject. To simulate the noninvasive procedure used for ankle arthroscopy a force of 135 N (30 lbs) was applied to the ankle through an ankle distraction harness for 60 minutes. Stimulation of all 3 sensory nerves was done proximal to the harness throughout the procedure. Note: on an average, for the seven control subjects tested, reduction in the sensory amplitudes began to occur approximately 30 minutes into the study at distraction forces greater than 90 N (20 lbs) with almost complete conduction block at 60 minutes. Full recovery of the SP and DP sensory nerve amplitudes was seen in all cases studied within 5 minutes after the release of the harness. The sural nerve was spared as a result of the harness design (From Dowdy, PA, Watson, BV, Amendola, A, Brown, JD (1996) Noninvasive ankle distraction: relationship between force, magnitude of distraction, and nerve conduction abnormalities. Arthroscopy, 12: 64–69. Reprinted by permission of W.B. Saunders Company, an imprint of Elsevier).

Destruction or misalignment of basement membrane tubes, characteristic of rejoined sectioned peripheral nerves, creates severe problems because many sprouts may fail to successfully cross the injury site. Furthermore, of those that do, many are unable to find their matching basement membrane tubes in the distal stump and thereby eventually fail to find their natural target postsynaptic cells or end organs. Faulty regeneration patterns can lead to lasting weakness, sensory impairment, and bizarre reinnervation patterns. Injuries extending over any appreciable distance along the path of a nerve and those accompanied by an overly vigorous connective response only increase the latent period required for the axons to cross the injury site and reduce the numbers of nerve fibers able to successfully bridge the gap (Sunderland, 1978). Characteristically only 1 or 2 of the many developing axon sprouts make it to the periphery contained within a single basement mem-

brane tube. Such branches, if they mature and reach the periphery, can form the basis for various axon reflexes whose presence can sometimes be electrophysiologically identified in the clinic. The earliest regenerating axons are very thin and devoid of myelin sheaths. Their stimulus thresholds are characteristically very high and their conduction velocities very low (< 1–2 m/s) (Berry et al., 1944; Devor and Govrin-Lippmann, 1979). Conduction in some of these regenerating axons may be continuous, at least prior to myelination, although discontinuous conduction between closely spaced high concentrations of sodium channels cannot be excluded (Feasby et al., 1981a). Some of the latter loci could become nodal regions. Once myelination becomes established, the inward currents become limited to the nodal regions similar to normal nerve (Feasby et al., 1981a, 1981b). Progressive distal regeneration is accompanied by

108 Fig. 4.9 Plots of the maximum hypothenar “M” potential negative peak area (mVms) in response to supramaximal stimulation between wrist and spinal roots 5, 10, 18, 32, 39, 54, 59 and 66 days following the onset of paralysis in a patient with Guillain–Barre’ polyneuropathy. Shown for reference are the mean and 2 standard deviations lower limiting values from control data. Conduction block was complete at 10 days, but, recovery was apparent at the 32-day mark (From Brown, WF and Watson, BV (2002) Recording of electrical activity in nerve trunks and conduction in human sensory and motor nerve fibers. In: Neuromuscular Function and Disease, New York, W.B. Saunders Company, pp. 21–55. Reprinted by permission of W.B. Saunders Company, an imprint of Elsevier).

Fig. 4.10 Examples of acute “axonal” conduction block in a 62-year-old female presenting with a 2-week history of mononeuritis multiplex secondary to a vasculitis. Note, the proximal reduction in the corresponding “M” potentials following stimulation of median (Ax), peroneal (PF) and tibial (PF) nerves. Two weeks later, the results were more in keeping with axonal degeneration as demonstrated by a marked reduction in the distal thenar (median), anterolateral (peroneal) and abductor hallucis (tibial) “M” potentials. W, wrist; E, elbow; Ax, axilla; FH, fibular head; PF, popliteal fossa; A, ankle.

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progressive increases in the axonal diameters and the thickness of their myelin sheaths. These two processes proceed in a proximal to distal manner. Lower stimulus thresholds and progressive increases in impulse velocities parallel the structural maturations. At maturity, conduction velocities approach and sometimes reach normal values (Berry et al., 1944; Sanders and Whitterridge, 1946; Berry and Hinsey, 1947; Cragg and Thomas, 1964; Devor and GovrinLippmann, 1979). The failure to reach normal conduction velocities may have several explanations. For example, the multiplication of Schwann cells resulting in shorter internode lengths, smaller axon diameters, and thinner myelin sheaths in the regenerated nerve fibers might singly or together account for the slower conduction velocities. In humans, maximum conduction velocities may reach 80 or even 100% of their normal mean values following regeneration (Hodes et al., 1948; Struppler and Huckauf, 1962; Ballantyne and Campbell, 1973; Donoso et al., 1979; Buchthal and Kuhl, 1979). The latent period in humans, between the injury and the earliest detectable evidence of reinnervation is

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close to what might be expected, based on regeneration rates of 1–3 mm per day (Jasper and Penfield, 1946; see review by Sunderland, 1978). For example, when the median nerve has been accidentally transected at the level of the wrist, the earliest detectable motor responses in the thenar muscles to electrical stimulation proximal to the transection may take as long as 120 days or more to appear. When allowances are made for the time necessary to cross the transection site and establish enough functional end plates to produce a detectable M-potential, and when account is taken of the distance between the muscle and the transection site, the estimated maximum rate of regeneration was about 1–2 mm per day. Not all axons reach the target tissues at the same time; indeed, there can be very appreciable lags between the arrival of the first few axons and the bulk of the remaining nerve fibers. It is not known whether this range reflects variations in the rate of regeneration among individual nerve fibers or simply wide differences between different nerve fibers in the times they require to cross the injury site.

Fig. 4.11 Experimental model of nerve fiber regeneration in the cat. In this model, the lateral (peroneal) division of the sciatic nerve was transected and immediately re-sutured. Shown are the monophasic dorsal root potentials (DRPs) recorded in oil in response to stimulation of the nerve both proximal and distal to the site of the transection. The nerve was mounted on a tri-polar stimulating electrode (cathode central) and attached to a horizontal calibrated drive. The latter made it possible to stimulate the nerve at successive intervals as short as one mm. The exposed nerve in the thigh was kept warm (37˚C) in the mineral oil. (A) One week following transection, no DRP potential could be detected in response to stimulation of the distal stump 8 or more millimeters distal to the original transection. The large DRPs in response to stimulation 2 and 4 mm distal to the transection probably reflect the proximal spread of the stimulating current to the much lower-threshold regions just proximal to the transection. Continued

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Fig. 4.11, Cont’d (B) The DRPs at two and eight weeks respectively. Note that while a few nerve fibers have reached as far as 25 millimeters beyond the transection site, the main body of the regenerating nerve fibers has not progressed nearly as far. At the eight-week mark, many more fibers have successively bridged the original “gap” and extended much further into the distal stump. Indeed, at this time reinnervation of the peroneal innervated muscles just below the knee had already begun as revealed by substantial reductions in amount of fibrillation activity in these muscles.

The rate of axonal regeneration is not constant, but diminishes as the regeneration proceeds distally. The rate of regeneration may also be somewhat slower following transection rather than crush injuries (Sunderland, 1978). The safety factor for neuromuscular transmission at the immature neuromuscular junctions is commonly reduced and may fail when end plate potentials become subthreshold. Moreover, in some of the immature axon terminals and preterminal branches, where the conduction velocities are characteristically

reduced, transmission may fail, especially at branch points. With time and further maturation, presynaptic conduction and neuromuscular transmission become more secure. Using needle EMG electrodes, the earliest detected motor unit action potentials (MUAPs) are often quite short in duration, their amplitudes small and their shapes may occasionally be polyphasic. Before long, the MUAPs become increasingly hypercomplex in shape and the durations of the MUAPs increase, sometimes

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Fig. 4.11, Cont’d (C). Twenty-four weeks following transaction, the size of the DRP is much larger, approaching values elicited proximal to the transection. Note also the reduced temporal dispersion in the DRP, the latter no doubt reflecting the increasing diameters of the nerve fibers and myelination of the regenerating nerve fibers at this stage. (D). Serial DRPs in a control nerve (From Brown, WF (1984) Conduction in abnormal nerve. In: The Physiological and Technical Basis of Electromyography. Boston, Butterworth-Heinemann, pp. 37–94. Reprinted by permission of ButterworthHeinemann, an imprint of Elsevier).

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dramatically so. These changes reflect the ever-increasing number of muscle fibers contacted by the motor nerve fiber as well as wide differences between the arrival times of their respective muscle fiber action potentials within the pick-up territory of the electrode. The latter differences in arrival times, in turn, reflect wide differences among conduction times of the various preterminal and terminal motor nerve fibers within the motor unit as well as differences among the conduction velocities of the muscle fibers in the unit. Later, as the presynaptic motor nerve fibers mature and muscle fiber diameters become more uniform within the reconstituted motor units, the associated MUAPs increase in amplitude and become less temporally dispersed. One of the earliest signs or reinnervation is a reduction in the number of fibrillation potentials. Further regeneration and reinnervation increases the size of the compound “M” potential and the force generated by the muscle in response to a supramaximal stimulus delivered to the presynaptic nerve. The number of voluntarily recruited motor units steadily increases, although the order of recruitment may be somewhat upset following a previous transection of a nerve (Milner-Brown et al., 1974). Equivalent maturation in sensory axons is harder to detect. For example, it is possible to detect the presence of even one motor axon once innervation of a sufficient number of muscle fibers in the motor unit has taken place. However, the compound nerve action potential associated with regenerating sensory fibers is simply too small and temporally dispersed to detect even with near-nerve needle electrode recording techniques (Donoso et al., 1979; Buchthal and Kuhl, 1979). Where large numbers of nerve fibers fail to cross the site of the injury many of the “frustrated” sprouts may form a tangled ball or neuroma. Spontaneous discharges have been clearly shown in such experimental neuromas (Wall and Gutnick, 1974a, 1974b; Devor and Bernstein, 1982). These spontaneous afferent barrages in human neuromas may, in part at least, explain the pains, paresthesia, and phantom limb sensations experienced by these patients. 4.12. Neuromuscular transmission and conduction block in regenerating nerve fibers During reinnervation, the surviving axons sprout new terminals which, in most cases, migrate toward the original endplate regions of the denervated muscle fibers. The density of acetylcholine receptors within the endplate zone remains relatively normal within

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weeks following denervation (Slater and Vincent, 1992), however, a portion of these receptors carry embryonic-like properties resulting in unstable neuromuscular junctions (Loring and Salpeter, 1980; Gu and Hall, 1988). As a result of these newly formed, immature junctions, neuromuscular transmission often fails, possibly as a result of sub-threshold endplate currents. As also, nerve conduction may be blocked along the immature preterminal branches of the regenerating nerve fiber. Neuromuscular transmission failure can occur in any condition where there has been evidence of denervation. An example of this is often seen in motor neuron diseases where decrement can be observed, even at low frequencies (1Hz), while performing routine motor nerve conduction studies. Needle electromyography also often demonstrates the immaturity of axonal collateral sprouting with blocking of satellite or link potentials, observed during voluntary motor unit activation. Single-fiber EMG can also be used to demonstrate neuromuscular block, or increased jitter, in cases of faulty neuromuscular transmission during nerve regeneration. 4.13. Neuronopathies and axonopathies The Wallerian degeneration, that follows transection or crush of a nerve, is not representative of the types of degeneration seen in more slowly progressive primary axonal peripheral neuropathies. Traumatic interruption of the axon shuts down in all those metabolic and structural processes in the distal stump, which depend on axoplasmic transport of molecular constituents originating in the cell body. This contrasts with the situation in degenerative neuronal and axonal neuropathies where only a part of the metabolic or transport processes may be affected. Moreover, the cell bodies and proximal axons are not directly injured in traumatic injuries and, except in very proximal axonal lesions, are able to sustain repair and regeneration of the axon. By comparison, in degenerative neuronal and axonal neuropathies, the cell bodies or axons may well be dysfunctional for a variety of reasons and, therefore, are less capable of regenerating and sustaining their end organs, peripheral synaptic connections, or postsynaptic cells. Furthermore collateral sprouting and reinnervation and hypertrophy of muscle fibers may mask the severity of axon loss in the axonopathies or neuronopathies because of the comparatively much slower pace of these disorders in contrast to that of traumatic lesions.

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The distinction between the neuronopathies, where the primary injury is thought to be in the cell body, and axonopathies, where the injury site is thought to be to the peripheral axonal process, is at best arbitrary. Too little is known about the basic mechanisms involved to make this distinction anything more than speculative in most diseases. The term “dying-back” neuropathy was introduced by Greenfield (1954) and Cavanagh (1964) to describe those diseases where the primary abnormalities were thought to reside in the cell bodies. In response to dysfunctional changes in the cell bodies, the earliest degenerative changes begin at the distal extremities of the axonal processes (both central and peripheral) and progress toward the parent cell bodies, leading as the terminal event to degeneration and loss of the neuron. Detailed examination of experimental and human toxic neuropathies has led, however, to questions about the dying-back hypothesis (see review by Spencer and Schaumburg, 1976). These authors claimed that the patterns of axonal degeneration seen in these intoxications differed from the patterns expected in a dying-back neuropathy. For example, when the neurons are the primary site of the attack, the earliest and most severe degeneration should begin at the distal extremities of those neurons that have the largest peripheral volume to maintain (and perhaps the largest metabolic load). Degeneration should also begin earliest at the distal extremities of those neurons that have the longest and largest-diameter peripheral axons. This simple view has not turned out to be true in the case of some of the toxic neuropathies where degeneration begins not at the axon terminals, but at multiple preterminal sites. Spencer and Schaumburg (1976) also pointed out that although the degeneration does primarily involve the longest and thickest myelinated axons, exceptions do exist to this model. For example, in experimental acrylamide neuropathy, pacinian corpuscles begin to degenerate prior to the primary endings of muscle spindles (Schaumburg et al., 1974). Moreover, in experimental n-hexane neuropathy, nerve fibers, that innervate the calf muscles, degenerate before nerve fibers of equivalent diameter but longer length, such as those that innervate the feet (Spencer and Schaumburg, 1976, 1980). Whatever their precise mechanism(s), the general pattern of these degenerations is common to a wide variety of peripheral neuropathies (Spencer et al., 1978; Spencer and Schaumburg, 1980). The accompanying electrophysiological abnormalities have been documented in the various experimental models and in humans. They include:

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(1) Early receptor failures. For example, abnormal responses in muscle stretch receptors have been observed prior to any detectable change in conduction proximal to the receptor endorgans (Sumner and Asbury, 1975; Lowndes et al., 1978a, 1978b). (2) Selective losses of the largest-diameter myelinated axons. These losses may explain reductions in maximum conduction velocities in acrylamide neuropathy (Hopkins and Gilliatt, 1971). (3) Centripetal progression of conduction failure in the affected nerve fibers Ever more proximal progression of the degeneration of the nerve fibers is associated with the centripetal advance of border zone proximal to which conduction can be sustained without failure. Associated with the latter is the finding that the thresholds of affected nerve fibers are highest at the extremities of the degenerating fibers. (4) Preferential involvement of certain physiological types of axons and relative sparing of other axons of equivalent diameter. For example, in acrylamide neuropathy, sensory nerve fibers of equivalent diameter tend to be more involved than the motor axons (Sumner, 1980). Human peripheral nerves cannot be examined with the same precision as in experimental models. However, equivalent electrophysiological abnormalities have been observed in human neuronopathies and axonopathies. These include: (1) Maximum conduction velocities tend to remain within the normal range. In cases where the maximum conduction velocities fall below the normal range, there may be several explanations. For example, there may have been selective degeneration and loss of the fastest conducting nerve fibers or sufficiently severe losses of nerve fibers overall, that even on a random basis, the fastest-conducting nerve fibers were lost. In these two scenarios the slower conduction velocities may simply reflect the velocities of the remaining, normally slower-conducting nerve fibers. In the latter regard it is well to remember that there is about a 30% range in the conduction velocities of motor axons in normal human motor nerves. Alternatively, slower conduction velocities might be an indication of slowing of

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conduction in normally more rapidly conducting nerve fibers. Conduction velocities in such primary axonal or neuronal diseases might also be slowed because of axonal atrophy (Baba et al., 1982) and perhaps secondary demyelination. For these reasons, caution should be exercised while interpreting maximum conduction velocities because even maximum conduction velocities as low as 20–30 m/s might be seen at the late stages of primary axonal or neuronal diseases in the absence of primary demyelination, especially where only a few motor nerve fibers remain. (2) Disproportionate increases in terminal conduction times may be observed. In these disorders, the earliest structural abnormalities begin at or near the terminal extremities of the central and peripheral processes of the sensory and motor neurons (Spencer and Schaumburg, 1976). These degenerative changes may be associated with secondary paranodal and segmental demyelination and, later, the formation of shorter internodes. Such structural changes would be expected to slow conduction velocities and prolong terminal latencies and would explain the disproportionate prolongation in distal latencies sometimes observed in certain human metabolic and toxic neuropathies, although in some diseases such as alcoholic neuropathy this has not been observed (Behse and Buchthal, 1977). Progressive centripetal degeneration described by Sumner (1980) in acrylamide neuropathy would be a better explanation for the centripetal progressive loss of excitability in sensory nerve fibers reported by Casey and LeQuesne (1972) in alcoholic neuropathy. (3) Denervation and reinnervation. These phenomena are simply a direct consequence of the axonal degeneration and, in general, are more severe in distal than in proximal muscles. Other neurological disorders characterized by primary involvement of the neuron are the motor neuron diseases of which amyotrophic lateral sclerosis (ALS) is the most common example. The etiology of these disorders is not known but has recently been reviewed (Brown et al., 2000). The electrophysiological abnormalities in ALS (Chan and Brown, 2000) are similar to those in the primary axonopathies, except in ALS the “upper” motor neurons as well as the bulbar and

spinal motor neurons are among the primary targets (Hudson, 1981). 4.14. Can axonal neuropathies be distinguished from demyelinating neuropathies based on their conduction velocities? Is there a conduction velocity below which electromyographers can confidently speak of a primary demyelinating neuropathy? The answer is no. The reason being some motor neuronopathies and axonopathies may be associated with conduction velocities less than 50% of control values. Conduction tends to be most slowed in the distal segments of the longest nerve fibers and most apparent where there are only a few surviving motor nerve fibers supplying distal muscles such as the extensor digitorum brevis muscle in the foot. Possible explanations for such slowed conduction velocities in axonal neuropathies include axonal atrophy as well as secondary demyelination. Maximum motor conduction velocities in the less severely affected nerves of such cases tend to be much more normal in contrast to primary demyelinating neuropathies such as Charcot–Marie– Tooth type IB, where the conduction velocities are much more uniformly slowed. References Aitkin, JT and Thomas, PK (1962) Retrograde changes in fiber size following nerve section. J. Anat., 96: 121–129. Allt, G (1976) Pathology of the peripheral nerve. In: DN Landon (Ed.), The Peripheral Nerve. John Wiley & Sons, New York, pp. 666–739. Arroyo, EJ and Scherer, SS (2000) On the molecular architecture of myelinated fibers. Histochem. Cell Biol., 113: 1–18. Asbury, AK, Arnason, BG and Adams, RD (1969) The inflammatory lesion in idiopathic polyneuritis. Medicine, 48: 173–215. Asbury, AK and Johnson, PC (1978) Pathology of peripheral nerve. In: JL Bennington (Ed.), Major Problems in Pathology. W.B. Saunders, Philadelphia. Baba, M, Fowler, CJ, Jacobs, JM and Gilliatt, RW (1982) Changes in peripheral nerve fibers distal to a constriction. J. Neurol. Sci., 54: 197–208. Ballantyne, JP and Campbell, MJ (1973) Electrophysiological study after surgical repair of sectioned peripheral nerves. J. Neurol. Neurosurg. Psychiatry, 36: 797–805.

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Feasby, TE, Bostock, H and Sears, TA (1981a) Conduction in regenerating dorsal root fibers. J. Neurol. Sci., 49: 439–454. Feasby, TE, Pullen, AH and Sears, TA (1981b) A quantitative ultrastructural study of dorsal root regeneration. J. Neurol. Sci., 49: 363–386. Fowler, TJ, Danta, G and Gilliatt, RW (1972) Recovery of nerve conduction after a pneumatic tourniquet: observations on the hind-limb of the baboon. J. Neurol. Neurosurg. Psych., 35: 638–647. Gilliatt, RW (1980a) Acute compression block. In: AJ Sumner (Ed.), The Physiology of Peripheral Nerve Disease. W.B. Saunders, Philadelphia, pp. 287–315. Gilliatt, RW (1980b) Chronic nerve compression and entrapment. In: AJ Sumner (Ed.), The Physiology of Peripheral Nerve Disease. W.B. Saunders, Philadelphia, pp. 316–339. Gilliatt, RW (1973) Recent advances in the pathophysiology of nerve conduction. In: JE Desmedt (Ed.), New Developments in Electromyography and Clinical Neurophysiology. Karger, Basel, pp. 2–18. Gilliatt, RW and Hjorth, RJ (1972) Nerve conduction during wallerian degeneration in the baboon. J. Neurol. Neurosurg. Psychiatry, 35: 335–341. Greenfield, JC (1954) The Spinocerebellar Degeneration. Blackwell, Oxford. Gu, Y and Hall, ZW (1988) Immunological evidence for a change in subunits of the acetylcholine receptor in developing and denervated rat muscle. Neuron, 1: 117–125. Haymaker, W and Kernohan, JW (1949) The LandryGuillain–Barré syndrome. Medicine, 28: 59–141. Hodes, R, Larrabee, MG and German, W (1948) The human electromyogram in response to nerve stimulation and the conduction velocity of motor axons. Arch. Neurol. Psychiatry, 60: 340–365. Hopkins, AP and Gilliatt, RW (1971) Motor and sensory nerve conduction velocity in the baboon: normal values and changes during acrylamide neuropathy. J. Neurol. Neurosurg. Psychiatry, 34: 415–426. Hudson, AJ (1981) Amyotrophic lateral sclerosis and its association with dementia, parkinsonism and other neurological disorders: a review. Brain, 104: 217–247. Huxley, AF and Stämpfli, R (1949a) Evidence for saltatory conduction in peripheral myelinated nerve fibers. J. Physiol., 108: 315–339. Huxley, AF and Stämpfli, R (1949b) Saltatory transmission of the nervous impulse. Arch. Sci. Physiol., 3: 435–448.

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Jasper, H and Penfield, W (1946) The rate of reinnervation of muscle following nerve injuries in man as determined by the electromyogram. Trans. R. Soc. Can., 5: 81–91. Joe, EH and Angelides, KJ (1993) Clustering and mobility of voltage-dependent sodium channels during myelination. J. Neurosci., 13: 2993–3005. Joe, EH and Angelides, K (1992) Clustering of voltagedependent sodium channels on axons depends on Schwann cell contact. Nature, 356: 333–335. Johnston, WL, Dyer, JR, Castellucci, VF and Dunn, RJ (1996) Clustered voltage-gated Na+ channels in aplysia axons. J. Neurosci., 16: 1730–1739. Joseph, BS (1973) Somatofugal events in wallerian degeneration: a conceptual overview. Brain Res., 59: 1–18. Kaji, R, Sumner, AJ (1989a) Ouabain reverses conduction disturbances in single myelinated nerve fibers. Neurology, 39: 1364–1368. Kaji, R and Sumner, AJ (1989b) Effect of digitalis on central demyelinative conduction block in vivo. Ann. Neurol., 25: 159–165. Koles, ZJ and Rasminsky, M (1972) A computer simulation of conduction in demyelinated nerve fibers. J. Physiol., 227: 351–364. Kuno, M, Miyaya, Y and Muñoz-Martinez, EJ (1974a) Properties of fast and slow alpha motoneurones following motor reinnervation. J. Physiol., 242: 273–288. Kuno, M, Miyata, Y and Muñoz-Martinez, EJ (1974b) Differential reaction of fast and slow alpha motorneurones to axotomy. J. Physiol., 240: 725–739. Kurdi, A and Abdul-Kader, M (1979) Clinical and electrophysiological studies of diphtheritic neuritis in Jordan. J. Neurol. Sci., 42: 243–250. Lieberman, AR (1974) Some factors affecting retrograde neuronal responses to axonal lesions. In: R Bellairs, EG Gray (Eds.), Essays On The Nervous System. Clarendon Press, Oxford, pp. 71–105. Lieberman, AR (1971) The axon reaction: a review of the principal features of perikaryal responses to axon injury. Int. Rev. Neurobiol., 14: 49–124. Loring, R and Salpeter, MM (1980) Denervation increases turnover rate of junctional acetylcholine receptors. Proc. Natl. Acad. Sci. USA, 44: 2293–2298. Lowndes, HE, Baker, T, Cho, E and Jortner, BS (1978a) Position sensitivity of de-efferented muscle spindles in experimental acrylamide neuropathy. J. Pharmacol. Exp. Ther., 205: 40–48.

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Lowndes, HE, Baker, T, Michelson, LP and VincentAblazey, M (1978b) Attenuated dynamic responses of primary endings of muscle spindles: a basis for depressed tendon responses in acrylamide neuropathy. Ann. Neurol., 3: 433–437. McComas, AJ, Sica, RE and Banerjee, S (1978) Longterm effects of partial limb amputation in man. J. Neurol. Neurosurg. Psychiatry, 41: 425–432. McDonald, WI (1982) Clinical consequences of conduction defects produced by demyelination. In: WJ Culp, J Ochoa (Eds.), Abnormal Nerves and Muscles as Impulse Generators. Oxford University Press, Oxford. McLeod, JG, Walsh, JC, Prineas, JW and Pollard, JP (1976) Acute idiopathic polyneuritis. J. Neurol. Sci., 27: 145–162. Mendell, LM, Munson, JB and Scott, JG (1976) Alterations of synapses on axotomized motoneurones. J. Physiol., 255: 67–79. Miledi, R and Slater, CR (1970) On the degeneration of rat neuromuscular junctions after nerve section. J. Physiol., 207: 507–528. Milner-Brown, HS, Stein, RB and Lee, RG (1974) Pattern of recruiting human motor units in neuropathies and motor neuron disease. J. Neurol. Neurosurg. Psychiatry, 37: 665–69. Neary, D and Eames, RW (1975) The pathology of ulnar nerve compression in man. Neuropathol. Appl. Neurobiol., 1: 69–88. Neary, D, Ochoa, J and Gilliatt, RW (1975) Subclinical entrapment neuropathy in man. J. Neurol. Sci., 24: 283–298. Nicholls JG, Martin AR, Wallace BG and Fuchs PA (2000) From Neuron to Brain. Sinauer Sunderland, MA. IVth Ed. Novakovic, SD, Deerinck, TJ, Levinson, SR, Shrager, P and Ellisman, MH (1996) Clusters of axonal Na+ channels adjacent to remyelinating Schwann cells. J. Neurocytol., 25: 403–412. Ochoa, J, Danta, G, Fowler, TJ and Gilliatt, RW (1971) Nature of the nerve lesion caused by a pneumatic tourniquet. Nature, 233: 265–266. Ochoa, J, Fowler, TJ and Gilliatt, RW (1972) Anatomical changes in peripheral nerves compressed by a pneumatic tourniquet. J. Anat., 113: 433–455. Ochoa, J and Marotte, L (1973) The nature of the nerve lesion caused by chronic entrapment in the guinea pig. J. Neurol. Sci., 19: 491–495. Peles, E and Salzer, JL (2000) Molecular domains of myelinated axons. Curr. Opin. Neurobiol., 10: 558–565.

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Rasband, MN, Peles, E, Trimmer, JS, Levinson, SR, Lux, SE and Shrager, P (1999) Dependence of nodal sodium channel clustering on paranodal axoglial contact in the developing CNS. J. Neurosci., 19: 7516–7528. Rasminsky, M (1982) Ectopic excitation, ephaptic excitation and auto excitation in peripheral nerve fibers of mutant mice. In: WJ Culp, J Ochoa (Eds.), Abnormal Nerves and Muscles as Impulse Generators. Oxford University Press, Oxford, pp. 344–362. Rasminsky, M (1978) Ectopic generation of impulses and cross talk in spinal nerve roots of “dystrophic mice”. Ann. Neurol., 3: 351–357. Rasminsky, M (1983) Ectopic impulse generation in pathological nerve fibers. Trends Neurosci., 6: 38–90. Rasminsky, M and Sears, TA (1972) Internodal conduction in undissected demyelinated nerve fibers. J. Physiol., 227: 323–350. Redford, EJ, Kapoor, R and Smith, KJ (1997) Nitric oxide donors reversibly block axonal conduction: Demyelinated axons are especially susceptible. Brain, 120: 2149–2157. Renganathan, M, Cummins, TR, Hormuzdiar, WN, Black, JA and Waxman, SG (1999) Nitric oxide is an autocrine regulator of Na+ currents in axotomized DRG neuron. J. Neurophysiol., 83: 2431–2443. Ritchie, JM (1995) Physiology of axons. In: SG Waxman, JD Kocsis, PK Stys (Eds.), Axon: Structure, Function and Pathophysiology. Oxford University Press, Oxford, pp. 68–96. Ritchie, JM (1979) Sodium channels in muscle and nerve. In: AJ Aguayo, G Karpati (Eds.), Current Topics in Nerve and Muscle Research. Excerpta Medica Amsterdam, Oxford, pp. 210–219. Rogart, RB and Stämpfli, R (1982) Voltage-clamp studies of mammalian myelinated nerve. In: WJ Culp, H Ochoa (Eds.), Abnorma Nerves and Muscles as Impulse Generators. Oxford University Press, Oxford, pp. 193–210. Ruch, TC, Patton, HD, Woodbury, JW and Towe, AL (1965) Neurophysiology. W.B. Saunders, Philadelphia. Rudge, P, Ochoa, J and Gilliatt, RW (1974) Acute peripheral nerve compression in the baboon. J. Neurol. Sci., 23: 403–420. Rushton, WAH (1951) A theory of the effects of fiber size in medullated nerve. J. Physiol., 115: 101–122. Saida, G, Saida, K, Saida, T and Asbury, AK (1981) Axonal lesions in acute experimental demyelination: a sequential teased nerve fiber study. Neurology, 31: 413–421.

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Saida, K, Sumner, AJ, Saida, T, Brown, MJ and Silberberg, DH (1979) Anti-serum-mediated demyelination: relationships between remyelination and functional recovery. Ann. Neurol., 8: 12–24. Sanders, FK and Whitterridge, D (1946) Conduction velocity and myelin thickness in regenerating nerve fibers. J. Physiol., 105: 152–174. Schaumburg, HH, Spencer, PS and Thomas, PK (1983) Disorders of Peripheral Nerves. FA Davis, Philadelphia. Schaumburg, HH, Wisniewski, HM and Spencer, PS (1974) Ultrastructural studies of the dying-back process. Neuropathol. Exp. Neurol., 33: 260–284. Selzer, M (1980) Regeneration of peripheral nerve. In: AJ Sumner (Ed.), The Physiology of Peripheral Nerve Disease. W.B. Saunders, Philadelphia, pp. 358–431. Shrager, P, Custer, AW, Kazarinoa, K, Rasband, MN and Mattson, D (1998) Nerve conduction block by nitric oxide that is mediated by the axonal environment. J. Neurophysiol., 79: 529–536. Slater, C and Vincent, A (1992) Structure and development of the neuromuscular junction. In: A Vincent, D Wray (Eds.), Neuromuscular Transmission: Basic and Applied Aspects. Pergamon Press Ltd., Oxford, England, pp. 1–26. Smith, KJ and Hall, SM (1980) Nerve conduction during peripheral demyelination and remyelination. J. Neurol. Sci., 48: 201–219. Spencer, PS, Sabri, MI, Schaumburg, HH and Moore, CL (1978) Does a defect of energy metabolism in the nerve fiber underlie axonal degeneration in polyneuropathies? Ann. Neurol., 5: 501–507. Spencer, PS and Schaumburg, H (1976) Centralperipheral distal axonopathy-the pathology of dying-back polyneuropathies. In: HM Zimmerman (Ed.), Progress in Neuropathology. Grune & Stratton, New York, pp. 253–295. Spencer, PS and Schaumburg, HH (1980) Experimental and Clinical Neurotoxicology. Williams & Wilkins, Baltimore. Stöhr, M, Schumm, F and Reill, P (1977) Retrograde changes in motor and sensory conduction velocity after nerve injury. J. Neurol., 214: 281–287. Struppler, A and Huckauf, H (1962) Propagation velocity in regenerated motor nerve fibers. Electroencephalogr. Clin. Neurophysiol., Suppl. 22: 58–60. Sumner, AJ (1980) The Physiology of Peripheral Nerve Disease. W.B. Saunders, Philadelphia, pp. 340–357.

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Peripheral Nerve Diseases Handbook of Clinical Neurophysiology, Vol. 7 J. Kimura (Ed.) © 2006 Elsevier B.V. All rights reserved

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CHAPTER 5

Principles of stimulation and recording Hiroyuki Nodera* and Ryuji Kaji Department of Clinical Neuroscience, Graduate School of Medicine, University of Tokushima, Japan

CHAPTER 1 5.1. Introduction

5.3. Electrodes

With the recent advances in computer technology, even a novice electromyographer can comfortably perform electrophysiological testing with the aid of a semi-automatic electromyography (EMG) device. However, a lack of basic knowledge in electronics or the appropriate settings of such a device prevents the operator from performing a proper assessment (or troubleshooting) of the obtained waveforms. This chapter deals with the practical aspects of instrumentation and recording, and presents a brief discussion on electronics.

Electrodes can be used for either stimulation or recording. This section will discuss only recording electrodes (stimulation through needle electrodes is discussed in the section on stimulators). Two types of recording electrode can be used: surface and needle electrodes. Both of these have strengths and drawbacks, and it is important to understand the unique characteristics of each type so that they can be used as appropriate. The electrical function of the electrodes is to allow the detection of action potentials generated by nervous tissue in the body. In general, a surface electrode is preferred for clinical nerve conduction studies (NCSs), since it records from all motor units (although with some limitations), whereas a needle electrode has a much smaller recording radius, making it less suitable for studying CMAPs.

5.2. Basic electrical components required for nerve conduction studies The minimal apparatus required to perform EMG/nerve conduction tests include electrodes, amplifiers, a visual display, loudspeakers, and a stimulator; the use of data storage devices is optional (Misulis, 1989; Gitter and Stolov, 1995; Kimura, 2001; Dumitru et al., 2002). The same apparatus can often be used for different techniques, but certain differences often exist in specific applications. For instance, a stimulator may not be necessary for surface EMG or sympathetic skin response. In addition, surface electrodes are used to obtain muscle action potentials (MAPs) from the many underlying motor units, whereas a needle electrode is used to target activities from the limited number of motor units located near to the tip of an electrode.

* Correspondence to: Ryuji Kaji, MD, PhD, Department of Clinical Neuroscience, Graduate School of Medicine, University of Tokushima, 2-50-1 Kuramotocho, TokushimaCity 770-8503, Japan. E-mail address: [email protected] Tel.: +81-88-633-7207; fax: +81-88-633-7208.

5.3.1. Surface electrodes By applying surface electrodes over the muscle or the nerve, it is possible to record transcutaneously the summated activity from many motor units or nerves, respectively. Given the different regions of the body that are recorded from and the possible purposes of those recordings, a variety of surface recording electrodes have been used in both the clinical and research setting (e.g., discs, bar electrodes with multiples discs embedded in plastic, self-adhesive disposable electrodes, and ring electrodes with adjustable steel electrodes). A disc electrode is made from a durable metal, which has good electrical conductivity for clinical use (e.g., silver, gold, stainless, nichrome, and platinum). Due to biological and environmental noise, the electrical activity from nervous tissue is usually obtained by making bipolar recordings with the reference electrode usually placed near to the recording site. The electrical activity of these two electrodes is processed and the difference between them is displayed on a cathode-ray tube (CRT) screen. Although this method presumes that the reference electrode is electrically inactive,

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waveforms can differ significantly by moving only the reference electrode, reflecting the electrical propagation into the reference. This is discussed further in the section on volume conduction. The biggest advantage of using surface electrodes over needle electrodes is that there is no pain involved because they are applied to the skin without puncturing it, but the waveforms obtained are less well defined due to the high impedance of the skin resulting in a reduction in the magnitude of the potential detected by the electrode. The interface between the skin and electrode ranges from 500 to 9 M (Swain et al., 1985; Panescu et al., 1993), and this can be effectively reduced by: (1) preparing clean electrodes; (2) using skin preparation and cleansing agents, if necessary, or rubbing with an alcohol-based preparatory solution; and (3) applying an electrode cream between the electrode and the skin surface. However, the use of excess amounts of electrode cream may result in unstable placement of the electrode or bridging between the active and reference electrodes. The choice of surface electrode has no definite answer. The electrical properties of each recording electrode can differ significantly, not only for a particular type of electrode, but also with respect to numerous factors such as its method of manufacture, the electrical material utilized, and the size of the electrode. Therefore, each laboratory should obtain normative data with their own particular recording electrodes. A disposable, self-adhesive electrode with a slightly larger surface area than the traditional disc electrode has recently been introduced, which obviates the need for electrode cream or adhesive tape. Whether the data obtained by using a disposable electrode are comparable to those obtained with a disc electrode is yet to be established. There is no agreement upon the appropriate size of the recording electrode (Bromberg and Spiegelberg, 1997), the most commonly used one being a flat or cup-shaped disc electrode with an average diameter of 1 cm and a recording radius of approximately 20 mm (Barkhaus and Nandedkar, 1994). The performance of larger electrodes has been studied, particularly in the field of motor conduction (Wee and Ashley, 1990; van Dijk et al., 1995; Tjon et al., 1996; Ferdjallah et al., 1999; Finsterer et al., 2002). Larger electrodes generally yield CMAPs with smaller amplitudes and areas, but with comparable conduction velocities and onset latencies. This is understandable because larger electrodes pick up potentials from larger regions with differing degrees of contribution to the CMAP (i.e.,

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a variable potential field in a region with high spatial gradients), thereby “averaging out” the electrical response and yielding a smaller-amplitude CMAP. The regular-sized “small” electrode has been used as an outcome measure for various therapeutic trials to monitor the progression of a disease (Hahn et al., 1996; Russell et al., 1996; Kaji et al., 1998). However, it is thought that the repeatability of the CMAP amplitude may be low due to inconsistent placement of the electrodes, because even the slightest shift in the placement of the active electrode can change the CMAP amplitude significantly (as the spatial gradient is high near the motor point). On the contrary, the longitudinal reliability of CMAP amplitudes appears to be slightly higher when using larger electrodes, which are presumably more suitable for longitudinal monitoring of CMAPs in clinical trials. Theoretically, a larger electrode “averages out” the potentials with different amplitudes underneath the electrode, and the relationship between the maximal motor point and the placement of the electrode may be less strict (unpublished data from Nodera and Kaji, see Fig. 5.1). 5.3.2. Needle electrodes The main advantages of using needle electrodes over surface electrodes are: (1) due to the close proximity of the active electrode to the potential generator and significantly lower impedance than surface electrodes, larger potentials are yielded for which the loss of highfrequency signals due to biological filtering is reduced (see discussion on filters); and (2) due to the smaller size of the electrode, potentials from a smaller number of motor units are obtained, making motor unit analysis easier. However, there are also drawbacks, such as: (1) the pain inflicted on the subject and the potential transmission of pathogens; (2) the high cost, whether using disposable or reusable (following autoclaving) electrodes; (3) lower reproducibility of the potentials, most significantly the action potential amplitude; and (4) the risk (albeit fairly low) of injuring nervous tissue or vasculature exists. Despite these drawbacks, the use of needle electrodes has been studied extensively, and it is believed that in normal individuals they do not usually cause an elevation of muscle enzyme levels (Finsterer et al., 2002). Still, great care should be taken when using needle electrodes in patients with significantly elevated levels of muscle enzymes or in patients with renal failure. Monopolar needle electrodes are made from stainless steel covered by a Teflon sleeve, and have an

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CMAP amplitudes (mV)

Repeatability of conventional (“small”) vs. “large” electrodes

B

12 11 10 9 8 7 6 5 4 3 2

R small L small R large L large

1

2

3

4

Week

Fig. 5.1 Median motor nerve conduction study performed using a large E1 electrode covering a large area of the muscle belly (A). The CMAP obtained by the large recording electrode is smaller than that obtained by the small recording electrode, since the spatial gradient is high near the motor point and the effect is averaged out by the large electrode. The thin and long E2 recording electrode was placed around the interphalangeal joint for test-retest consistency. Compared with serial CMAP recordings achieved with a conventional disc recording electrode (a “small” electrode), those achieved with a large recording electrode are smaller and, more importantly, are more reproducible (B).

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average diameter of approximately 0.8 mm. The entire length of the electrode is insulated, apart from a 0.2-mm to 0.4-mm length at the distal tip, which is used for the recording. It has been shown that needles with only their tip bared provide a better signal-tonoise (S/N) ratio than those with a completely bare shaft, and are thus recommended for clinical use (Schoonhoven and De Weerd, 1984; Chu et al., 1987). Furthermore, since varying the length of the bared section of the needle electrode significantly alters the action potential parameters of motor units, particularly the amplitude and the area of the motor unit potentials (Chu et al., 1987), the length of the bared tip should always be taken into account when analyzing data. A surface electrode or a second needle electrode inserted into the subcutaneous tissue serves as a reference electrode. Coaxial concentric needle electrodes comprise a stainless-steel cannula with a wire in the center of the shaft. The difference is obtained between the electrical discharges recorded from the inside wire and the outside rim, the distance between which is very short, so that this needle electrode records potentials that have a lower amplitude and a smaller number of phases than a monopolar needle, which has a restricted recording radius (Howard et al., 1988; Pease and Bowyer, 1988; Nandedkar et al., 1990; Nandedkar and Sanders, 1991; Kohara et al., 1993; Dumitru et al., 1997; Joynt, 1998; Trojaborg, 1998). Hence, this type of the electrode is rarely used for NCSs, with the possible exception of studies that focus on single or only a few motor units. In NCSs, a needle electrode is typically used for near-nerve sensory recording, where an active electrode is inserted subcutaneously and the tip of the needle is set in close proximity to a nerve in order to obtain subtle discharges from that nerve (Rosenfalck, 1978; Oh et al., 1984; Smith, 1998; Wexler et al., 1998; Seo and Oh, 2002). A similar application is in microneurography, in which a thin tungsten needle electrode is inserted into a nerve trunk to obtain discharges from a single unmyelinated nerve. This method is used primarily for studies of the autonomic nervous system (including itching) and of sensory systems, with emphasis on neuropathic pain (to investigate myelinated and unmyelinated sensory fiber activities; Hallin and Torebjork, 1970, 1973; Handwerker et al., 1991; Schmelz et al., 1997; Kuwabara et al., 2000; Weidner et al., 2002; Schmelz et al., 2003), and for recording somatosensory, auditory, and visual evoked potentials.

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5.4. Electrode amplifiers In order to properly visualize biological electric signals ranging in size from microvolts to millivolts on visual displays, the signals must be significantly amplified. For example, if the actual amplitudes of the signal are 1 μV and 1 mV, amplification factors of ×1 000 000 and ×1000, respectively, are required to register on an oscilloscope display set at 1 V/cm. Several methods are currently employed to accomplish this significant degree of amplification. The first method is to multiply the amplifying effects by using a preamplifier with a gain of, for example, ×500, followed by several amplifier and attenuator stages to produce a gain of ×2–2000, depending on the range of the required output. The second method involves the use of differential amplifiers. Since the major source of signal noise originates from the surrounding environment and the current power lines, the magnitudes of which far exceed those obtained from biological tissues, amplifiers have been developed that cancel out the nonbiological noise recorded from both the active and reference electrodes (the common signal) whilst amplifying the remaining signals from both electrodes. This cancelled potential is called the common signal. The use of these differential amplifiers also effectively eliminates unwanted distant MAPs that affect equally both the active and reference electrodes. The effectiveness of a differential amplifier can be quantified using the common-mode rejection ratio (CMRR) (Perreault et al., 1993). The CMRR is defined as the ratio of the amplification of the difference signal to the amplification of the common-mode voltage. For example, a CMRR of 100000 means that the biological signals are amplified 100 000 times more significantly than unwanted noises appearing as the common-mode voltage. However, even when using a differential amplifier, it may not be possible to completely eliminate the unwanted noise because of, for example, unequal noise on the two electrodes due to different degrees of electromagnetic interference, or the existence of unequal contact impedance between the electrodes, which is very difficult to avoid. 5.5. Filters Most biological electrical potentials consist of a combination of complex discharges with variable frequencies. Fourier analysis of these complex discharges reveals the sum of sine waves of different frequencies as their constituents. In general, discharges from

HIROYUKI NODERA AND RYUJI KAJI

neurological tissue have frequencies ranging from 10/20 KHz. Activities outside of this range are most likely to be artificial and hence should not be included in analyses. For this purpose, a filter can effectively wipe out electrical activities below or above certain frequency: a low-frequency (high-pass) filter cutting out low-frequency activity and a high-frequency (lowpass) filter cutting out high-frequency activity (Maccabee and Hassan, 1992). Since setting the cutoff frequencies of filters inappropriately significantly alters waveforms, it is important to understand the effects of filters (Pease and Pitzer, 1990; Dumitru et al., 2002). If a low-frequency filter (LFF) is set too low, the most notable effect would be an unstable baseline due to the inclusion of very-low-frequency artificial discharges (from, for example, body movement). Conversely, if a LFF is set too high, the lowfrequency component of the biological discharge is removed, with the net effect of lowering the discharge amplitude and quicker return of the waveform to the baseline. If a high-frequency filter (HFF) is set too low, the high-frequency component of the biological discharge is omitted, resulting in delay of the onset latency (at which point the high-frequency component predominates for the discharge to rise quickly from baseline) and a less-steep rise from the baseline, a slight delay in the peak latency (which is less noticeable than that in the onset latency, since high-frequency discharges are not dominant at the peak), and, as an advantage, less-notable high-frequency background noise. Conversely, if the HFF is set too high, too much noise will be included in the waveform (Fig. 5.2). With these points in mind, although there is no universal agreement on this matter, the recommended filter settings for NCSs are as follows: (1) motor NCSs, LFF at 2–10 Hz and HFF at 10 kHz; and (2) sensory NCSs, LFF at 2–10 Hz and HFF at 2 kHz. In general, a body tissue acts as a low-pass (highfrequency) filter, resulting in smoothing of the signals. Hence, an electrode placed far away from the generator of the potential preferentially picks up lowfrequency signals, resulting in waveforms with a slow rise time and a low-pitched sound, erroneously interpreted as “neurogenic, reinnervated” potentials. 5.6. Visual displays As for other electrical visual aids (e.g., the computer screen and television), the CRT has long played a central role in the visual displays used for EMG/nerve

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A

Default filter setting

B

Lower LFF

C

Higher LFF

D

Lower HFF

E

Higher HFF

Fig. 5.2 Effects of varying the filter setting in a model of a CMAP recording. If the low-frequency filter (LFF) is set too low, the baseline becomes less stable due to inclusion of low-frequency discharges (B). Setting the LFF too high reduces the amplitude of the CMAP due to the omission of the slow component of the response (C). If the high-frequency filter (HHF) is set too low, the onset latency becomes longer, the rise time increases, and there is a slight decrease in the peak amplitude (D). If the HFF is too high, the signal contamination with artificial high-frequency noise occurs (E).

conduction tests. Although the digital screen has recently largely replaced the CRT, a basic understanding of how it works would help us to appreciate the basic principles underlying electrical visual displays. The CRT consists of an electron gun, deflecting apparatus, and luminescent screen, which together function essentially as a pointer that can be moved rapidly in two dimensions, whose intensity can be modulated. The electron gun produces a beam of high-speed electrons from a thermionic cathode that are focused to a small spot on the screen. The intensity of these electrons is regulated by the negative potential on a thimble, which is set in front of the cathode and has a small hole through which the electrons pass. A second anode set at a high potential, more than 500 V in most sets, then accelerates the electrons. The electron beam is guided by two deflecting plates set at 90˚ angles to one another (horizontal and vertical deflection plates). An electric field is established between these plates and the electrons are deflected accordingly onto a luminescent screen. When the electrons hit the screen, the phosphor is excited, and emits light.

A delay line is employed in the triggered sweep oscilloscope to retard the arrival of the vertical input waveform at the vertical deflection plates until the trigger circuit and time-base generator have had a chance to start the sweep of the beam. The effect of the delay line is that the leading edge of the vertical input waveform can be viewed on the screen. Thus, one can initiate the horizontal sweep on command for an electromyographer to set successive and recurring motor unit potentials at the beginning of each sweep, enabling detailed analysis. 5.7. Loudspeaker A loudspeaker is a device that converts electric signals into sound waves, and houses a driver that converts electrical audio signals into sound waves. Among the many types of drivers, by far the most common type is the moving-coil electrodynamic piston driver. It has a moving diaphragm that acts like a piston to move air backwards and forwards, thereby creating sound waves. Because the diaphragm can

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only operate effectively over a limited frequency range, multiple diaphragms are usually required for the faithful representation of sound. The sounds produced by different kinds of potentials during examination with a needle electrode are important, and are used for proper identification and analysis of the waveforms. Each potential has its own characteristic sound, and an experienced operator often relies heavily on these sounds rather than the waveforms themselves (Okajima et al., 2000). Owing to the HFF effect of body tissue, a potential that arises some distance from the recording electrode creates a low-pitched sound. The position of the needle must be adjusted until a clear, crisp sound (reflecting a short rise time due to a reduction of the HFF effect) from a nearby unit is obtained. 5.8. Artifacts Not all observed electrical activity originates from nerve or muscle. Although most of the discharges that arise from nonneural tissue or from outside the body have distinct discharge patterns, some others mimic those arising from neural tissue, and even an experienced examiner may have difficulty determining its origin. These artifacts may originate from several sources; for example, from a nonneural tissue, artificial devices, amplifier noise, defective apparatus, movement artifacts, and interference. Each of these are discussed separately here (Kimura, 2001; Dumitru et al., 2002). Artifacts from nonneural tissue can originate from any body tissue with the capability of generating electric signals, but only a few selected body systems can generate discharges that are intense enough to affect EMG or nerve conduction tests. The most notable artifact comes from the heart. Due to the regularity of the cardiac rhythm, it is fairly easy to identify this particular source, but artifacts from a patient with an irregular heart rhythm may be challenging. The cardiac artifact may be pronounced if the reference electrode is placed on the opposite limb to the recording electrode by crossing the midline of the body. Another potential source of the cardiac artifact is the pacemaker potential, which, because of its regularity, is sometimes easily mistaken for a fibrillation signal. Careful inquiry, simultaneous ECG recording, and the presence of synchronization with the pulsation of the blood vessels allow the identification of pacemaker potentials. As discussed above, the detected electric signal must be amplified 1000 to 1000 000 times for effec-

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tive visualization. Hence, even the slightest electrical noise inherent in an amplifier can produce a significant problem. These artifact can be generated by its components, including the resistors, transistors, and integrated circuits. For example, thermal noise, defined as the noise generated by thermal agitation of electrons in a conductor, is proportional to the square root of the absolute temperature and increases with the impedance in the input stage. Another sort of noise, microphonic noise, is caused by the unstable capacitance of various components, and is due mainly to mechanical vibration. In practice, amplifier noise is most evident when the tip of the needle electrode is in fatty tissue (which has a higher impedance than muscle), and high-frequency noise disrupts the baseline. This noise can often be decreased by repositioning of the needle. The most commonly encountered artifacts are attributable to defective apparatus. For example, a broken wire creates bizarre, often very-high-frequency noise that fluctuates with manipulation of the wire. In addition, a partially defective conductor may generate artifacts that occur simultaneously with muscle twitches. Inadvertent insulation of the electrode by blood protein or electrode cream may produce similar artifacts, requiring routine care of the surface of the electrodes by, for example, alcohol-based preparations. The largest source of noise may be the electrolyte-skin interaction, thus emphasizing the need for proper treatment of the skin prior to application of an electrode (Huigen et al., 2002). Movement artifact is produced when a patient contracts a muscle and the surface electrode slides over the skin, resulting in a change in the impedance between the skin and the surface electrode (Conforto et al., 1999). This artifact is often accompanied by sudden deflection of the baseline immediately following muscle twitches. Movement artifacts can be suppressed by encouraging the patient to remain still, immobilization of the region being studied (if possible), and proper fixation of the electrode by tape and, if necessary, by manual compression. Sweat may also alter the input impedance between the skin and the recording electrode, resulting in unsteadiness of the baseline. The sympathetic skin response takes advantage of this artifact in the assessment of autonomic nervous system function (Gutrecht, 1994; Arunodaya and Taly, 1995; Marchello et al., 1996). Another source of artifact that is often problematic for clinical evaluation is the 50- or 60-Hz interference generated by the surrounding electrical equipment (electrostatic

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and electromagnetic interference). Possible sources include, for example, electric fans, lamps, unused power cords plugged into wall outlets, CRT screens, and fluorescent lights. Occasionally, an electric source some distance from the EMG device can cause a noticeable interference; for example, the ignition system of an automobile in a nearby driveway. Similarly, interference from radio broadcasts and mobile phones can be recorded and lessened by relocating the EMG device in the examining room and refraining from using mobile phones. However, given the increasing prevalence of remote communications, these sources of interference may not be completely abolished (Uludag et al., 1997). This type of interference may be exaggerated when an examiner touches the subject, because the examiner’s body acts as an antenna. This type of potential interference may be of enormous clinical significance in bedside testing, especially in an intensive care unit, where the activity of numerous monitoring and therapeutic devices generate significant interference, and in some cases no satisfactory recording can be obtained. The best approach to avoid such interference would be to shield the room and unplug all of the electric apparatus which is, of course, impossible in practice. Thus, the most reasonable and practical approach for obtaining satisfactory waveforms is as follows: (1) to bundle the lead wires from the pickup and ground electrodes to minimize the area susceptible to the field of interference; (2) to attempt to relocate the wires relative to the patient and recording apparatus; and (3) to set a certain distance between the CRT monitor and the patient to avoid the interference from the monitor (Kimura, 2001). The stimulus artifact is another commonly observed artifact, potentially posing a major technical challenge in routine nerve conduction testing. It is essentially a volume-conducted potential of the electrical stimulation (see below on volume conduction) that propagates instantly throughout the body independent of nerve or muscle tissue. Hence, the onset of the stimulus artifact is earlier than that of neural responses, and if the magnitude of the artifact is large enough, the background activity fails to return to baseline, resulting in an illdefined onset latency and amplitude of the responses. The potential reasons for a large stimulus artifact are: (1) poor stimulus isolation from the ground; (2) insufficient separation between the stimulus and recording sites; (3) too great a distance between the active and reference electrodes; (4) excess use of electrolyte cream, resulting in surface spread of the potential;

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(5) high impedance of the stimulating skin region (e.g., callous skin); and (6) and the active and reference electrodes located on different isopotential lines produced by a situation often improved by rotating the anode while maintaining the position of the cathode over the intended nerve. Technical tips for reduction of the stimulus artifact are listed in Table 5.1. 5.9. Stimulators 5.9.1. Types of stimulation Various modes of stimulation can elicit a nerve action potential. For example, several research groups have managed to obtain sensory nerve action potentials (SNAPs) by mechanical, air-puff stimulation (Buchthal, 1980; Hashimoto et al., 1991; Kuwabara et al., 2000; Mizobuchi et al., 2000; Fig. 5.3). Although this “natural” stimulus allows the assessment of mechanoreceptors that are completely bypassed by electrical stimulation, the SNAPs obtained by nonelectrical stimulation are significantly smaller in amplitude than electrically elicited responses, presumably due to the activation of a smaller proportion of the nerve fibers. In practical neurophysiology, electrical stimulation is easy to perform and considered to be reliable method of eliciting potentials; hence, electrical stimulation is the focus of this section. Magnetic stimulation of the central and peripheral nervous systems is a unique alternative mode of stimulation, in that it can be used to stimulate practically any part of the nervous system with relative safety (please refer to Chapter 19).

Table 5.1 Stimulus artifact reduction (Modified from Dumitru et al., 2002) (1) Remove perspiration from skin between stimulator and recording electrodes (2) Use only a small amount of electrolyte cream beneath all electrodes (3) Place ground electrode next to active recording electrode between it and the cathode (4) Only use current strength and duration sufficient to achieve a supramaximal response (5) Reduce the impedance between the skin and all electrodes (6) Elevate the low-frequency filter (use sparingly, if at all because of tendency to distort the waveform) (7) Use a needle cathode (anode)

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Normal

Electric

10 μV 5 ms

Air-puff

5 μV 5 ms

Fig. 5.3 SNAPs obtained by conventional electrical stimulation (top) and air-puff stimulation (bottom) (Kuwabara et al., 2000). The SNAP obtained by air-puff stimulation is characterized by: (1) a smaller amplitude than the electrically stimulated response, since a more limited proportion of the nerve fibers can be stimulated; and (2) a longer distal latency, since air-puff stimulation follows the physiological sensory pathway through sensory receptors, as opposed to electrical stimulation that bypasses these receptors.

5.9.2. Constant-voltage and constant-current stimulators Successful recording relies strongly on the stability of the electrical stimulation, and application of constantvoltage and constant-current electrical stimulation provides two ways of achieving this. Constant-voltage stimulation involves delivering an adjustable voltage across the stimulators, independent of the current, whereas with constant-current stimulation a fixed level of electric current is delivered. As Ohm’s law implies (V = I × R, where V, I, and R denote voltage, current, and resistance, respectively), the level of electric current between the cathode and anode (and hence at the nerve) may vary significantly with fluctuating resistance (e.g., sweating or movement of the stimulator). Constant-voltage and constant-current stimulation generally allow a greater control of the stimulus, especially for studies requiring trains of stimuli. 5.9.3. Basis of electrical stimulation The basis of electrical stimulation is discussed briefly here; a more detailed description can be found in Chapter 2 of the book, “Physiology and Function.” Electrical stimulation is accomplished by applying a

cathode (the negative terminal of a source of electrical current, conventionally colored in black in neurophysiology) and an anode (the positive terminal, conventionally colored in red) in close proximity either transcutaneously or subcutaneously. A cathode attracts positive ions (cations) and an anode attracts negative ions (anions). An electric current will be set up in the underlying tissue in the direction of (by convention) the positive ions (i.e., from anode to cathode). The current of sufficient magnitude enables an underlying nerve to be stimulated as an uneven capacitor. The extracellular surface facing a nerve near the anode is discharged more positively by accumulating negative ions on the surface of a nerve (i.e., hyperpolarization) due to a capacitative effect. Similarly, the extracellular tissue under the cathode is less positively charged, with a less negative membrane potential existing intracellularly. Once the membrane potential under the cathode reaches a certain threshold, that portion of the nerve depolarizes to produce an action potential. In a normal myelinated nerve, the node of Ranvier offers the least resistance to current flow and is the most easily depolarized part of the nerve. Although in conventional neurophysiology the cathode has been recognized as “active” in terms of producing an action potential in an axon beneath it, the effect of anodal stimulation has been debated. “Anodal block” is a local block of nerve conduction caused by hyperpolarization of the nerve cell membrane by an electric stimulus produced by an anode. For example, one might wonder whether reversing the placement of the anode and cathode in a NCS would block the propagation of the action potential; prior studies have argued against the presence of anodal block in routine neurophysiological testing (Dreyer et al., 1993; Kirshblum et al., 1998; Wee et al., 2000). The effective use of an electrical stimulator will result in depolarization of a large area of an underlying nerve. An electrical square-pulse stimulus lasting less than 50 μs may fail to activate some high-threshold fibers, and is not recommended for routine use. On the other hand, electrical stimuli lasting longer than 1 ms tend to be poorly tolerated and are usually unnecessary (except for special techniques required for nerve excitability testing, for example). In a routine clinical setting, most electromyographers use a default stimulation duration of 0.1–0.2 ms, increasing this parameter until a supramaximal response is obtained. However, stimuli of short duration may fail to elicit a response in a patient with severe edema, adiposity, or demyelination (see discussion below), in which case, starting

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with a longer-duration stimulus may be necessary in order to avoid the administration of an excessive number of stimuli. In cases of edema and adiposity, and for research in which prolonged recording is required, the use of subcutaneous stimulation through a pair of needle electrodes could be advantageous. The use of electrodes placed subcutaneously avoids the highresistance cutaneous and transcutaneous tissues and can elicit nerve potentials more easily because of their proximity to the nerve, particularly in cases of edema and adiposity, or when testing a nerve that is located deep below the skin surface. It is not uncommon to develop a skin abrasion after applying a surface stimulator firmly enough to edematous skin in order to obtain nerve responses. In addition, since it is generally easier to achieve a more stable placement using a subcutaneous stimulator than a surface stimulator, the former would be more suitable for lengthy recording sessions, even though it is slightly invasive. A study in which the effects of different types of surface electrodes were investigated demonstrated that there were considerable differences in the torques and electrical impedances obtained between electrodes (i.e., selfadhering pregelled pads, solvent-activated conductive tape, carbonized conductive silicone rubber, and feltcovered metal plates). Therefore, the effective use of a stimulating electrode requires an understanding of its unique characteristics (Nelson et al., 1980). Due to the increased risk of electrical nerve injury, great care should be taken while performing the “nearnerve” technique. Attention should be paid to factors such as a short stimulus duration (e.g., less than 0.05 ms), low stimulus strength (e.g., less than 1.0 mA), and careful clinical observation (e.g., paresthesia and pain) to limit the strength of the stimulation (Rosenfalck, 1978). Particular caution should be exercised when stimulating in the constant-current condition with a needle that has only the tip bared, because the small tip area yields a local current of high intensity, possibly resulting in heat- or current-induced injury of the surrounding tissue (Pease et al., 1989). For the same reason, use of concentric needle electrodes with very short interelectrode distances should be avoided. Since activation thresholds change according to the diameter and degree of myelination (if any) of the axons, stimulation using either a transcutaneous or intradermal electrode generally results in activation of the thicker, heavily myelinated A-alpha fibers (which have a low stimulus threshold) first, followed at increasing stimulus intensities by smaller-diameter fibers with less myelination. Recently, an interesting attempt was made

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to selectively stimulate nociceptive A-delta or C-fibers by using an intraepidermal needle electrode (0.2 mm in depth) to stimulate only the free endings of a nerve located in a region of the epidermis where only nociceptive fibers are found (Inui et al., 2002). By applying electrical currents for longer periods, the current required to elicit the same amplitude of the response decreases. The intensity of an electric current of infinite duration necessary to produce a minimal response is called the rheobase. As the rheobase of a sensory nerve is lower than that of a motor nerve, longer-duration stimuli (e.g., 1 ms) preferentially stimulate sensory nerves, which is the theoretical basis of using long-duration stimuli to analyze the H-wave. Conversely, short-duration stimuli predominantly excite motor nerves. In a research setting, applying different modes of stimulation reveals the otherwise unidentifiable properties of nerves and axons (Bostock et al., 1998; Burke et al., 2001). First, applying two successive electrical stimuli reveals the aftereffects of the stimulation in a nerve (i.e., the recovery cycle). For up to 4 ms after the first stimulation, the stimulus intensity required to elicit a second response from a nerve increases significantly (i.e., the threshold of activation increases), suggesting decreased excitability (absolute and relative refractoriness) due to the transient inactivation of the Na+ channels that were opened during the action potential. Then, at 7 ms poststimulus (or slightly earlier), the nerve is more excitable than at baseline (supernormality), a phenomenon that can be explained by depolarization of the paranodal region due to propagation of the depolarization into the region. Another mode of stimulation, called threshold electrotonus, involves applying very long subthreshold electrical stimuli (e.g., 100–300 ms), which enables the activation of various ion channels to compensate for the changes in the membrane polarization induced by lengthy stimulation. Thus, threshold tracking, takes full advantage of the utility of different modes of stimulation, has shed light on the properties of axon membranes in various neurological conditions (discussed in detail in Chapter 17). Most modern EMG devices have several modes of stimulation that allow the stimulus frequency to be altered. By simply pressing a switch, one may apply either a single stimulus with increasing stimulation intensity or trains of stimuli at a regular frequency (usually 1–2 Hz for most routine NCSs) while increasing the intensity of the stimuli until a maximal response is obtained. Only one of these stimulation

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modes should be used in order to avoid the application of unnecessary stimuli to the subject. The most suitable frequency of stimulation depends on the type of testing being performed. For example, for a microneurographic recording of an unmyelinated sensory fiber, an initial stimulation frequency of 0.25 Hz (once every 4 s) or less is preferred due to its low conduction velocity (approximately 1 m/s) and frequency-dependent slowing (slower nerve conduction velocity with higher stimulation frequency due to a hyperpolarizing afterpotential; Hallin and Torebjork, 1970, 1973; Torebjork and Hallin, 1974; Weidner et al., 2002). In a patient with neuromuscular junction disorder, the preferred stimulation frequency is either 2–3 Hz (for the study of slow repetitive stimulation) or 20–50 Hz (for a rapid repetitive stimulation), but these may elicit a falsely low CMAP amplitude due to fatigue. In general, subjects are intolerant of high-frequency electrical stimulation at greater than 3–4 Hz, although this figure varies significantly between individuals. 5.9.4. Stimulation of demyelinated nerves In a demyelinated nerve (e.g., in chronic inflammatory demyelinating polyneuropathy and Charcot–Marie– Tooth disease type 1), the electrical threshold is often markedly increased, requiring stronger stimuli than a normal nerve so as to elicit a response (Cappelen-Smith et al., 2001). This is most likely due to demyelination in the paranodal region. In a normal state, the paranodal region is covered by layers of myelin sheaths that exhibit a very high electrical resistance. Hence, as Ohm’s law implies, the current through the paranode is very small and the majority of the current is directed to the node (with lower resistance), enabling efficacious stimulation at the node. In a demyelinated nerve, on the other hand, the high resistance at the paranode is not maintained and the electrical stimuli pass through both the nodal and paranodal regions, failing to achieve efficacious stimulation through the node, and thereby requiring much higher-intensity electrical stimuli for depolarization to occur. Hence, in severely denervated nerves (e.g., Dejerine–Sottas disease), even electrical stimulation at the strongest current available to the EMG device and with a long stimulus duration may fail to elicit a maximal response (Kaji, 2003). 5.10. S/N Ratio and averaging The relative amplitudes of the signal and the background noise (i.e., the S/N ratio) must be taken into

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account when assessing the observed signal, to avoid falsely interpreting the unwanted noise as a real neural response. Even when using the most appropriate technique and taking the greatest care, it may not be possible to abolish the background noise, and hence the intended nerve activity remains embedded in the noise (i.e., there is a low S/N ratio), especially in sensory NCSs due to the already relatively small response amplitude that is elicited. A simple and widely used method for removing background noise (and hence increasing the S/N ratio) is by averaging many epochs of the same evoked potential. By averaging several recordings with the same repeating stimuli, the activities unrelated to the stimuli (with no relationship in time) will cancel out and only evoked “true” potentials remain visible because they are time locked to the onset of the stimulation. In general, the S/N ratio improves in proportion to the square root of the number of epochs averaged. For example, averaging four responses improves the S/N ratio twofold. The recommended degree of averaging depends on the S/N ratio of the intended responses, and may range from few times to up to 30. Obviously, unnecessarily large numbers of epochs averaged should be avoided. When the amplitude of the intended response is very small, one should be very cautious about interpreting the observed response. The biggest source of error might be intermittent muscle activity (e.g., voluntary movements or tremor), because the amplitude of the MAP far exceeds that from a sensory nerve, and hence contamination by even a single MAP would not be completely canceled out and thus may be falsely interpreted as sensory activity. To avoid this, one should concentrate on the loudspeaker during the averaging for any possible MAP or other noise as a potential source for contamination. 5.11. Volume conduction and waveforms As mentioned in Section 5.3, a pair of electrodes is placed on the surface of a nerve or a muscle to record its action potential. For a successful recording, there must be a difference in electrical potential between the active electrode (E1) and the reference electrode (E2), where E1 is the electrode placed near to the nerve or muscle and E2 is placed away from the generator of the action potential (so that the direct potential change from the electronic generator is smaller at E2). The theory behind this is that E2 is electrically inactive; however, a significant degree of electrical activation of

PRINCIPLES OF STIMULATION AND RECORDING

E2 may occur (e.g., in ulnar motor conduction studies) and may affect the waveforms significantly, depending on the placement of E2 (Kincaid et al., 1993; Kincaid, 1999; McGill and Lateva, 1999). Traditionally, in electroneurophysiology, negativity at E1 relative to E2 is expressed as an upward deflection of the trace (“negative up, positive down”), although exceptions do exist. In practice, using either extracellular surface or needle electrodes, the extracellular (not the intracellular) potential is measured, and so that the recorded polarity of the extracellular component is the opposite of the intracellular one (i.e., in depolarization, the recorded potential is negative, and vice versa). Volume-conduction theory provides us with an understanding of the origins of the diphasic or triphasic waveforms of a nerve action potential or a MAP. First, imagine a nerve with its polarizing phase described as “inside negative, outside positive” and depolarizing phase as “inside positive, outside negative.” An intracellular nerve action potential has a duration of approximately 0.5 ms (Paintal, 1966), and given the normal conduction velocity of a nerve of 50 m/s and assuming that the length of the depolarized portion of a nerve is 25 mm [length (mm) = time (ms) × conduction velocity (m/s)], one can assume that a 25-mm length of “inside positive, outside negative” segment propagates an action potential at the speed of 50 m/s. As no potential difference is produced between the two isopotential segments in either the depolarizing or polarizing portion, the only regions that are potentially electrically significant enough to be recorded are both ends of the depolarized segment where portions of positive and negative polarity are located next to each other (a dipole, expressed as “+ −”). Hence, the electrical model that best represents the passage of a depolarized portion can be illustrated as quadripoles (“+ − − +”), or more simplified as tripoles (“+ − +”) that move in the nerve or the muscle (also described as leading and trailing dipoles; Stegeman et al., 1997; Dumitru, 2000). As the magnitude of a potential is proportional to the distance of an electrode from the source, a larger change in the action potential is detected in E1 (rather than E2), such that passage of a “+ − +” tripole by E1 is recorded as a triphasic waveform of an initial positive deflection, followed by negative and positive polarities (hence, shown as initial downward deflection from the baseline followed by the upward deflection, then again a downward deflection). If E1 is placed over the motor point where a MAP is generated, the lack of effect of the leading positive polarity changes the morphology

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a

G1

G2

G1

G2

b c d e

a⬘ b⬘ c⬘ d⬘

Fig. 5.4 Diphasic (top) and monophasic (bottom) recording of an action potential, that is represented by the shaded area. The impulse propagates from left to right, and no difference of potential was detected in a, c, and e. In b, the active electrode, G1 (E1) becomes negative relative to the reference electrode, G2 (E2). Similarly, G1 (E1) becomes positive relative to G2 (E2) in e. In the bottom trace, the shaded area on the right illustrates the killed end due to permanent depolarization, making G1 (E1) positive relative to G2 (E2) in a′, b′, and d′. In b′, there is no potential difference between G1 (E1) and G2 (E2), causing an upward deflection from the positive baseline to zero potential (modified from Kimura, 2001).

of the waveform from triphasic to biphasic (an initial negative followed by a positive deflection; Kimura, 2001; Dumitru et al., 2002; Fig. 5.4). 5.12. Field theory: “near-field” and “far-field” potentials The term “far-field potential” was taken from antenna theory and incorporated into clinical neurophysiology for use as a theoretical means of explaining brainstem auditory evoked potentials, where apparently potentials far from the recording electrodes are recorded (Stegeman et al., 1997). The terms “near-field” and “far-field” potentials are frequently used, but this is not technically correct, since all potentials are recorded at some distance from the potential generator and there is no consistent distinction between the two terms (AAEE, 1987). Still, with this caveat in mind,

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the concept of near-field and far-field potentials helps in the understanding of the different mechanisms of potential generation. Since the introduction of the concept into clinical neurophysiology, far-field potentials have been actively investigated and have been detected using other techniques (e.g., using somatosensory evoked potentials (SSEPs) and in peripheral nerve conduction studies). To understand far-field potentials, the opposite concept—near-field potentials—should first be understood. A near-field potential is defined as a waveform that clearly changes in amplitude, polarity, morphology, and usually latency when the active electrode is serially repositioned over small distances (Stegeman et al., 1997; Dumitru, 2000), as the source of the electric potential is presumably generated in close proximity to the active electrode. Conversely, far-field potentials can be defined as waveforms that do not fulfill the definition of a near-field potential (i.e., it does not change with repositioning the active electrode over small distances (i.e., a potential that is fairly stable in amplitude, polarity, and morphology, and usually latency). As discussed earlier, an action potential is recorded by the volume conduction of dipoles, with the assumption that the nerve has constant characteristics longitudinally (e.g., identical diameter, conduction velocity, and no curvature). Imbalance between the positive and negative poles in a dipole can generate a potential that can be detected not only by an active electrode in close proximity, but also by an electrode that is some distance from the site of action potential generation. This is considered to be the source of the far-field potentials observed in peripheral NCSs (Kimura et al., 1983; Machida et al., 1983; Kimura et al., 1984; Yamada et al., 1985; Kimura et al., 1986). Clinically, in motor unit action potentials and compound action potentials, a far-field potential can be recorded at the terminal phase of repolarization, and is observed as a long-lasting downward deflection. Examples of generators of far-field potentials include the following: (1) Fixed neural generators (e.g., the cardiac pacemaker). (2) “Boundary” or “junctional” potentials, that occur when leading/trailing dipoles propagate along a nerve with a varying diameter (e.g., from small to large cylinder) or different tissue resistances. As shown in Fig. 5.5 for an experimental antidromic sensory conduction study, along with the apparent sensory action potential (the latency

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of which increases as the nerve is stimulated more proximally), there are two potentials with consistent latencies irrespective of the stimulation site. These are considered to be far-field potentials generated at the wrist and the base of the digit. As to the reason why a potential generated at the junction has a constant latency irrespective of the stimulation site may be best understood by using the following analogy. An oncoming train (the axonal volley) becomes simultaneously visible (recorded as a far-field potential) to all bystanders at a distance (a series of recording electrodes) as it emerges from a tunnel (partition of the volume conductor), whereas the same bystanders see the train pass by at different times (recorded as a near-field potential)

Fig. 5.5 Stimulation of the left radial nerve 10 cm proximal to the styloid process of the radius, and serial recording of antidromic sensory potentials along the course of the radial nerve (A). There was an abrupt change of the volume conduction at the “0” level at the base of the second digit.

PRINCIPLES OF STIMULATION AND RECORDING

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ND (−3) − (−4)

PW

NW

PD

(−3) − v

(−2) − (−3) (−1) − (−2)

(−2) − v

(0) − (−1)

(−1) − v

(+1) − (0)

(0) − v

(+2) − (+1)

N1

(+1) − v

(+3) − (+2) (+2) − v

(+4) − (+3)

(+3) − v

(+5) − (+4) (+6) − (+5)

(+4) − v

(+7) − (+6)

(+5) − v

(+8) − (+7)

(+6) − v 5.0 μV 1.0 μs

(+7) − v (+8) − v

P1

P2

Bipolar Recording

5.0 μV 1.0 ms

B

Referential Recording

Fig. 5.5, Cont’d (B). In the reference recording, the ring electrode was placed around the fifth digit, whereas the ring electrode was placed around the second digit distally in the bipolar recording. Note that in the reference recording, there were biphasic peaks (far-field potentials) with a stationary latency irrespective of the recording sites along the digits, first appearing near the wrist and at the base of the digit, respectively. This observation supports the theory that the far-field potential is produced at the boundary between two surrounding media with different conduction characteristics along the nerve.

depending on their position along the railroad (Kimura et al., 1986). (3) A change in the direction of neural propagation (e.g., bending of a joint, anatomical curvature). (4) The termination of excitable tissue, when sudden termination of propagation occurs resulting in the disappearance of the positive pole, then a surviving negative pole is no longer counterbalanced and a potential is generated (Stegeman et al., 1997). It is commonly believed that far-field potentials are generated at synapses, reflecting potentials generated at the cell bodies of second-order neurons. However, the SSEP studies have revealed that, on the contrary, the far-field potentials reflect primarily the physical

relationship between the nerve and the surrounding conducting medium, for example, at the shoulder and pelvic girdles where simple bending the shoulder joint alters the latency and morphology of the waveforms slightly, but significantly (Kimura et al., 1983, 1984). References Arunodaya, GR and Taly, AB (1995) Sympathetic skin response: a decade later. J. Neurol. Sci., 129: 81–89. Barkhaus, PE and Nandedkar, SD (1994) Recording characteristics of the surface EMG electrodes. Muscle Nerve, 17: 1317–1323. Bostock, H, Cikurel, K and Burke, D (1998) Threshold tracking techniques in the study of human peripheral nerve. Muscle Nerve, 21: 137–158.

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Bromberg, MB and Spiegelberg, T (1997) The influence of active electrode placement on CMAP amplitude. Electroencephalogr. Clin. Neurophysiol., 105: 385–389. Buchthal, F (1980) Action potentials in the sural nerve evoked by tactile stimuli. Mayo. Clin. Proc., 55: 223–230. Burke, D, Kiernan, MC and Bostock, H (2001) Excitability of human axons. Clin. Neurophysiol., 112: 1575–1585. Cappelen-Smith, C, Kuwabara, S, Lin, CS, Mogyoros, I and Burke, D (2001) Membrane properties in chronic inflammatory demyelinating polyneuropathy. Brain, 124: 2439–2447. Chu, J, Chan, RC and Bruyninckx, F (1987) Progressive teflon denudation of the monopolar needle: effects on motor unit potential parameters. Arch. Phys. Med. Rehabil., 68: 36–40. Conforto, S, D’Alessio, T and Pignatelli, S (1999) Optimal rejection of movement artefacts from myoelectric signals by means of a wavelength filtering procedure. J. Electromyogr. Kinesiol., 9: 47–57. Dreyer, SJ, Dumitru, D and King, JC (1993) Anodal block V anodal stimulation. Fact or fiction. Am. J. Phys. Med. Rehabil., 72: 10–18. Dumitru, D (2000) Physiologic basis of potentials recorded in electromyography. Muscle Nerve, 23: 1667–1685. Dumitru, D, Amato, A and Zwarts, M (2002) Electrodiagnostic Medicine. Hanley & Belfus, Philadelphia. Dumitru, D, King, JC and Nandedkar, SD (1997) Motor unit action potentials recorded with concentric electrodes: physiologic implications. Electroencephalogr. Clin. Neurophysiol., 105: 333–339. Ferdjallah, M, Wertsch, JJ and Harris, GF (1999) Effects of surface electrode size on computer-simulated surface motor unit potentials. Electromyogr. Clin. Neurophysiol., 39: 259–265. Finsterer, J, Mittendorfer, B, Neuhuber, W and Loscher, WN (2002) Influence of disposable, concentric needle electrodes on muscle enzyme and lactate serum levels. J. Electromyogr. Kinesiol., 1: 329–337. Gitter, AJ and Stolov, WC (1995) AAEM minimonograph #16: instrumentation and measurement in electrodiagnostic medicine—Part II. Muscle Nerve, 18: 812–824. Gutrecht, JA (1994) Sympathetic skin response. J. Clin. Neurophysiol., 11: 519–524. Hahn, AF, Bolton, CF, Pillay, N, Chalk, C, Benstead, T, Bril, V, Shumak, K, Vandervoort, MK and Feasby, TE (1996) Plasma-exchange therapy in chronic

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inflammatory demyelinating polyneuropathy. A double-blind, sham-controlled, cross-over study. Brain, 119 (Pt 4): 1055–1066. Hahn, AF, Bolton, CF, Zochodne, D and Feasby, TE (1996) Intravenous immunoglobulin treatment in chronic inflammatory demyelinating polyneuropathy. A double-blind, placebo-controlled, cross-over study. Brain, 119 (Pt 4): 1067–1077. Hallin, RG and Torebjork, HE (1970) Microneurographic analysis of the fibre spectrum of human sensory nerve fascicles. Acta Physiol. Scand., 80: 24A–25A. Hallin, RG and Torebjork, HE (1973) Electrically induced A and C fiber responses in intact human skin nerves. Exp. Brain Res., 16: 309–320. Handwerker, HO, Forster, C and Kirchhoff, C (1991) Discharge patterns of human C-fibers induced by itching and burning stimuli. J. Neurophysiol., 66: 307–315. Hashimoto, I, Yoshikawa, K, Sasaki, M, Gatayama, T and Nomura, M (1991) Conduction velocity and temporal dispersion of the nerve volleys evoked by air-puff stimulation of the index finger and palm. Electroencephalogr. Clin. Neurophysiol., 81: 102–107. Howard, JE, McGill, KC and Dorfman, LJ (1988) Properties of motor unit action potentials recorded with concentric and monopolar needle electrodes: ADEMG analysis. Muscle Nerve, 11: 1051–1055. Huigen, E, Peper, A and Grimbergen, CA (2002) Investigation into the origin of the noise of surface electrodes. Med. Biol. Eng. Comput., 40: 332–338. Inui, K, Tran, TD, Hoshiyama, M and Kakigi, R (2002) Preferential stimulation of Adelta fibers by intra-epidermal needle electrode in humans. Pain, 96: 247–252. Joynt, RL (1998) The concentric versus the monopolar needle electrode. The case for monopolar needles. Muscle Nerve, 21: 1804–1806; discussion 1809. Kaji, R (2003) Physiology of conduction block in multifocal motor neuropathy and other demyelinating neuropathies. Muscle Nerve, 27: 285–296. Kaji, R, Kodama, M, Imamura, A, Hashida, T, Kohara, N, Ishizu, M, Inui, K and Kimura, J (1998) Effect of ultrahigh-dose methylcobalamin on compound muscle action potentials in amyotrophic lateral sclerosis: a double-blind controlled study. Muscle Nerve, 21: 1775–1778. Kimura, J (2001) Electrodiagnosis in diseases of nerve and muscle: principles and practice. Oxford University Press, New York.

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Kimura, J, Kimura, A, Ishida, T, Kudo, Y, Suzuki, S, Machida, M, Matsuoka, H and Yamada, T (1986) What determines the latency and amplitude of stationary peaks in far-field recordings? Ann. Neurol., 19: 479–486. Kimura, J, Mitsudome, A, Beck, DO, Yamada, T and Dickins, QS (1983) Field distribution of antidromically activated digital nerve potentials: model for far-field recording. Neurology, 33: 1164–1169. Kimura, J, Mitsudome, A, Yamada, T and Dickins, QS (1984) Stationary peaks from a moving source in far-field recording. Electroencephalogr. Clin. Neurophysiol., 58: 351–361. Kincaid, JC (1999) The compound muscle action potential and its shape. Muscle Nerve, 22: 4–5. Kincaid, JC, Brashear, A and Markand, ON (1993) The influence of the reference electrode on CMAP configuration. Muscle Nerve, 16: 392–396. Kirshblum, S, Cai, P, Johnston, MV, Shah, V and O’Connor, K (1998) Anodal block in F-wave studies. Arch. Phys. Med. Rehabil., 79: 1059–1061. Kohara, N, Kaji, R and Kimura, J (1993) Comparison of recording characteristics of monopolar and concentric needle electrodes. Electroencephalogr. Clin. Neurophysiol., 89: 242–246. Kuwabara, S, Mizobuchi, K, Toma, S, Nakajima, Y, Ogawara, K and Hattori, T (2000) “Tactile” sensory nerve potentials elicited by air-puff stimulation: a microneurographic study. Neurology, 54: 762–765. Maccabee, PJ and Hassan, NF (1992) AAEM minimonograph #39: digital filtering: basic concepts and application to evoked potentials. Muscle Nerve, 15: 865–875. Machida, M, Yamada, T and Kimura, J (1983) [“Far field potentials” after stimulation of the median and tibial nerve in man]. Nippon Seikeigeka Gakkai Zasshi, 57: 271–284. Marchello, L, Donadio, V and Montagna, P (1996) The sympathetic skin response: a neurological perspective. Funct. Neurol., 11: 283–299. McGill, KC and Lateva, ZC (1999) The contribution of the interosseous muscles to the hypothenar compound muscle action potential. Muscle Nerve, 22: 6–15. Misulis, KE (1989) Basic electronics for clinical neurophysiology. J. Clin. Neurophysiol., 6: 41–74. Mizobuchi, K, Kuwabara, S, Toma, S, Nakajima, Y, Ogawara, K and Hattori, T (2000) Single-unit responses of human cutaneous mechanoreceptors to air-puff stimulation. Clin. Neurophysiol., 111: 1577–1581.

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Nandedkar, SD and Sanders, DB (1991) Recording characteristics of monopolar EMG electrodes. Muscle Nerve, 14: 108–112. Nandedkar, SD, Tedman, B and Sanders, DB (1990) Recording and physical characteristics of disposable concentric needle EMG electrodes. Muscle Nerve, 13: 909–914. Nelson, HE, Jr., Smith, MB, Bowman, BR and Waters, RL (1980) Electrode effectiveness during transcutaneous motor stimulation. Arch. Phys. Med. Rehabil., 61: 73–77. Oh, SJ, Kim, HS and Ahmad, BK (1984) Electrophysiological diagnosis of interdigital neuropathy of the foot. Muscle Nerve, 7: 218–225. Okajima, Y, Tomita, Y, Ushijima, R and Chino, N (2000) Motor unit sound in needle electromyography: assessing normal and neuropathic units. Muscle Nerve, 23: 1076–1083. Paintal, AS (1966) The influence of diameter of medullated nerve fibers of cats on the rising and falling phases of the spike and its recovery. J. Physiol., 184: 791–811. Panescu, D, Cohen, KP, Webster, JG and Stratbucker, RA (1993) The mosaic electrical characteristics of the skin. IEEE Trans. Biomed. Eng., 40: 434–439. Pease, WS and Bowyer, BL (1988) Motor unit analysis. Comparison between concentric and monopolar electrodes. Am. J. Phys. Med. Rehabil., 67: 2–6. Pease, WS, Fatehi, MT and Johnson, EW (1989) Monopolar needle stimulation: safety considerations. Arch. Phys. Med. Rehabil., 70: 412–414. Pease, WS and Pitzer, NL (1990) Electronic filter effects on normal motor and sensory nerve conduction tests. Am. J. Phys. Med. Rehabil., 69: 28–31. Perreault, EJ, Hunter, IW and Kearney, RE (1993) Quantitative analysis of four EMG amplifiers. J. Biomed. Eng., 15: 413–419. Rosenfalck, A (1978) Early recognition of nerve disorders by near-nerve recording of sensory action potentials. Muscle Nerve, 1: 360–367. Russell, JW, Karnes, JL and Dyck, PJ (1996) Sural nerve-myelinated fiber density differences associated with meaningful changes in clinical and electrophysiologic measurements. J. Neurol. Sci., 135: 114–117. Schmelz, M, Schmidt, R, Bickel, A, Handwerker, HO and Torebjork, HE (1997) Specific C-receptors for itch in human skin. J. Neurosci., 17: 8003–8008. Schmelz, M, Schmidt, R, Weidner, C, Hilliges, M, Torebjork, HE and Handwerker, HO (2003)

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Chemical response pattern of different classes of C-nociceptors to pruritogens and algogens. J. Neurophysiol., 89: 2441–2448. Schoonhoven, R and De Weerd, JP (1984) On the optimal choice of a recording electrode in electroneurography. Electroencephalogr. Clin. Neurophysiol., 58: 308–316. Seo, JH and Oh, SJ (2002) Near-nerve needle sensory conduction study of the medial calcaneal nerve: New method and report of four cases of medial calcaneal neuropathy. Muscle Nerve, 26: 654–658. Smith, T (1998) Near-nerve versus surface electrode recordings of sensory nerve conduction in patients with carpal tunnel syndrome. Acta Neurol. Scand., 98: 280–282. Stegeman, DF, Dumitru, D, King, JC and Roeleveld, K (1997) Near- and far-fields: source characteristics and the conducting medium in neurophysiology. J. Clin. Neurophysiol., 14: 429–442. Swain, ID, Wilson, GR and Crook, SC (1985) A simple method of measuring the electrical resistance of the skin. J. Hand Surg. [Br], 10: 319–323. Tjon, ATAM, Lemkes, HH, van der Kamp-Huyts, AJ and van Dijk, JG (1996) Large electrodes improve nerve conduction repeatability in controls as well as in patients with diabetic neuropathy. Muscle Nerve, 19: 689–695. Torebjork, HE and Hallin, RG (1974) Responses in human A and C fibers to repeated electrical intradermal stimulation. J. Neurol. Neurosurg. Psychiatry, 37: 653–664.

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Trojaborg, W (1998) The concentric versus the monopolar needle electrode. The case for concentric needles. Muscle Nerve, 21: 1806–1808. Uludag, B, Koklu, F and On, A (1997) A new source of electromyographic artifact: mobile phones. Muscle Nerve, 20: 121–122. van Dijk, JG, Tjon-a-Tsien, A and van der Kamp, W (1995) CMAP variability as a function of electrode site and size. Muscle Nerve, 18: 68–73. Wee, AS and Ashley, RA (1990) Relationship between the size of the recording electrodes and morphology of the compound muscle action potentials. Electromyogr. Clin. Neurophysiol., 30: 165–168. Wee, AS, Leis, AA, Kuhn, AR and Gilbert, RW (2000) Anodal block: can this occur during routine nerve conduction studies? Electromyogr. Clin. Neurophysiol., 40: 387–391. Weidner, C, Schmelz, M, Schmidt, R, Hammarberg, B, Orstavik, K, Hilliges, M, Torebjork, HE and Handwerker, HO (2002) Neural signal processing: the underestimated contribution of peripheral human C-fibers. J. Neurosci., 22: 6704–6712. Wexler, I, Paley, D, Herzenberg, JE and Herbert, A (1998) Detection of nerve entrapment during limb lengthening by means of near-nerve recording. Electromyogr. Clin. Neurophysiol., 38: 161–167. Yamada, T, Machida, M, Oishi, M, Kimura, A, Kimura, J and Rodnitzky, RL (1985) Stationary negative potentials near the source vs. positive farfield potentials at a distance. Electroencephalogr. Clin. Neurophysiol., 60: 509–524.

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CHAPTER 6

Motor nerve conduction studies Kerry H. Levin* Cleveland Clinic Foundation, OH, USA

6.1. Introduction Motor nerve conduction studies constitute a major element of the electrodiagnostic examination. They can identify features of demyelination and axon loss, they can define defects of neuromuscular junction transmission, and they can reflect severe muscle fiber loss. Stimulation is applied at distal and proximal sites along a nerve trunk. Recording of the resulting action potential occurs with surface electrodes overlying a muscle belly innervated by the stimulated nerve (Fig. 6.1). Features of a motor conduction study that can be analyzed include the amplitude, area, and configuration of the response waveform, as well as its latency and conduction velocity. 6.2. The compound muscle action potential Since the active recording electrode overlies a muscle belly, the recorded response is a compound muscle action potential (CMAP). In comparison, the sensory nerve action potential (SNAP) is recorded directly from a nerve trunk. The CMAP represents a summation in the volume conductor of all the individual muscle fiber action potentials activated by the stimulus and within the pick up territory of the active recording electrode. Normally, the CMAP is a biphasic waveform with an initial negative deflection. This configuration results from placement of the active recording electrode at the motor point of the muscle belly, where the muscle fibers are initially depolarized and their action potentials generated. In comparison, the sensory nerve action potential is normally a triphasic waveform with initial positive deflection, because the propagated

* Correspondence to: Kerry H. Levin MD, Department of Neurology, Desk S-90, Cleveland Clinic Foundation, 9500 Euclid Avenue, Cleveland OH 44195, USA. E-mail address: [email protected]. Tel.: +1-216-444-8370; fax: +1-216-445-4653.

depolarization along the nerve trunk approaches the recording electrode from the stimulation site. The CMAP is measured in millivolts, compared to microvolts for a sensory action potential, because the size of an action potential is proportional to the diameter of the excitable tissue from which it is derived. A number of factors contribute to the size and shape of a normal CMAP, including the geographical relationship between the active and reference recording electrodes, active recording electrode size, the effect of recording distance on CMAP size, the effect of filtering, and the effect of temperature. 6.2.1. Relationship between recording electrodes and CMAP shape Manipulating the electrode placement in a differential amplifier recording system can affect the amplitude of the recorded potential. This is especially necessary when recording sensory nerve action potentials in the microvolt range. Because the CMAP size is measured in millivolts, it is not necessary to space the active and reference recording electrodes so that the potential at the reference electrode site adds amplitude to the waveform. Rather, by minimizing the reference electrode’s contribution to the CMAP response, variability in response size due to imprecise electrode placement can be theoretically minimized, and standardization of the technique improved. This has traditionally been accomplished by applying the reference electrode over the tendon distal to the muscle belly on which the active electrode is applied. Recent work has confirmed that the reference electrode may not be electrically silent in relation to the active electrode. This is especially true for the ulnar and tibial motor nerve conduction studies, where the traditional reference electrode position can contribute a greater amount toward the main negative peak of the CMAP than does the active electrode (Kincaid et al., 1993; Brashear and Kincaid, 1996). This effect correlates with the double-peak appearance of the ulnar and

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Distance = 270 mm

7 mv

S

CV = 270 = 54 m/s 5

A 8.5 ms

Fig. 6.1 Median motor nerve conduction study, after stimulation at the elbow (A), and wrist (B). The measured distance between the two stimulation sites is 270 mm. (From Levin, 1993. Common focal mononeuropathies and their electrodiagnosis. J. Clin. Neurophysiol. 10: 181–189, reproduced with permission)

6.5 mV

S

B

3.5 ms

tibial CMAP. In comparison, the reference electrodes do not significantly contribute to the main negative component of the median and peroneal CMAPs. The large reference electrode contribution to the tibial response may result from the large amount of tibial muscle mass in the foot and its wider geographical distribution in the foot, compared to peroneal musculature. Far-field muscle potentials may also be contributing to this effect (Dumitru and King, 1991). An attempt to neutralize the effect of the reference electrode by placing it on the contralateral limb leads to further complications. With that montage, short latency components are recorded at the onset of the tibial CMAP when proximal stimulation studies are performed. These extra potentials are thought to be farfield components generated from muscle action potentials in the distal leg. They do not appear when the active and reference electrodes are ipsilateral, due to common mode rejection (Brashear and Kincaid, 1996). The size and shape of the ulnar CMAP obtained with the active electrode overlying the hypothenar region depend on the action potentials from all ulnar innervated hand muscles. Stimulating individual muscles, McGill and Lateva showed that the first peak of the negative phase of the CMAP is derived from hypothenar muscles, while the second peak is due to a volume conducted potential from interosseus muscles (McGill and Lateva, 1999). This contribution was altered by changing the relative positioning of the fingers, and could be affected by the presence of a temperature gradient in the hand. A reduction in the

interosseus contribution was obtained with the use of a combined reference from two electrodes, one on each tendon of the hypothenar muscle group. (Fig. 6.2) Muscle fiber length and muscle position have an effect on the shape of the CMAP. Examining the thenar CMAP, Lateva and colleagues showed that changes in CMAP shape and amplitude with thumb abduction are due to changes in termination time (point at which the muscle fiber action potential reaches the muscle-tendon junction) resulting from changes in muscle fiber lengths (Lateva et al., 1996). (Fig. 6.3) 6.2.2. Relationship between electrode size and CMAP There is a complicated relationship between the recording electrode size CMAP characteristics. This topic is treated in detail in Chapter 5. One study showed an advantage to the use of a larger electrode size (comparing a standard 10 mm diameter tin disc electrode with a disposable adhesive ground electrode measuring 2.9 × 2.65 cm) in regard to inter-operator and inter-study reproducibility of results (Tjon-ATsien et al., 1996). Their study did not show a significant change in CMAP amplitude, but larger electrodes tended to produce slightly longer distal latencies and slower mean conduction velocities. However, other authors identified the opposite; over the range of round disc electrode diameter from 4 to 14 mm, there was a 10% decrease in amplitude but no change in latency,

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141

b/dt

b/dt

A

b/pt

b/pt

b/br

b/br

C

Neutral d2-d4 flexed

dt

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b/dt

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b/pt

b/pt

b/br subj KM.I

B

4 mV

b/br subj SK.r

D

b

− +

b/br

pt

E

Fig. 6.2 Cancellation of the interosseus contribution using a balanced reference. (A, C) Sums of the directly stimulated interosseus signals recorded by the b/dt and b/pt electrode configurations, and the “balanced reference” signal (b/br) formed by adding 1/3 times the signal to 2/3 times the b/pt signal, for two subjects. The interosseus contributions are almost completely canceled in the b/br signals. (B, D) The effects of length manipulation (solid line = neutral, dashed line = flexion of digits 2–4) are reduced in the b/br CMAP compared to the b/dt and b/pt CMAPs. (E) Electrode configuration for recording the balanced-reference signal. Reference electrodes are placed over both the distal and proximal tendons and are connected in parallel. The distal electrode has been cut in half to give an approximate 1 to 2 ratio between the dt and pt contributions (From McGill and Lateva, 1999, reproduced with permission from John Wiley and Sons, Inc.).

simultaneously electrically evoked components of the CMAP, due to different rates of conduction along motor nerve fibers (Schulte-Mattler et al., 2001). The lesser degree of temporal dispersion seen in CMAPs compared to SNAPs is attributed to the narrower range of conduction velocities along motor nerve fibers in a nerve trunk, and the broader duration of CMAPs (Fig. 6.5). Based on a mathematical model of the CMAP as a sum of asynchronous single nerve fiber action potentials, it has been estimated that there may be a 25-m/s difference between fastest and slowest fibers in a sensory nerve trunk, versus 11 m/s for motor fibers (Dorfman, 1984). For a SNAP, the lower

when recording over the abductor digiti minimi (Wee and Ashley, 1990). 6.2.3. Effects of distance from point of stimulation to recording site The CMAP generated at a proximal site of stimulation differs modestly from the CMAP obtained with distal simulation. (Fig. 6.4) The measurable differences include reduced amplitude and area, increased duration, and shallower initial negative slope of the proximally generated CMAP. These changes result from temporal dispersion, or desynchronization, of the

Abducted 90⬚ Abducted 45⬚ Relaxed Extended

2 mV 2 ms pt/p (x5)

tt/t(x10) b/t

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subj FS

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Fig. 6.3 CMAPs recorded at different thumb positions (extended, relaxed, abducted by 45˚, and abducted by 90˚) for 2 subjects (From Lateva and McGill, 1996, reproduced with permission from John Wiley and Sons, Inc.).

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Fig. 6.4 Ulnar CMAPs recording from the hypothenar eminence, resulting from stimulation at the wrist (1), below the elbow (2), above the elbow (3), at the axilla (4), and at the supraclavicular site (5). Vertical division = 5 mV; horizontal division = 5 ms.

amplitude obtained after proximal stimulation is likely related to physiological phase cancellation due to the velocity-related temporal dispersion of sensory action potentials over a nerve segment and the short duration

of sensory action potentials (Kimura et al., 1986). (Figs. 6.6 and 6.7) For motor NCS in normal subjects, most laboratories expect less than 20% decrease in the amplitude between the distal stimulation site and

Fig. 6.5 Median sensory nerve action potentials recording from digit 2, resulting from stimulation at the wrist (1), above the elbow (2), at the axilla (3), and at the supraclavicular site (4). Vertical division = 20 μV; horizontal division = 2 ms.

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Individual responses

Summated responses

F

S

F

F

S

Fig. 6.6 A model for phase cancellation between fast (F) and slow (S) conducting sensory fibers. With distal stimulation, two unit discharges summate in phase to produce a sensory action potential twice as large. With proximal stimulation, a delay of the fiber causes phase cancellation between the negative peak of the fast fiber and positive peak of the slow fiber, resulting in a 50% reduction in size of the summated response (From Kimura et al., 1986, with permission.).

elbow or knee, with the exception of the tibial study, where there can be up to 50% decrease in amplitude with proximal stimulation. The degree of temporal dispersion in a CMAP is related to the length of the conducted segment and the range of conduction velocities in the nerve trunk. Therefore, the impact will be greater along nerves in the leg, especially the tibial nerve. Using clinical measurements and Fourier analysis from 86 subjects, Schulte-Mattler et al., found that amplitude loss was most marked for the tibial (36%) and ulnar (21%) nerves, area loss for the median (12%) and ulnar (18%) nerves, and duration prolongation for the tibial (32%) and ulnar (17%) nerves (Schulte-Mattler et al., 2001). None of these findings was age dependent. Their data showed that the dependence of these variables on length was approximately linear. In a disease

state, the range between fastest and slowest motor fibers increases, and phase cancellation becomes more prominent along motor nerves (Lee et al., 1975). 6.2.4. Effects of filtering on the compound muscle action potential Electrophysiological recordings obtained from a differential amplifier can be altered by manipulating the bandwidth of frequencies allowed to contribute to the recording. Filter settings are chosen to maximize the contribution of frequencies close to the recording electrodes and minimize contamination of the recording by remote generators. Because amplitude, latency, and waveform configuration change as filter settings are altered, standard settings are required.

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Summated response

F

S

F S

Fig. 6.7 Same arrangement as in Fig. 6.6 to show the relationship between fast (F) and slow (S) conduction motor fibers. With distal stimulation, two unit discharges representing motor unit potentials summate to produce a muscle action potential twice as large. With proximal stimulation, long-duration motor unit potentials still superimpose nearly in phase despite the same latency shift of the slow motor fiber as the sensory fiber shown in Fig. 6.6. Thus, a physiological temporal dispersion alters the size of the muscle action potential only minimally, if at all (From Kimura et al., 1986, with permission.)

Increasing the low frequency filter setting tends to reduce CMAP amplitude, peak latency, and total waveform duration (Fig. 6.8). These effects are related to the filtering out of important low frequency components contributing to the overall size of the CMAP waveform. There is no effect on the onset latency of the CMAP (Dumitru and Walsh, 1988). Decreasing the high frequency filter tends to increase peak and onset latencies, and increase the negative spike component of the CMAP (Fig. 6.9). Onset latency becomes delayed because highfrequency components over a short duration at the onset of the waveform are filtered out, causing a “drop out” of the first part of the potential. There is little effect on the amplitude down to a frequency of 500 Hz. 6.2.5. Effects of temperature on the compound muscle action potential A decrease in muscle temperature corresponds with increased CMAP amplitude, duration, rise time, and area. Local cooling leads to increased voltage of muscle fiber action potentials (Ricker et al., 1977). This is the result of delayed voltage-gated sodium channel inactivation, leading to excess depolarization of the muscle fiber membrane and, subsequently, increased propagated action potential duration, spike height, and area (Scheopfle and Erlanger, 1941).

6.2.6. Sources of error in compound muscle action potential recordings Of all the motor NCS parameters, CMAP amplitude appears to have the least reproducibility (Bleasel and Tuck, 1991; Claus et al., 1993; van Dijk et al., 1994, Kohara et al., 2000). While mean motor nerve conduction velocities are highly reproducible, the coefficient of variation for CMAP amplitude and latency has ranged from 8.5–46% (Tjon-A-Tsien et al., 1996). Inability to reproducibly localize the optimum recording site appears to be the main cause of variability. Van Dijk et al. showed that a variability of 5–10 mm in the recording site could have a profound effect on CMAP amplitude (van Dijk et al., 1994). Variability in CMAP amplitude and latency recordings is even higher in patients with diabetic neuropathy (Tjon-A-Tsien et al., 1996). Other sources of error can be identified in the routine laboratory on a daily basis, although their contributions have not been studied scientifically. Submaximal stimulus intensity, imprecise stimulator placement, stimulus spread to adjoining nerve trunks, and temperature fluctuations all contribute to CMAP recording variability and error. The normal CMAP amplitude, configuration, or both may be altered in the presence of anomalous innervation. In the case of the median-to-ulnar

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145 Fig. 6.8 Ulnar CMAPs recording from the hypothenar eminence, after stimulation at the wrist. Low filter setting is 2 Hz (1), 10 Hz (2), 20 Hz (3), 50 Hz (4), and 500 Hz (5). The high filter setting remains constant at 10 KHz. Vertical division = 10 mV; horizontal division = 5 ms.

crossover in the forearm (estimated to occur in 5–30% of arms), ulnar motor nerve fibers reside in the median nerve at the elbow, but switch back into the ulnar nerve trunk somewhere in the forearm. This leads to co-stimulation of median and ulnar motor fibers at the elbow but not at the wrist, giving rise to disparity between the CMAP waveforms at the two sites of stimulation (Chapter 14). 6.3. Motor conduction latency and velocity The speed with which an electrical impulse passes along a motor nerve trunk is a composite of the speeds of individual motor nerve fiber action potential propagations to the recorded muscle motor point. These are, in turn, related to the diameter of nerve fibers, temperature, stimulation intensity, and age.

6.3.1. Effects of temperature on motor conduction velocity Temperature has a marked effect on nerve conduction (see above). Nerve conduction velocity decreases in proportion to the decrease in temperature, without a significant influence from nerve fiber size. Conduction velocity can be measured to decline to as low as 1–2% of normal velocity, before conduction fails entirely at 7–8˚C (Paintal, 1965). The degree of change in conduction velocity with progressive change in temperature for upper extremity nerve trunks has been defined (Halar et al., 1980, 1983). For the median motor conduction velocity, there is a 1.5 m/s decline for every 1˚ C temperature decrease. The median motor distal latency increases by 0.2 m/s for each 1˚ C decrease in temperature. The

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Fig. 6.9 Ulnar CMAPs recording from the hypothenar eminence, after stimulation at the wrist. The response with a latency of 2.8 ms corresponds to a high filter setting of 10 KHz, 2.9 ms corresponds to 2 KHz, 3.0 ms corresponds to 1 KHz, and 3.2 ms corresponds to 500 Hz. The low filter setting remains constant at 2 Hz. Vertical division = 5 mV; horizontal division = 2 ms.

ulnar motor conduction velocity decreases by 2.1 m/s for every 1˚ C temperature decrease, and the ulnar motor distal latency increases by 0.2 m/s for each 1˚C decrease in temperature. 6.3.2. Effects of age on motor conduction velocity Peripheral nerves at birth are not fully myelinated. Motor nerve conduction velocities in the neonate are about one half the adult values. The developmental process of myelination continues until the age of 3 to 5 years, when conduction velocities approach adult values (Wagner and Buchthal, 1972). Over the age of 50 years, conduction velocity declines by 1–2 m/s per decade. There is an associated progressive increase in distal latency. 6.3.3. Sources of error in obtaining motor conduction velocity A number of sources of error exist in the performance of NCS that can lead to spurious conduction velocities. Foremost among these is measurement of the dis-

tance over which the conduction is occurring. Landmarks easily shift on extremities, giving rise to significant variation in the same patient and among patients. These errors tend to be amplified over shortconduction distances (since the error is a larger proportion of the whole distance), and when conduction velocities are higher (Maynard and Stolov, 1972). Anatomy and nerve trunk position issues also predispose to a lack of reliability. When analyzing nerve conduction around a joint (classically the ulnar nerve as it courses around the elbow), the conduction distance varies according to whether the joint is flexed (the nerve is stretched to its fullest extent and lengthened) or extended (the nerve is kinked on itself and shortened). However, the latency of conduction remains the same, so that in the flexed elbow position, the calculated conduction velocity will be slower than in the extended elbow position. Stimulus spread can introduce error in the conduction velocity measurement. When the stimulus intensity is increased beyond the ideal supramaximal level, a shift in the location of nerve trunk depolarization can

MOTOR NERVE CONDUCTION STUDIES

change by 3–5 millimeters (Buchthal and Rosenfalck, 1966). If the stimulus spread occurs in the direction of the recording electrode, the true conduction distance is shorter than the measured distance, leading to a falsely increased conduction velocity. A theoretical consideration is the effect of passive lengthening or shortening of muscle fibers on latency and velocity of conduction. Measuring over a segment of a muscle fiber, Trontelj used single fiber recordings to show that stretching a muscle fiber could reduce its conduction velocity by 22%, while shortening the muscle fiber could increase the velocity by up to 33% (Trontelj, 1993). In another study, intramuscular stimulation of nerve fibers in the biceps brachii showed no significant change in impulse conduction velocity with muscle fiber length change when recording over the center fiber length, but a decrease in latency by as much as 24% occurred with muscle fiber shortening (Brown et al., 1996). In shortened muscle, the plasmalemma of a muscle fiber is folded and there are inpouchings of membrane known as caveolae (Dulhunty and Franzini-Armstrong, 1975). This may explain why the time taken for impulses to travel between individual sarcomeres, and to reach the end of a muscle is least in the shortened position. 6.4. Performing routine motor nerve conduction studies 6.4.1. Overview Nerve conduction studies and the needle electrode examination constitute the major components of the electrodiagnostic examination. In most clinical laboratories, the NCS are performed first, and guide the physician in the approach to the needle electrode examination. NCS require no active participation from the patient. Although the electrical stimulations are uncomfortable, they are tolerable to all but the most sensitive patients. They are indispensable for the electrodiagnosis of focal nerve lesions in the extremities and are the only way to gain physiological confirmation of the presence of demyelination. There are a number of limitations regarding the use and interpretation of motor NCS. First, very proximal portions of nerve trunks cannot be reliably accessed for stimulation. Second, because of unavoidable technical imprecision in performing motor NCS and because of a wide range of normal for CMAP amplitudes, detection of motor axon loss by NCS is relatively insensitive, unless over 50% of

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nerve fibers have been lost. Comparison with the contralateral limb can improve the sensitivity, if the condition is unilateral. Third, motor NCS are also relatively insensitive to the presence of muscle fiber loss in myopathy. Individual clinical EMG laboratories differ in the technical aspects of performing motor nerve conduction studies. However, the basic approaches are similar in most laboratories. The following descriptions are considered standard techniques. 6.4.2. Setting up the electrodiagnostic machine Machine settings must be adjusted for proper motor nerve conduction study results. Settings to be adjusted include the filters, sweep speed, and gain. Filter settings for motor NCS should allow a wide recording bandwidth. The low filter setting is usually set at 2 Hz, and the high filter setting is 10 KHz. Because the CMAP response is large (measured in millivolts), the effect of contamination from CMAPs of neighboring muscle bellies is less important. Thus, the objective is to include as many of muscle fiber action potentials as possible in the summated CMAP of the recorded muscle. For each nerve trunk stimulation, the sweep speed needs to be adequate in order to capture the produced CMAP. Reliable measurement of the latency depends on a precise waveform offset from baseline. The visual characteristics of that take-off point change with changes in the sweep speed. Thus, in order to optimize reproducibility of technique, the sweep speed should be fast enough to capture CMAPs at all planned points of stimulation. Normal distal motor latencies recorded after stimulation at the wrist or ankle range from 3 to 6 ms in normal adults. In the arm, latencies measured at the hand muscles after stimulation at the elbow and supraclavicular sites of stimulation range from 6 to 10 ms, and 13 to 15 ms, respectively. In the leg, latencies measured at the foot muscles after stimulation at the knee range from 8 to 10 ms in normal adults. These values correlate directly with the length of the arm and leg. In disease states, latencies may increase by a factor of two or more. When no motor response is recorded after nerve stimulation, it is important to increase the sweep speed to identify the presence of a pathologically slowed response. The gain setting should also be chosen to maximize CMAP visibility and accuracy of measurement of amplitude and latency. Like the sweep speed, the gain

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setting should be the same for all stimulation points of a given nerve trunk. This will avoid inaccuracies in the measurement of latency. Increasing the gain tends to increase onset latency (Dumitru and Walsh, 1988) (Table 6.1). 6.4.3. Stimulus guidelines and pitfalls For motor NCS, stimulus duration should be 0.2 ms initially. The stimulus voltage should be increased gradually until there is no further increase in CMAP amplitude. At that point, the stimulus voltage should be increased by another 10–20%, the so-called supramaximal stimulus intensity. If a maximum CMAP is not achieved, the duration should be increased, and the process repeated. Failure to use a supramaximal stimulus will result in the possibility of a less than maximal CMAP amplitude. On the other hand, excessive stimulation increases the possibility that nearby nerve trunks will be co-stimulated, leading to a falsely increased CMAP amplitude or a spurious waveform configuration. Co-stimulation is likely to occur between the median and ulnar nerve trunks at the wrist and above the elbow; among the median, ulnar, and radial nerve trunks at the axilla; and between the peroneal and tibial nerve trunks in the popliteal fossa. Excessive stimulation can also lead to stimulus jump beyond the region immediately under the stimulating electrodes. In general both the cathode and anode of the stimulator should be placed on the nerve trunk, with the cathode distal (and closest) to the recorded muscle. The anode may be swiveled off the nerve trunk when there is excessive shock artifact and when the usual placement causes direct muscle belly stimulation, as is the case when stimulating the facial nerve and recording over the nasalis muscle. Table 6.1 Effect of gain setting on the CMAP latency after median nerve stimulation at the wrist, recording over the abductor pollicis brevis muscle* Gain setting (mV/division)

Onset latency (m/s)

Amplitude (mV)

500 1000 5000 10 000

3.4 3.5 3.7 3.8

23 23

*The high and low frequency filters remained constant at 10 000 Hz and 10 Hz, respectively (Dumitru and Walsh, 1988).

Points of stimulation along a nerve trunk are limited to sites close to the skin surface. Routinely, these sites are at the wrist, elbow, ankle, and popliteal fossa. Nerve trunks can also be stimulated in the axilla and in the supraclavicular fossa. Specific nerve trunks are also accessible, such as the phrenic and spinal accessory nerves in the neck, and the facial and trigeminal nerves face. Needle electrodes used for stimulation may be able to reach some deep nerves, such as extra-spinal nerve roots. 6.4.4. Electrode placement and pitfalls Electrodiagnostic standard practice requires that the active recording electrode be placed over the motor point of the muscle belly innervated by the nerve trunk being stimulated. This location is identified by moving the electrode with each stimulus until a CMAP of maximal amplitude and initial negative deflection occurs. This task is relatively straightforward in the distal hand and foot muscles. The reference electrode is traditionally placed distal to the active recording electrode, on a relatively electrically silent structure such as a muscle tendon. As noted above, in actuality there is no electrically inactive site. The reference electrode is especially likely to contribute to the ulnar and tibial CMAP waveform appearance (Brashear and Kincaid, 1996). The task is more difficult when recording proximally (such as the forearm), where there are layers of overlapping muscles, either innervated by the same nerve trunk, or innervated by a nerve trunk that may be co-stimulated with the nerve trunk under investigation. In either case, there is a potential for generation of a spurious CMAP waveform that has an initial volume-conducted positive deflection, a falsely enlarged amplitude, or an altered configuration. A spurious CMAP may also be generated from hand muscles, such as the thenar group, when stimulating proximally at the axilla and supraclavicular site, where co-stimulation of the ulnar nerve is sometimes unavoidable. In this setting, volume-conducted components from closely positioned ulnar innervated muscles contaminate the thenar (median-derived) CMAP. Contamination of the thenar recording may be reduced by selectively recording from the thenar muscle group with an intramuscular needle electrode, thus decreasing the recording distance and minimizing the distant volume–conducted contributions. When stimulating the ulnar nerve proximally and recording from the hypothenar group in the hand, there is less chance of contamination from median nerve costimulation, since there are no large median innervated muscles in close proximity to the hypothenar group.

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6.4.5. Calculating the conduction velocity Conduction velocity can be calculated easily after stimulating at two sites along a nerve trunk. The distal latency is the time elapsed from the point of distal (wrist or ankle) stimulus to the time of initial CMAP onset. This value is subtracted from the latency obtained when stimulating at the proximal site and recording from the muscle belly. After measuring the distance between the proximal and distal stimulation sites, it is simply a matter of completing the equation v = distance/time. The resulting value represents the conduction velocity in the forearm and foreleg, since the distal conduction has been subtracted out (Fig. 6.1). When a nerve trunk remains superficial over a whole segment of its length, it is possible to sequentially stimulate at short intervals. Examples include the median nerve at the wrist and palm, and the ulnar nerve across the elbow. Segmental conduction studies can be performed along short segments, as the stimulator is moved at 1 cm increments along the nerve, a technique described as inching. Inching has the potential to localize a focal nerve lesion (site of compression, entrapment, or focal demyelination) by demonstrating slowing or amplitude drop over a short nerve segment (Chapter 8). In general, a distal conduction velocity (from the distal-most site of stimulation to the muscle belly recording electrode site) is not calculated, because the latency of conduction over that segment includes conduction time along the small-diameter terminal axon branches, the neuromuscular junction transmission time, the time to depolarize the muscle membrane, and the time to generate a muscle fiber action potential. The distal latency is, therefore, longer than the latency over the same distance along a more proximal segment of the nerve. The difference between the distal latency and a more proximal latency over the same distance is described as the residual latency, and reflects the conduction time through terminal axons and the neuromuscular junction transmission time. Representative normal limits of some motor nerve conduction studies are displayed in Table 6.2. 6.4.6. Practical guidelines A number of practical guidelines will improve the reliability and sensitivity of motor NCS. At the hand the skin temperature should be maintained at 35˚C or above; at the foot the temperature should be at least 32˚C. When applying stimulation, it is critical to

149 Table 6.2 Normal limits of motor nerve conduction studies in a 40-year-old

Median Ulnar Peroneal Tibial

Amplitude Distal (mV) Latency (ms)

Conduction velocity (m/s)

>6 >7 >4 >6

>50 >50 >40 >40

<3.9 (8 cm distance) <3.0 (8 cm distance) <5.5 (5 cm distance) <5.8 (8 cm distance)

observe the degree and distribution of muscle contraction. This assures that adequate stimulation is being applied and that the expected muscle contraction is occurring. Co-stimulation of neighboring nerve trunks will be observable by visual inspection of the muscle groups contracting. The failure to achieve a normal response requires a careful search for technical causes. These include over or under stimulation, off-nerve stimulation, improper recording electrode placements, and improper set up of the voltage and/or duration of stimulus. Any response obtained that is considered abnormal must be compared with the response on the opposite side when the symptoms are unilateral. When a response is at the lower end of the normal range, it is appropriate to study the opposite side, looking for subtle pathology that may not constitute an abnormality based its absolute value. 6.4.7. Advanced techniques Several uncommonly used techniques may be useful in special situations. With motor NCS a needle electrode can at times be useful, both for stimulation and recording. A needle electrode can be used to penetrate deep to the skin surface to stimulate a nerve trunk not accessible with percutaneous stimulation. Use of a needle electrode to stimulate the median or ulnar nerve branches in the forearm can provide an additional distal or proximal site of stimulation to confirm the precise location of a conduction block. A needle electrode can also be used for recording purposes. In the setting of severe nerve trauma, a CMAP from an involved muscle may not be obtainable using disc electrodes applied to the overlying skin. A needle electrode within the muscle may be able to record small CMAP responses otherwise not obtainable. CMAPs can be obtained after stimulation from sites proximal to the peripheral nervous system.

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Trans-cranial magnetic stimulation produces recordable muscle potentials with latencies that reflect transmission along central and peripheral motor pathways (see Chapter 19). Intra-operatively electric motor evoked potentials provide similar information when used for cortical and spinal cord stimulation. 6.5. Basic patterns of pathology 6.5.1. Electrodiagnostic correlates of weakness Although a number of symptoms bring patients to the electrodiagnostic laboratory, weakness is the primary symptom being explored with motor NCS. Among all the motor parameters examined, CMAP amplitude is the only one that correlates closely with weakness. While changes in conduction velocity and latency may be seen in patients who are weak, weakness is directly related to interruption of impulses along motor nerves, leading to failure to depolarize muscle fibers. Established axon loss of motor nerve fibers produces CMAPs of reduced amplitude at all points of stimulation along the nerve, irrespective of the location of initial injury to the nerve fibers. Conduction block is the result of demyelination along motor nerve fibers to the extent that essentially all action potential propagation across the demyelinated

segment ceases, even though the underlying motor axons remain intact. The result of either axon loss or conduction block along a nerve fiber is the failure of action potential arrival at the nerve terminal and, ultimately, failure of muscle fiber contraction. Less complete demyelination along a nerve fiber may cause disruption of saltatory conduction without complete interruption of action potential transmission. The result is slowing of conduction, but the action potential reaches the nerve terminal, and ultimately its muscle fiber contracts. Thus, slowing of conduction velocity and prolongation of latency do not by themselves cause weakness. 6.5.2. Evolution of CMAP loss in an acute axon loss process After nerve transection, a series of pathological events takes place culminating in Wallerian degeneration of the nerve fibers. In spite of complete disconnection of the distal nerve fibers from their proximal portions and their anterior horn cells, and also in spite of cessation of axonal transport, the distal nerve fiber membranes can still be depolarized and action potential propagation continues for some days. Progressively, nerve fiber disintegration and neuromuscular junction transmission failure result in loss of nerve excitability and muscle fiber depolarization.

Injury 100

80

60 NCS amp. (%)

40

20

0

0

1

2

3

Motor and sensory, stimulating proximal to lesion

6 4 5 Days post-injury

7

8

Motor, stimulating distal to lesion

9

10

11

12

Sensory, stimulating distal to lesion

Fig. 6.10 The time course of changes in the CMAP and sensory nerve action potential amplitudes following a severe axon loss lesion (From Wilbourn, AJ, 1986 AAEE Case Report #12: Common peroneal mononeuropathy at the fibular head. Muscle Nerve 9: 825–836, reproduced with permission from John Wiley and Sons, Inc.)

MOTOR NERVE CONDUCTION STUDIES

After a severe proximal motor nerve trunk transection, with stimulation of the nerve distal to the point of transection, the CMAP amplitude remains normal until day 3 after transection. From that point, until day 5–8, there is a nearly linear decline in the evoked CMAP amplitude until the lowest amplitude is obtained, determined by the proportion of motor nerve fibers transected (Wilbourn, 1986) (Fig. 6.10). During this process, there is no significant change in the motor latencies or conduction velocities. In studies of progressive motor neuron disease of the ALS type (amyotrophic lateral sclerosis), Lambert demonstrated that the mean ulnar motor conduction velocity of 322 patients did not fall below 50 m/s until >75% of CMAP amplitude had been lost, corresponding to loss of the last fastest-conducting motor nerve fibers. (Lambert, 1962) In comparison, the sensory nerve action potential amplitude falls at a different rate after nerve fiber transection. With stimulation and recording distal to the point of nerve transection, the sensory nerve action potential amplitude remains normal until day 5. From that point, until day 8–10, there is a nearly linear decline in the evoked sensory nerve action potential amplitude until the lowest amplitude is obtained, determined by the proportion of sensory nerve fibers

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transected (Wilbourn, 1986) (Fig. 6.10). Decline of the sensory nerve action potential amplitude reflects failure of nerve fiber action potential propagation alone, since the response is recorded from the nerve fibers themselves. The faster failure of motor conduction as measured from the CMAP likely reflects earlier failure of neuromuscular junction transmission. 6.5.3. Patterns of abnormality on routine nerve conduction studies Disorders affecting peripheral nerves may be primarily axon loss in etiology, primarily demyelinating, or mixed. Motor NCS are a critical component of the assessment of nerve pathophysiology. With axon loss processes, the NCS pattern differs for acute and subacute/chronic lesions. With an acute complete transection of the median nerve trunk in the forearm of 3 days’ duration or less, stimulation of the median nerve at the elbow, recording over the thenar eminence will yield no CMAP response. With stimulation anywhere below the site of transection, the CMAP will have normal amplitude. After 8 days from the time of transection, stimulation of the nerve anywhere along its course will yield no CMAP response (Fig. 6.11).

Axon loss

Distal stimulation site

Proximal stimulation site

Normal

Acute discontinuity conduction block between proximal and distal stimulation sites

Axon loss anywhere along nerve, >8 days

Fig. 6.11 CMAPs with various axon loss lesions. The first waveform in each line results from distal motor nerve trunk stimulation, and the second waveform results from proximal nerve trunk stimulation.

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Demyelination Distal stimulation site

Proximal stimulation site

Normal or conduction block proximal to stimulation sites

Generalized, synchronized slowing

Segmental slowing between sites of stimulation

Focal conduction block distal to distal stimulation site

Focal conduction block between sites of stimulation

Fig. 6.12 CMAPs with various demyelinating lesions. The first waveform in each line results from distal motor nerve trunk stimulation, and the second waveform results from proximal nerve trunk stimulation.

With predominantly demyelinating processes, the NCS pattern varies according to the nature and severity of the demyelinating lesion. A focal conduction block (such as is seen with compressive radial neuropathy at the spiral groove) will resemble the pattern noted above in the acute stage of a partial or complete nerve trunk transection. A diffusely and homogeneously demyelinating process (such as Charcot–Marie–Tooth Disease Type 1a) will demonstrate relatively preserved CMAP configurations with prolonged latencies and slowed conduction velocities. A non-uniform, segmental demyelinating process (such as acute inflammatory demyelinating polyneuropathy; i.e., Guillain–Barré syndrome) may demonstrate conduction block and variable slowing within the same nerve trunk, resulting in CMAPs with reduced amplitude and dispersed configuration at proximal sites of stimulation (Fig. 6.12). Focal or generalized slowing of conduction as the sole manifestation of a peripheral nerve disorder is uncommon. Median neuropathy at or distal to the wrist (carpal tunnel syndrome) of mild-moderate severity may be the only common example of this pattern. Mixed patterns of demyelination and axon loss are common. This is because: (1) progressive demyelination is often associated with damage to the underlying axons; and (2) axon loss processes develop electrophysiological features of demyelination as a reflection of nerve regeneration. Some axon loss disorders, such as diabetic polyneuropathy, have an electrophysiological signature which includes mild features of demyelination. Focal entrapment of the peroneal nerve at the fibular head is an example of a focal nerve disorder that is characterized most commonly by the

Mixed axon loss and conduction block

Distal stimulation site

Proximal stimulation site

Normal

Axon loss with focal conduction block between sites of stimulation

Fig. 6.13 CMAPs with mixed axon loss and demyelinating lesions. The first waveform in each line results from distal motor nerve trunk stimulation, and the second waveform results from proximal nerve trunk stimulation.

MOTOR NERVE CONDUCTION STUDIES

presence of focal demyelinating conduction block across the fibular head, with associated features of axon loss. With NCS, peroneal motor CMAPs are reduced in amplitude at all sites of stimulation, but there is superimposed additional loss of amplitude with stimulation proximal to the fibular head (Fig. 6.13). References Bleasel, AF and Tuck, RR (1991) Variability of repeated nerve conduction studies. Electroencephalogr. Clin. Neurophysiol., 81: 417–420. Brashear, A and Kincaid, JC (1996) The influence of the reference electrode on CMAP configuration: leg nerve observations and an alternative reference site. Muscle Nerve, 19: 63–67. Brown, T, Galea, V and McComas, AJ (1996) Muscle shortening, response latency, and conduction velocity. Muscle Nerve, 19: 1493–1495. Buchthal, F and Rosenfalck, A (1966) Evoked action potentials and conduction velocity in human sensory nerves. Brain Res., 3: 1–122. Claus, D, Mustafa, C, Vogel, W, Herz, M and Neundorfer, B (1993) Assessment of diabetic neuropathy: definition of norm and discrimination of abnormal nerve function. Muscle Nerve, 16: 757–768. Dorfman, LJ (1984) The distribution of conduction velocities (DCV) in peripheral nerves: a review. Muscle Nerve, 7: 2–11. Dulhunty, AF and Frazini-Armstorng, C (1975) The relative contributions of the folds and caveolae to the surface membrane of frog skeletal muscle fibres at different sarcomere lengths. J. Physiol. (Lond.), 250: 513–539. Dumitru, D and King, JC (1991) Far-field potentials in muscle. Muscle Nerve, 14: 981–989. Dumitru, D and Walsh, NE (1988) Practical instrumentation and common sources of error. Am. J. Phys. Med. Rehabil., 67: 55–65. Halar, EM, DeLisa, JA and Brozovich, FV (1980) Nerve conduction velocity: relationship of skin, subcutaneous, and intramuscular temperatures. Arch. Phys. Med. Rehabil., 61: 199–203. Halar, EM, DeLisa, JA and Soine, TL (1983) Nerve conduction studies in upper extremities: skin temperature corrections. Arch. Phys. Med. Rehabil., 64: 412–416. Kimura, J, Machida, M and Ishida, T et al. (1986) Relation between size of compound sensory or

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muscle action potentials and length of nerve segment. Neurology, 36: 647–652. Kincaid, JC, Brashear, A and Markand, ON (1993) The influence of the reference electrode on CMAP configuration. Muscle Nerve, 16: 392–396. Kohara, N, Kimura, J, Kaji, R, Goto, Y, Ishii, J, Takiguchi, M and Nakai, M (2000) F-latency serves as the most reproducible measure in nerve conduction studies of diabetic polyneuropathy: multicenter analysis in healthy subjects and patients with diabetic polyneuropathy. Diabetologia, 43: 915–921. Lambert, EH (1962) Diagnostic value of electrical stimulation of motor nerves. Electroencephalogr. Clin. Neurophysiol., Suppl. 22: 9–16. Lateva, ZC and McGill, KC (1996) Anatomical and electrophysiological determinants of the human thenar compound muscle action potential. Muscle Nerve, 19: 1457–1468. Lee, R, Ashby, P, White, D and Aguayo, A (1975) Analysis of motor conduction velocity in human median nerve by computer simulation of compound action potentials. Electroencephalogr. Clin. Neurophysiol., 39: 225–237. Levin, KH (1993) Common focal mononeuropathies and their electrodiagnosis. J. Clin. Neurophysiol., 10: 181–189. Maynard, RM and Stolov, WC (1972) Experimental error in determination of nerve conduction velocity. Arch. Phys. Med. Rehabil., 53: 362–372. McGill, KC and Lateva, ZC (1999) The contribution of the interosseus muscles to the hypothenar compound muscle action potential. Muscle Nerve, 22: 6–15. Paintal, AS (1965) Block of conduction in mammalian myelinated nerve fibers by low temperature. J. Physiol., 180: 1–19. Ricker, K, Hertel, G and Stodieck, G (1977) Increased voltage of the muscle action potential of normal subjects after local cooling. J. Neurol., 216: 33-38. Scheopfle, GM and Erlanger, J (1941) The action of temperature on the excitability, spike height and configuration, and refractory period observed in the responses of single medullated nerve fibers. Am. J. Physiol., 134: 694–704. Schulte-Mattler, WJ, Müller, T, Georgiadis, D, Kornhuber, ME and Zierz, S (2001) Length dependence of variables associated with temporal dispersion in human motor nerves. Muscle Nerve, 24: 527–533.

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Tjon-A-Tsien, AML, Lemkes, HHPJ, van Der KampHuyts, AJC and van Dijk, JG (1996) Large electrodes improve nerve conduction repeatability in controls as well as in patients with diabetic neuropathy. Muscle Nerve, 19: 689–695. Trontelj, JV (1993) Muscle fiber conduction velocity changes with length. Muscle Nerve, 16: 506–512. van Dijk, JG, van der Kamp, W, van Hilten, BJ and van Someren, P (1994) Influence of recording site on CMAP amplitude and on its variation over a length of nerve. Muscle Nerve, 17: 1286–1292.

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Wagner, AL and Buchthal, F (1972) Motor and Sensory conduction in infancy and childhood: reappraisal. Dev. Med. Child Neurol., 14: 189–216. Wee, AS and Ashley, RA (1990) Relationship between the size of the recording electrodes and morphology of the CMAPs. Electromyogr. Clin. Neurophysiol., 30: 156–168. Wilbourn, AJ (1986) AAEM Case Report #12: common peroneal mononeuropathy at the fibular head. Muscle Nerve, 9: 825–836.

Peripheral Nerve Diseases Handbook of Clinical Neurophysiology, Vol. 7 J. Kimura (Ed.) © 2006 Elsevier B.V. All rights reserved

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CHAPTER 7

Sensory nerve conduction studies Tulio E. Bertorini* University of Tennessee Health Science Center and Wesley Neurology Clinic, TN, USA

1. Introduction Sensory nerve conduction studies (SNCS) are an integral element in clinical electrodiagnostic testing of peripheral nerve disorders, as they provide useful and direct information of the population and function of the sensory axons and their involvement in disease processes. Conventional SNCS using surface electrodes were performed initially by Dawson in 1956, (Dawson, 1956) although he had previously recorded sensory nerve action potentials (SNAPs) by stimulating fingers immersed in a beaker of saline solution (Dawson, 1950). Even earlier, in 1949, Dawson and Scott had recorded a mixed nerve conductions in the arm (Dawson and Scott, 1949). Later, Gilliatt and Sears documented abnormalities of SNCS in peripheral neuropathy, demonstrating an absence of the SNAP in half of their patients (Gilliatt and Sears, 1958). In vitro nerve conduction tests were first studied in surgically excised nerves by investigators who determined the velocities of the different nerve fiber types (Collins et al., 1960; Hensel and Bowman, 1960). Also, in elegant studies, Dyck, Lambert and collaborators compared the characteristics of the various components of the in vitro SNAPs with the results of the histological findings in peripheral neuropathy (Dyck et al., 1971; Dyck and Lambert, 1974). Other researchers studied the in vivo SNAP of the sural nerve, and compared the findings with morphological analysis of their biopsies, determining that the main component of the SNAP was produced by the summation of action potentials of large myelinated nerve fibers, providing a direct assessment of their popu* Correspondence to: Tulio E. Bertorini, MD, Professor of Neurology and Pathology, University of Tennessee Health Science Center and Wesley Neurology Clinic, PC, Memphis, TN, USA. E-mail address: [email protected]. Tel.: +1-901-725-8920; fax: +1-901-725-8934.

lation (Buchthal and Rosenfalck, 1971; Buchthal et al., 1971 and 1978; Tackmann et al., 1977). There are several important differences between SNCS and motor nerve conduction tests. One is that the SNAP recorded with surface electrodes is of three orders of magnitude smaller than the compound motor action potentials (CMAP) obtained by motor nerve stimulation. This is because the SNAP represents the summation of the action potentials of the large myelinated axons of the studied nerve, whereas the CMAP is formed by the summation of the individual action potentials of the many muscle fibers from the motor units recorded in the studied muscle. Another difference is that in motor conduction tests, the measured latencies include the physiological delay at the neuromuscular junction, for this reason their conduction velocity (CV) is calculated by subtracting the difference in the latencies from two stimulating sites and not from the stimulus to the response. The sensory CV determines the velocity only of the stimulated axons, and thus, this is calculated not only from the difference in latencies at two stimulation sites, but also and more often, directly from the stimulus to the SNAP. In routine clinical testing, sensory CV is measured by the use of electrical stimulation of the nerves distally, for example, at the hand digits using ring electrodes, and recording the response proximally at the nerve trunk, for example, the median nerve at the wrist for orthodromic conduction (from the Greek ortho–straight and dromo–course). The test can also be done by stimulating the nerves proximally, for example, the median nerve at the wrist and recording distally at the digits (antidromic conduction). (Fig. 7.1A and B) Sensory CV can also be calculated by the more physiological method of tactile stimulation (Buchthal, 1982a; Simonetti et al., 1998). The SNAPs produced with tactile stimulations, however, are smaller and more dispersed (Krarup and Trojaborg, 1994) and are not used in routine clinical practice.

156

TULIO E. BERTORINI Fig. 7.1 (A) Antidromic stimulation of the median nerve at the wrist and recording at the index finger. (B) Orthodromic stimulation of the median nerve on the index finger and recording at the wrist.

SNCS are more sensitive than motor conduction tests in the diagnosis of peripheral neuropathy (Gilliatt and Sears, 1958; Buchthal and Rosenfalck, 1971) and nerve entrapments (Buchthal et al., 1974). Sensory nerve conduction studies also help in differentiating plexopathies in which the SNAP are usually either of low amplitude or absent, from radiculopathies in which they are usually normal (Kimura, 2001). Sometimes, however, spinal canal disorders can also affect the posterior root ganglia causing a decreased SNAP amplitude (Kikuchi et al., 1994; Levin, 1998; Hurdle et al., 2003), but because the dermatomal territory of individual roots is more limited than those of the plexus, SNCS are still helpful in their differentiation. For example, a cervical radiculopathy affects the SNAP in single digits, whereas in plexopathy multiple digital SNAPs could be involved (Newman and Nelson, 1983; Ferrante and Wilbourn, 1995). Because the SNAPS are of shorter duration and smaller size than the CMAP, they are more sensitive to the effects of temporal dispersion and phase cancellation that occur with increasing the distance between stimulation and recording electrodes (Fig. 7.2). This

produces a drop in the proximal amplitude, and the determination of conduction block is less reliable in SNCS (Wilbourn, 1994). Another problem with SNCS is that with increased distance between the stimulation and recording sites, there could be a disappearance or a poor demarcation of the initial positive wave when using orthodromic stimulation, or a less clear demarcation of the onset of the negative wave when using the antidromic technique, making measurement of the exact maximum conduction velocity difficult, particularly in proximal limbs. Sensory CV measured at the beginning of the SNAP is about 54 m/s or above, and at its end is about 34 m/s. There is a clear relationship between the size of the myelinated fibers and their conduction velocity with a correction factor of four (Boyd and Davey, 1968; Buchthal et al., 1984). Thus, the large myelinated fibers that conduct at these velocities have diameters of about 9 μm or above. The potentials of these fibers form the main component of the SNAP (Fig. 7.3) because they have a larger membrane area that produces a higher current flux in the generation of the action potential (Shefner, 2002).

SENSORY NERVE CONDUCTION STUDIES

157

Fig. 7.2 Antidromic median nerve SNAP obtained by stimulating at the index finger and recording at the wrist (A1) and the elbow, (A2) 20 μV per division and 2 m/s per division.

Peripheral nerves also contain more abundant small myelinated and unmyelinated fibers (Ochoa and Mair, 1969; Buchthal and Behse, 1978; Behse, 1990), and as most of the small myelinated, or Aδ fibers, conduct at velocities slower than 34 m/s, they do not contribute to

the SNAP. Neither do the much slower-conducting unmyelinating fibers, and for this reason they are not assessed with conventional CNCS. The near-nerve needle recording technique (NNNRT), using orthodromic stimulation and signal averaging, is a useful method to study the action potentials generated by smaller myelinated Aδ fibers that conduct as slow as 15 m/s (Shefner and Logigian, 1994; Shefner, 2002). Measurement of the cutaneous silent period obtained by stimulation of a sensory nerve during voluntary muscle contraction is another method to study this fiber population (Luciano, 2002). Microneurography is a technique applied to test the function of various types of nerve fibers, including the unmyelinated C-fibers (Wallin and Elam, 1997). This, however, is an invasive test that is not used in clinical practice. Measurement of the sympathetic skin response, on the other hand, is used to assess the function of the unmyelinated autonomic postganglionic C-fibers that may be affected in neuropathies that also affect small unmyelinated sensory fibers (Wang and Kaufman, 2002). Quantitative sensory tests are psychophysical methods that determine the perception of the stimuli carried by different fiber populations (Verdugo and

–− 10 μV

Sura + 62

21 m/s

33

0.5 μV

Popl. fossa

Fibers 600

400

200 n = 2600

n = 4000 0

14

12

10

6 8 Diameter, μm

4

2

0

Fig. 7.3 Normal sural nerve. The compound nerve action potential was recorded with near-nerve needle electrodes 50 cm (top) and 15 cm (middle) proximal to the site of stimulation at the level of the lateral malleolus. For each recording, 200 traces were electronically averaged. The fiber size distribution of the nerve is shown (bottom, as well as the probable correspondences among fiber diameter, conduction velocity, and the various components of the nerve action potential. The main potential, recorded 15 cm away, corresponds to nerve fibers with diameters 7 μm and more and conduction velocities between 33 and 62 m/s, and some slower components are apparent in the recordings, especially in the recording at the popliteal fossa. (From Buchthal, F, Rosenfalck, A, Behse F (1975) Sensory potentials of normal and diseased nerve. In: PJ Dyck, PK Thomas, EH Lambert (Eds.) Peripheral Neuropathy. WB Saunders, Philadelphia, pp. 442–462 with permission).

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Dubrovsky, 2002), but these are not direct electrodiagnostic tests and will be discussed in another chapter. Finally, measurements of the refractory periods during paired sensory nerve stimulation is another useful test of axonal function that can be applied in the diagnosis of polyneuropathies and entrapment syndromes (Oh, 2002). 7.2. Stimulating techniques During routine SNCS, the sensory nerve is studied using rectangular electrical stimulus with a duration of about 0.1 ms generated by surface discs, or by ring electrodes applied to the digits. Needle stimulation is sometimes used in antidromic sensory testing to allow the study of nerves that are not easily accessible by surface electrodes. In order to activate all large myelinated axons, the stimulus should be about 12 times threshold or just supramaximal. This method is different from motor conduction tests that require stimulation of about 20% above supramaximal to stimulate all motor axons. An excessive stimulus strength causes a larger area of effective stimulation or “stimulus lead,” making this stimulation site to be, in reality, closer to the recording electrode than the site where the distance is measured, producing falsely shorter latencies and faster conduction velocities (Gilliatt et al., 1965). Antidromic and orthodromic techniques produce similar conduction velocities (Buchthal and Rosenfalck, 1966) and latencies (Tackmann et al., 1981; Goddard et al., 1983; Cohn et al., 1990). Some authors, however, report longer latencies and slower conductions with the antidromic method (Murai and Sanderson, 1975; Tashjian et al., 1987). This is likely to be caused by lower temperatures in the more distal recording sites; another reason is that the latencies of the orthodromic SNAP are measured to the peak of the initial positive wave, while in the antidromic method, this is measured at the beginning of the negative wave. Another difference in the techniques is that the SNAP amplitude obtained with orthodromic stimulation is smaller (Fig. 7.4), unless needle-recording electrodes are used (Buchthal and Rosenfalck, 1966). The larger amplitude antidromic response is caused by the close proximity of the ring electrodes to the digital nerves in the hands and by the lower distal limb temperatures. One disadvantage of the antidromic technique is that the SNAP could be distorted by volume-conducted muscle action potentials generated by stimulating the mixed nerves.

Fig. 7.4 (A) Antidromic SNAP of the median nerve obtained by stimulating at the wrist and recording at the index finger, gain 20 μV per division, 2 ms per division. (B) Orthodromic SNAP of the median nerve obtained by stimulating at the index finger and recording at the wrist, gain 20 μV per division, 2 ms per division.

When performing sensory conduction tests, the examiner should be careful to avoid excessive stimulus strength, which could not only produce falsely shorter latencies, but also distort the SNAP that is more sensitive to this effect than the CMAP because of its smaller size. This distortion might be secondary to capacitance coupling between the stimulating and recording electrodes, strong capacitive currents between the stimulating electrode and ground, and by polarization capacitances between the electrodes and skin (Brown and Watson, 2002). Several methods are used to reduce the stimuli artifacts, including the placement of the ground electrode between the stimulating and recording sites, by maintaining a low impedance of the skin by removing surface skin oil with alcohol or

SENSORY NERVE CONDUCTION STUDIES

acetone and by light abrasion using sandpaper with application of electrode paste. The stimulating electrode should be placed as close as possible to the nerve, allowing low intensity stimulation to produce maximal SNAP. Also, properly increasing the distance between stimulation and recording sites diminishes stimulus artifacts (Brown and Watson, 2002). 7.3. The characteristics of the sensory nerve action potential The SNAPs obtained with antidromic techniques are characteristically biphasic with an initial negative wave (Fig. 7.4A). The orthodromic response is usually triphasic with an initial positive wave (Fig. 7.4B). Recordings could apply bipolar or monopolar montages. In the former, both recording electrodes are placed along the length of the nerves; with this they are both electrically active. In monopolar recordings, the reference electrode is placed about 3 cm remote from the nerve. As will be discussed later, a monopolar montage is also used in the NNNRT. Bipolar montages at 2 cm distance between the recording electrodes produce a higher amplitude SNAP because of summation of the action potentials generated from the active and reference electrodes. With an excessive increase in the interelectrode distance, the action potential could have two peaks because the response from the first electrode terminates before the second response. This is also seen in nerves with slow nerve conduction from demyelination. Some use a 4-cm distance between stimulating electrodes because this produces less SNAP distortion (Edwards and Burke, 1988; Lee et al., 1993; Glitter and Stolov, 1995). With a separation of more than 3 cm, the action potential appears tetraphasic with a final negativity (Buchthal and Rosenfalck, 1966). A 3-cm separation produces less noise and is easier to apply in the digits (Wagman and Lesse, 1952), and for this reason it is favored by others, while some use a distance of 2 cm (Kimura, 2001). The values obtained with either montage should be compared with exactly the same techniques used to obtain normal control values. Orthodromic nerve stimulation with less than maximal strength also produces a potential with two peaks. This phenomenon was initially thought to be caused by variation in nerve conduction velocities along different axons (Buchthal and Rosenfalck, 1966), but recordings at different sites produce similar conduction velocities of the two peaks, and most likely, the double-peaked response is generated by the

159

stimulus being generated both at the cathode (first peak) and the anode (second peak). Using less than maximal stimulation, with the application of local anesthetics known to block sensory receptors causes a disappearance of the second response, and thus it is likely that this peak is generated by the sensory receptors that are stimulated by the anode (Aprile et al., 2003). Conventionally, the amplitude of the SNAP is measured from the onset to the peak of the negative wave, or from the peak of the positive wave to the peak of the negative wave. The duration is measured from the onset of the negative wave to its termination as it crosses the baseline. The area under the curve of the SNAP can be measured automatically with the use of modern equipment, providing a better assessment of the population of the large myelinated fibers contributing to the action potential (Fig. 7.5). 7.4. Determination of latencies and conduction velocities of sensory nerves When using antidromic stimulation, the SNAP latency is usually measured at the onset of the negative wave (Bertorini, 1970) as this represents the conduction of the faster fibers. In orthodromic techniques, the latency to the positive peak is used frequently, although some use the negative peak (Joynt, 1983). Measurement of the latency to the negative peak does not represent the latency of the faster fibers (Martinez et al., 1980), but this method is used for practical

Peak latency

Onset latency

Area

A m p l i t u d e

Duration

Fig. 7.5 Diagram of the SNAP with its various measurements.

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TULIO E. BERTORINI

purposes because the initial positive wave is sometimes difficult to identify, particularly when studying proximal sites (Gilliatt and Sears, 1958). This difficulty is now somewhat overcome with the use of newer higher resolution equipment and signal averaging. Normal sensory nerve conduction velocities (CV) are 5–10% faster than the motor fibers (Dawson, 1956), and as was discussed before, they can be calculated by measuring the latency from the stimuli artifact to the response, or between the latency from two recording sites. Determination of velocities using two recording sites provides a faster CV because the latency of activation has a fixed delay of 0.15 m/s at the stimulus site (Krarup et al., 1992). When distances are taken under consideration, the velocities obtained with the orthodromic and antidromic techniques are similar (Cohn et al., 1990). 7.4.1. Factors that influence sensory conduction studies Interpretation of SNCS should take into consideration several individual variations that influence the results, such as, patient’s age, height and limb temperature, and also several technical factors that could affect the results and produce inaccurate or poorly reproducible values. 7.4.1.1. Age Sensory conduction velocities vary with age and are slower at birth, particularly in premature babies (Cerra and Johnson, 1962). The values in newborns are about half that of adults (Desmedt et al., 1973), and by the age of 5 to 8, they reach adult values (Gamstorp, 1963; Buchthal et al., 1975). This slower conduction is likely the result of incomplete nerve myelination in infants (Thomas and Lambert, 1960), as myelination normally starts around the fourth month of gestation (Ochoa and Mair, 1971) and is not completed until approximately the age of 5 (Gutrecht and Dyck, 1970).

Sensory conduction velocities start to decline after the age of 20 (Norris et al., 1953; Desmedt and Cheron, 1980) (see Table 7.1). This occurs at a rate of about 1 m/s per decade from 20 to 55 years. Beyond 55 years, the decline is at about 3 m/s per decade (Brown and Watson, 2002). The effects of aging in conduction velocity are likely related to a loss of myelinated fibers (Swallow, 1966; O’Sullivan and Swallow, 1968), variations of internodal lengths, and due to segmental demyelination that occur in older persons (Lascelles and Thomas, 1966). 7.4.1.2. Height Individual heights also influence the results of nerve conduction tests (Stetson et al., 1992; Salerno et al., 1998); for example, increased height is associated with lower SNAP amplitudes and prolonged distal latencies in normal people (Soudman et al., 1982; Gadia et al., 1987; Rivner et al., 1990; Stetson, 1992; Robinson et al., 1993; Hyllienmark et al., 1995). The influence of height and conduction velocities has also been documented in peripheral neuropathy (Gadia et al., 1987). A correlation between increased arm length with slower nerve conduction velocity has been reported by some (Stetson et al., 1992; Robinson et al., 1993; Wu et al., 1999), but not by others (Soudman et al., 1982; Hennessey et al., 1994). The effects of height in conduction velocities might be related in part to lower temperature of the distal limb and also to the smaller distal axonal diameter and shorter internodes. Height, thus, should be taken into consideration prior to interpreting the results of the SNCS as abnormal, particularly in very tall individuals. 7.4.1.3. Gender Faster conduction velocities and higher amplitude SNAPs have been reported in women (Robinson et al., 1993). The increased amplitude might be related to

Table 7.1 Maximum Orthodromic Sensory Conduction Velocities Nerve Median nerve

Age (yr)

n

Wrist to Elbow

Elbow to Axilla

18–25 48–64 70–88

66 11 24

64.8 ± 5.2 55.5 ± 2.6 53.5 ± 4.7

68.7 ± 7.2 67.6 ± 10.2 60.9 ± 7.8

Modified from Buchthal F, Rosenfalck A: Evoked action potentials and conduction velocity in human sensory nerves. Brain Res 1966;3:1-122.

SENSORY NERVE CONDUCTION STUDIES

women having smaller finger circumference, making the ring electrodes to be closer to the axonal branches in digits when conduction velocities are measured through antidromic technique in the hand (Bolton and Carter 1980). Conduction velocities also appear to normalize when they are corrected to the person’s height (Stetson et al., 1992; Robinson et al., 1993). 7.4.1.4. Temperature Electrical activity of the peripheral nerve is influenced by changes in temperature (Hodgkin and Katz, 1949), and a failure of conduction due to conduction block has been observed in temperatures as low as 9˚ C (Paintal, 1965), while increasing temperature to above 45˚ C produces similar effects (Davis and Jacobson, 1971). The effects of temperature in nerve function are related to ionic channel function. For example, voltage-gated sodium channels activate and deactivate very slowly at lower temperatures, producing action potentials of larger amplitude and longer duration until there is conduction block (Frankenhauser and Moore, 1963). At high temperatures, channel activation and deactivation occur faster, so that the action potential is reduced in amplitude (Shefner, 2002). Conduction block occurs when the sodium channel deactivation is so rapid that there is insufficient depolarization to excite the next node (Rutkove et al., 1997). The effects of temperature have been documented in several human studies (Bolton et al., 1981; Lang and Puusa, 1981; Bolton et al., 1982; Denys, 1991; Lee et al., 1993; Phillips, 1996). For example, lower temperatures have been shown to increase the SNAP amplitude (Bolton et al., 1981) while raising the skin temperature to above physiological levels causes a reduction amplitude (Chaudhry et al., 1993; Rutkove et al., 1997). In one study, the amplitude of the median nerve SNAP dropped from 80 μV to 40 μV with an increase in temperatures from 20 to 30˚C (Bolton and Carter, 1981), while in another, increasing temperatures from 32 to 42˚ C also caused a drop in SNAP amplitude by an average of about 50% (Rutkove et al., 1997). At lower temperatures, there is a prolongation of the distal latency and slower conduction velocities (Bolton and Carter, 1981; Geerlings and Mechelse, 1985; Miyamoto et al., 1990), and Buchthal and Rosenfalck reported a linear increase of CV of about 2 m/s per degree of temperature between 22 and 32˚C

161

(Buchthal and Rosenfalck, 1966). In demyelinated nerves, increasing temperature raises the CV until there is a conduction block (Franssen et al., 1999), and similar effects have been observed in compressive neuropathy (Wang et al., 1999). Temperature has less effect in the conduction of small myelinated fibers; as was demonstrated with the use of near needle recordings (DeJesus et al., 1973). 7.4.1.5. Ischemia Although ischemia is not a physiological factor, its effects on SNCS should be discussed as this may play a role in nerve entrapments. Ischemia affects nerve excitability and causes progressive slowing of conduction velocity accompanied by a decreased amplitude and duration of the SNAP (Seneviratne and Peiris, 1968). These effects, as observed with the use of pneumatic tourniquets, occur more rapidly in carpal tunnel syndrome (Fullerton, 1963). Elderly individuals and those with diabetes and uremia have a greater resistance to ischemia than normals (Caruso et al., 1973), and this appears to be related to chronic hypoxemia (Hampton et al., 1989) and related to the levels of glycemia (Parry and Kohzu, 1990). 7.4.1.6. Technical factors Several technical factors can affect the results of SNCS. These include, among others, improper measurement of the latency time, particularly when using older analog techniques, this is now a lesser problem with the use of modern digital equipment. Also, latencies should be recorded at exactly the same point of the SNAP in both proximal and distal recordings, which is facilitated by the proper use of signal averaging. The interelectrode distance should be measured accurately, particularly because the short segments used in calculating sensory CV could cause errors from improper tape placement and skin movements (Oh, 2002). To minimize this, the distance should be measured from the center of the stimulating electrode to the center of the recording electrode with flexible tapes. Ideally, the tested nerve segment should be over 10 cm. This is obviated when using inching techniques. As discussed before, the stimulus strength should be just at supramaximal level. Excessive strength produces falsely shorter latencies, because the effective stimulating site occurs closer to the recording electrode than the site where it is measured. Filters should be properly selected to avoid distortions of the SNAPs and to reduce background noise

162

TULIO E. BERTORINI

that might affect proper recording of the exact latencies. The low frequency filter is usually set at 20 Hz to minimize baseline fluctuations, and the high frequency filter should be set at approximately 2000 Hz to reduce high frequency noise (Oh, 2002). The same sensitivity and sweep speeds should be used to measure the latencies from proximal and distal stimulating points. A higher sensitivity is ideal to determine the exact SNAP measure point. Temperature affects latencies, CV and amplitudes. For this reason, the laboratory temperature should be maintained between 21 and 23˚ C, while the patient’s skin surface temperature should be at 32˚C or slightly above, and if it is lower than this, the limb should be warmed with the use of infrared lamps or warm water for about 30 min. If the temperature cannot be raised using these techniques, the results can be normalized by adding 5% of the measured CV for each degree of temperature lower than 32˚ C. This and other similar conversion factors, however, can produce misleading interpretation in sick nerves (Chodoroff et al., 1985; Baysal et al., 1993; Notermans et al., 1994; Ashworth et al., 1998).

7.5. Special techniques used in the evaluation of sensory nerve function 7.5.1. Inching Inching techniques are applied in studies of motor nerves for the diagnosis of entrapments by stimulating with short distance increments of 1 to 2 cm, to determine the exact site of compression (Miller, 1979; Campbell et al., 1988; Kanakamedala et al., 1988). This technique can also be applied in the study of sensory and mixed nerves (Fig. 7.6). For example, Kimura used inching with antidromic and orthodromic stimulation and recorded at different points of the median nerve to diagnose carpal tunnel syndrome (Kimura, 2001). Others reported that this technique is more sensitive than routine tests in the diagnosis of that condition (Seror, 2000). Similarly, the exact location of ulnar nerve entrapment can be detected by stimulating the mixed ulnar nerve while recording the potential at different short distances below and above the elbow using NNNRT (Odabasi et al., 1999).

Inching study A Before surgery

−6 −4 −2 0 2

−6 −5 −4 −3 −2 −1 0 1 0

0.5

1.0

1.5 ms

0

0.5

1.0

1.5 ms

B After surgery −6 −5 −4 −3 −2 −1 0 1 50 μV 1 ms

Fig. 7.6 Sensory nerve potential in a patient with the carpal tunnel syndrome before and after surgery. Preoperative study showed a localized slowing from −4 to −3 with a calculated segmental conduction velocity of 8 m/s, which normalized in a repeat study conducted six months postoperatively. Reprinted from Kimura J (2001) Electrodiagnosis in Diseases of Nerve and Muscle: Principles and Practice, Oxford University Press, Oxford, IIIrd ed. with permission.

SENSORY NERVE CONDUCTION STUDIES

7.5.2. Mixed nerve conduction Similar to SNCS, mixed nerve stimulation produces compound nerve action potentials that can be measured directly. These potentials are larger than the SNAPs because they incorporate motor axons, and particularly axons from muscle afferents, which are larger and conduct electricity faster than sensory axons, and are more sensitive to the effects of compression and demyelination (Raynor et al., 1994; Odabasi et al., 1999). To test mixed nerve conduction, the nerve is stimulated orthodromically, and the biphasic response is recorded at different sites over the nerve (Chang et al., 2002). Recordings are made either with surface electrodes or using NNNRT (Odabasi et al., 1999). (Fig. 7.7).

47.0 32.4

A

163

Palmar stimulation of the median nerve and plantar stimulation of the tibial nerve are mixed conduction tests used in the diagnosis of carpal tunnel syndrome (Chang and Lien, 1991) and tarsal tunnel syndrome (Saeed and Gatens, 1982; Ponsfor, 1988). 7.5.3. Near nerve needle recording technique The NNNRT is a relatively simple and minimally invasive method of sensory nerve testing in the assessment of polyneuropathy (Oh et al., 1992, 2001; Claussen et al., 1996) (Fig. 7.8) and entrapment syndromes (Odabasi et al., 1999). The method consists in orthodromic sensory nerve stimulation with an intensity of 0.5 to 1 mA for a stimulus of 0.2 m/s, and 1–5 mA for stimulus of a 0.05 msec duration. Recordings are made using a Teflon-coated steel needle electrode with an uncoated tip area of 2 mm and a reference electrode with an uncoated tip of 3.5 mm. The recording electrode is placed near the nerve, and the reference electrode at 3–4 cm of transverse distance. The high frequency filter is set at

3μV 2 m/s

B

A

23.8

c II–III

11.3

II−III 27.5

42.6

24.7 25.7 IV−V

c 12.8

32.1

1μV 2 ms

III−IV

B

28.5

1 μV 59.0

26.5

2 m/s

Fig. 7.7 Mixed nerve conduction in the forearm segment in the ulnar nerve in a case of chronic inflammatory demyelinating polyneuropathy. (A). Two peaks of mixed compound nerve action potential indicative of demyelination. Numbers at the top of two peaks represent nerve conduction velocities (m per second). Clearly, the nerve conduction velocity with the second negative peak is slow. (B). Near-nerve needle mixed compound nerve action potential showing dispersion phenomenon. Numbers at the positive and negative peaks represent nerve conduction velocities (m per second). From Oh, SJ (2002), Nerve conduction velocity tests: their clinical applications, In Bertorini TE, Clinical Evaluation and Diagnostic Tests for Neuromuscular Disorders. Butterworth Heinemann, New York, Chapter 5.

V 33.3

24.3

c V

6.7 28.7

Fig. 7.8 Compound nerve action potential responses with the near-nerve needle sensory nerve conduction of the digital and interdigital nerves of the plantar nerves. (A). Axonal neuropathy. (B). Demyelinating neuropathy. Both cases had a distal sensory neuropathy with normal routine nerve conduction study. Roman numerals indicate the digit number. Arabic numbers under and above each compound nerve action potential denote maximum and negative peak nerve conduction velocity. Number between two bars indicates compound nerve action potential duration. (Reprinted with permission from SJ Oh, et al. (2001) Large-fiber neuropathy in distal sensory neuropathy with normal routine nerve conduction. Neurology, 56: 1570–1572.)

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5–10 KHz to allow recording of fast components. This induces noise is hard to eliminate, but can be improved by averaging 500–1000 responses. Near nerve needle recording technique allows the study of slow myelinated fibers with velocities as low as 15 m/s (Behse and Buchthal, 1971), and is useful in the diagnosis of peripheral neuropathies in which the SNAPs are not recordable with surface electrodes (Oh, 2002). This method also helps to diagnose involvement of large myelinated fibers in patients with apparent “small fiber neuropathy” that have normal routine SNCS. In these, the NNNRT could show slowing and dispersion of the slow components, indicating demyelination (Oh, 2001) (Figs. 7.8 and 7.9). The NNNRT has been valuable to diagnose patients suspected to have demyelinating neuropathy not supported by standard motor conduction tests (Claussen et al., 1996; Oh et al., 2001), and to diagnose tarsal tunnel syndrome, plantar neuropathy and interdigital neuropathies of the foot (Oh et al., 1985, 2001) and, as discussed, to localize ulnar nerve entrapments (Odabasi, 1999) (Fig. 7.10). 7.5.4. Measurement of refractory periods of sensory nerves After the generation of an action potential, the axon becomes inexcitable for a period of time called the refractory period (RP). This has two components, the absolute refractory period (ARP), which is the time

Fig. 7.9 Dispersion phenomenon of the sensory compound nerve action potential indicative of demyelination in near-nerve needle sensory nerve conduction of medial plantar nerve in a case of Guillain–Barré syndrome. Nerve conduction in the other nerves shows motor neuropathy without any feature of demyelination. Notice the normal maximum and negative peak nerve conduction velocities (numbers at the positive and negative peaks) and the long duration (16 msec) of the compound nerve action potential. From Oh, SJ (2002), Nerve conduction velocity tests: their clinical applications. In TE Bertorini (Ed.), Clinical Evaluation and Diagnostic Tests for Neuromuscular Disorders. Butterworth Heinemann, New York, Chapter 5.

TULIO E. BERTORINI

wherein the nerve is completely inexcitable to a second impulse, and the relative refractory period (RRP), which is the period of time when a response is obtained only with stimulus of increased intensity. Measurement of the RP of sensory, motor and mixed nerves is useful in the evaluation of peripheral nerve function and the diagnosis of neuropathies and entrapments syndromes (Oh, 1993, 2002). The ARP is caused by inactivation of the sodium current moving inward during the depolarization (Shefner, 2002), and lasts for about 1 m/s after the action potential (Gilliatt and Willison, 1963; Stohr, 1981). The RRP is caused by an increased permeability of the voltage-gated potassium channels that make the axon relatively inexcitable for an additional 3 m/s. After the RRP there is a time during which the axon becomes hyperexcitable; this is called the supernormal period and lasts from 4 to 20 m/s from the initial action potential. During this time, the threshold of activation is reduced by 10–20% lower than the initial stimuli (Bostock and Grafe, 1985; Potts et al., 1994; Kiernan et al., 1996; Shefner et al., 1996). The supernormal period is produced by depolarization after potentials that occur after the initial action potential (Bowe et al., 1987). Depolarization is followed by a longer period of relatively decreased excitability, called the subnormal period, which lasts up to 100 m/s, and is caused by slow potassium conductance and membrane hyperpolarization (Shefner, 2002).

32

16

1 μV 3m

37

SENSORY NERVE CONDUCTION STUDIES

165

Near-nerve needle sensory nerve conduction T R C

VF-BE(6)

1 $ BE(6)-BE(1.5) 2

BE(1.5)-AE 4 ms delay

0.62 μV 3

5 m/s Near-nerve needle mixed nerve conduction

4 W-BE(6) 10 μV 5

BE(6)-BE(1.5) 5 μV BE(1.5)-AE

6 5 m/s

2.5 μV

Fig. 7.10 Near-nerve sensory and mixed nerve conductions in a case with tardy ulnar sensory neuropathy. The most prominent change is noted with the sensory nerve conduction in BE (1.5)-AE segment: conduction block and slow nerve conduction velocity. Routine nerve conduction showed an absent mixed nerve compound nerve action potential at the elbow-axilla segment and normal motor nerve conduction study across the elbow. Inching technique was not able to localize the lesion. (AE-above elbow; BE-below elbow; VF-fifth finger; W = wrist). From Oh, SJ (2002) Nerve conduction velocity tests: their clinical applications. In: Bertorini, TE, Clinical Evaluation and Diagnostic Tests for Neuromuscular Disorders. Butterworth Heinemann, New York, Chapter 5.

In routine clinical electromyography, testing the ARP and RRP of sensory nerves is relatively simple with paired stimulation of various time intervals using surface stimulating and recording electrodes (Fig. 7.11). The ARP is measured by using interstimulus intervals starting at 2 m/s that are shortened consecutively by 0.1 m/s until the second stimulus produces no response. The normal ARP is about 0.51 m/s in the mixed ulnar nerve and 0.76 m/s in the sural nerve (Oh, 1993). To determine the RRP, the stimulus interval starts at 5 m/s and is shortened consecutively by 0.25 m/s. The RRP is determined as the interval by which the

7

8

TEMP: 33.0⬚C

Fig. 7.11 Refractory period for the sensory median nerve. Paired stimuli of the median nerve, antidromic stimulation with recording at the third digit. Notice the first appearance of the second response (absolute refractory period) at the bottom (long arrow). The antidromic response becomes equal in amplitude to the first response at the top (short arrow). The time between the two stimuli that produced equal amplitude response is the relative refractory period. Calibration: 20 μV/cm; 2 ms/cm. The stimulation was done at 0.5 millisecond (bottom line, 8), increasing up to 5-m/s (top line, 1) interstimuli intervals. From Oh, SJ (2002) Nerve conduction velocity tests: their clinical applications. In: Bertorini, TE (Ed.), Clinical Evaluation and Diagnostic Tests for Neuromuscular Disorders. Butterworth Heinemann, New York, Chapter 5.

amplitude of the second response decreases or has a prolonged latency. This period ranges from 2.50 m/s in the mixed ulnar nerve to 3.45 m/s in the ulnar sensory nerve (Oh, 1993, 2002).

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Measurements of the RP, particularly ARP appear to be more sensitive than conventional nerve conduction tests in the diagnosis of peripheral neuropathy (Lowitzsch and Hopf, 1973; Schutt et al., 1983; Braune, 1999). Tackmann and Lehmann, for example, demonstrated a prolonged latency and decreased amplitude of the second response in carpal tunnel syndrome. In another study (Holden et al., 1987), the ARP was prolonged, and the RRP had a 50% reduction in amplitude and was prolonged in this condition. An absent or prolonged second response was documented by Gilliatt and Meer in 79% of hands with carpal tunnel syndrome (Gilliatt and Meer, 1990), and these abnormalities were detected even when sensory conduction was normal. 7.5.4.1. Measurement of the cutaneous silent period The silent period is the time in which stimulation of a peripheral nerve causes a suppression of the electromyographic activity of voluntary contracting muscles. The MIXED NERVE SILENT PERIOD is obtained by stimulating a mixed nerve, whereas the CUTANEOUS SILENT PERIOD (CSP) is obtained by stimulating sensory nerves. The mixed nerve silent period lasts from 100 to 200 m/s and has three distinct periods. The first is an initial suppression of electromyographic activity between the CMAP and the onset of the F-wave. This lasts about 30 ms in the hand and is caused from a collision of the antidromic stimulus volley with the volitional orthodromic motor impulse. The second silent period follows the F-wave, lasts about 30–75 m/s and is likely the result of recurrent (Renshaw cell) inhibition from the antidromic motor volley (Shahani and Young, 1973). The third period of suppression of muscle activity follows a long latency reflex and appears at about 60–100 m/s. This results from the activation of high threshold cutaneous afferents, which are similar to those activated by the cutaneous silent period and is mediated by polysynaptic or oligosynaptic reflexes (Stetkarova et al., 2001). The mixed nerve silent period is used in the assessment of motor nerve hyperexcitability, such as in tetanus, but not in the evaluation of sensory nerves, and thus will not be discussed further. The CSP is elicited by a stimulation of either a digital or other sensory nerve, which produces a suppression of voluntary activity in muscles innervated by those nerves or different nerves or roots. The stimuli required for the suppression of muscle activ-

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ity should be in the nociceptive range, as the CSP is likely to be mediated by small myelinated Aδ fibers (Uncini et al., 1991; Shefner and Logigian, 1993; Leis, 1998). The CSP lasts about 75–140 m/s on the hand. (Luciano, 2002). To obtain the CSP, single shocks with intensity of about 10–12 times above the perception threshold (producing pain) are delivered to cutaneous nerves. In order to determine the onset and end of the CSP, the rectified EMG signal is averaged to include recordings of background EMG activity for about 50–150 m/s before the stimulation in order to determine the mean amplitude. The onset of the silent period is the first time in which the EMG activity decreases in amplitude to a certain percentage, for example 80% of the background, and the end is the time when the amplitude returns to the baseline level. The CSP has been used in the diagnoses of radiculopathy, root avulsion (Leis, 1998), entrapment syndromes (Aurora et al., 1998) and particularly to detect small fiber dysfunction in neuropathies (Leis, 1994) (Fig. 7.12). 7.5.4.2. Measurement of the sympathetic skin response The sympathetic skin response (SSR) assesses the function of preganglionic and postganglionic sympathetic activity and for this reason is used to test postganglionic unmyelinated fibers, which are similar to the small unmyelinated sensory axons. The SSR is a transient change in the electrical potential of the skin due to a change in conductance from sweat glands that are innervated by postganglionic cholinergic nerve cells. This response can be produced by stimulation of peripheral nerve afferents and deep breathing, as well as magnetic stimulation of the central nervous system (Wang and Kauffman, 2002). Conventionally, the SSR is obtained by stimulating the median nerve at the wrist or the tibial nerve at the ankle with recording of the palm of the hand or sole of the foot respectively, although stimulation in other sites and even the opposite limb provides similar results (Shahani et al., 1984). To obtain the SSR, the stimulus intensity should be at about 10–30 mA, and the recording sweep speed should be 0.5–1 second per division. Using sensitivity of about 100 μV to 1 mV per division the low frequency filter should be set at 0.1–2 Hz and the high-frequency filter at 4000 Hz. The response is a biphasic or triphasic potential (Fig. 7.13) with variable amplitude, and because of this variability, some measure an averaged response

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1. Control

2. Fabry’s

3. Fabry’s

4. Fabry’s Stimulus

50 m/s

Fig. 7.12 (A) Mixed nerve silent period. Stimulation of the median nerve at the wrist results in three periods of suppression of electromyographic activity. The first period (S1) ends approximately 30 m/s after the stimulus with the appearance of the F-waves. The second period (S2) follows the F-waves and lasts from 30 to 75 m/s until the appearance of the long latency reflex. The third period (S3) follows the long latency reflex and is thought to be equivalent to the cutaneous silent period. (B) Examples of waveforms showing the cutaneous silent period (CSP) over a period of 50 m/s in the lower extremity in control subject (1) and three patients with Fabry’s disease (2–4). Each trace represents the rectified average of 10 CSPs. Markers indicate the beginning and end of the CSPs, also indicated by asterisks. The CSP was absent in patient 4 (4). From Ali Syed, N, Sandbrink, F, Luciano, CA et al., 2000, Cutaneous silent periods in patients with Fabry’s disease. Muscle Nerve, 23: 1179–1186, with permission from John Wiley & Sons, Inc.

(Schondorf, 1997) while others measure the larger response, in clinical testing (Baba et al., 1988). The normal palmar SSR has been reported to have a latency of 1.51 ± 0.165 sec and an amplitude of about 479 ± 105 μV, whereas the plantar response has a latency of 2.07 ± 0.165 second and amplitude of 101 ± μV (Knezevic and Bajada, 1985). Similar values have been reported by others (Elie and Guiheneuc, 1990; Oh, 2003). Shahani and co-workers calculated the CV of the SSR by measuring the proximal and distal segments of a limb and obtained a velocity of 1.57 m/s in the arm and 1.02 to 1.49 m/s in the leg (Shahani et al., 1990). The SSR increases in amplitude and latency at higher temperatures (Deltombe et al., 1998). Some clinicians use the absence of the response or a drop in amplitude to define abnormalities in neuropathy (Toyokura, 1998), while others diagnose neuropathy based on prolonged latency of the SSR (Denisilic and Meh, 1997).

Fig. 7.13 Sympathetic skin response studying median nerve at the wrist recording at the palm reference electrode instead of the hand, 500 m/s per division, gain 500 μV.

Abnormalities of SSR have been documented in a variety of neuropathies (Elie and Guiheneuc, 1990; Wang et al., 1994; Brown et al., 1996; Ferrante et al., 1997) particularly in diabetes (Soliven et al., 1987; Levy et al., 1992; Zgur et al., 1993) and amyloidosis (Shivji and Ashby, 1999). The response has been found to be normal in diseases that affect large myelinated fibers, such as Friedreich’s Ataxia (Ingall and McLeod, 1991), but abnormal in hereditary motor sensory neuropathy, which also affects primarily large myelinated fibers (Denisilic and Trontelj, 1993). Abnormal SSR have also been reported in disorders that do not affect the peripheral nerves but likely impair central sympathetic activity (Haapaneimi et al., 2000). Variable results and even normal SSR have been seen in some patients with peripheral neuropathy (Evans et al., 1988; Montagna et al., 1988). As a result, its habituation and absence is noticeable in some normal people, particularly the elderly (Elie and Guiheneuc, 1990; Shahani et al., 1990; Hoeldtke et al., 1991). Therefore, the results of the SSR measurement should be interpreted with caution and taken in context of the patient’s clinical presentation and the findings of other electrodiagnostic tests. 7.5.5. Microneurography Microneurography is a technique used to assess the function of postganglionic sympathetic fibers (Birkett et al., 1992; Fagius and Berne, 1994; Wallin et al., 1994; Mano et al., 1996; Sugiyama et al., 1996), as well as cutaneous afferents and muscle afferents (Ochoa, 1996), providing information of the function of the various types of nerve fibers (Navarro et al., 1989; Burke, 1992).

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Recordings are made using tungsten microelectrodes with 200 μm diameter with an insulated tip of several μm inserted into a major nerve trunk or cutaneous nerve branch, using a subcutaneous reference electrode. The same electrode can also be used to elicit an evoked sympathetic response such as sweat, or changes in blood flow. The perception of different sensation types in the stimulated receptive field (Macefield et al., 1990) is used to assess the involvement of different types of nerve fibers. Also, conduction velocity of large axons can be measured by stimulating isolated single axons at different sites, while calculations of the conduction of Aδ and C-fibers can be made by stimulating the nociceptive and thermo receptive fields (Kakuda, 1992; Simone et al., 1994; Schmelz et al., 1998; Wallin and Elam, 1999). Microneurography is a valuable research tool used to study somatosensory and autonomic function, but it is an invasive procedure that is not applied in routine clinical electrodiagnosis. 7.5.6. In vitro nerve conduction tests In vitro nerve conduction tests (IVNCT) are performed on surgically excised nerve segments of about 4–5 cm, placed in a humidified, temperature-controlled chamber attached to a small platinum or silver wires that are used for stimulation with recording at the opposite end. Variations of the stimulus intensity, as well as sweep speeds, allow the study of action potentials of different nerve fiber types (Lambert and Dyck, 1975). Nerve action potentials generated during the IVNCT include firstly a large Aα potential conducting about 60 m/s, corresponding to large myelinated axons. This is followed by a small Aδ potential, corresponding to a small myelinated fiber conducting at about 15–20 m/s. A much later potential produced by a higher stimulus intensity is recorded using slower sweep speed. These potentials correspond to unmyelinated axons (C-fibers), which conduct at about 1 m/s (Fig. 7.14) (Hursh, 1939; Gasser, 1960; Lambert and Dyck, 1975). Dyck, Lambert and collaborators performed detailed analysis of IVNCT and identified characteristics of the potentials in various types of neuropathy (Dyck et al., 1971; Dyck and Lambert, 1974; Lambert and Dyck, 1975). They found an absent Aα potential that corresponded to absence of large myelinated fibers in Friedreich’s Ataxia, and a decreased Aα, and an absent

Fig. 7.14 In vitro nerve conduction of a normal sural nerve recorded with a TECA-TE42 electromyograph machine. (A). A large response represents the Aα potential; a smaller response, the Aδ potential. Sweep velocity = 1 ms/cm; sensitivity = −1 mV/cm. (B). C potential. Sweep velocity = 5 ms/cm; sensitivity −50 μV/cm. From Oh, SJ (2002), Nerve conduction velocity tests: their clinical applications. In: TE Bertorini (Ed.), Clinical Evaluation and Diagnostic Tests for Neuromuscular Disorders. Butterworth Heinemann, New York.

C potential with a reduced Aδ potential in patients with small fibers neuropathies. All potentials were reduced in amplitude in hereditary sensory neuropathy type I, although this was more pronounced in the Aδ and C potentials, corresponding to markedly decreased number of unmyelinated and small myelinated fibers in histology (Dyck et al., 1971) INVCT are not used in routine clinical practice. 7.6. Application of sensory nerve studies in diagnosis of peripheral nerve disorder As discussed before, SNCS has been found to be more sensitive than motor studies in peripheral neuropathy (Gilliatt and Sears, 1958; Buchthal and Rosenfalck, 1971), particularly in the lower extremities (Burke

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et al., 1974). This is expected in axonopathies that are length dependent and affect mainly the longer axons. The demonstration of low amplitude or absent sural SNAPs in patients with normal radial nerve SNAPs in these types of axonal neuropathy reinforces this concept (Rutkove et al., 1997). The diagnosis of distal axonopathies is made easier with stimulation of the toes with NNNCT, allowing the detection of abnormalities even in patients with normal routine sensory conduction tests (Park et al., 2003). NNNCT also provides an opportunity to study minimal conduction from smaller myelinated fibers. Slowing of minimal conduction could indicate the presence of small regenerating axons in axonopathy (Shefner et al., 1991). Similar findings, however, can also be found in demyelinating neuropathy (Krarup and Trojaborg, 1996). Measurements of the CSP allow the study of small myelinated fibers and help in the diagnosis of neuropathies that affect these fibers (Leis, 1994; Syed et al., 2000). The unmyelinated C-fibers, on the other hand, can be tested by the measurement of the SSR (Levy et al., 1992; Denisilic and Trontelj, 1993; Claus and Schondorf, 1999; Shivji and Ashby, 1999; Nazhel et al., 2002). Microneurography is also used to study unmyelinated fibers (Simone et al., 1994; Schmelz et al., 1998). Measurements of the refractory periods help in the assessment of axonal dysfunction prior to their detection using conventional diagnostic tests (Tackmann et al., 1974; Schutt et al., 1983; Braune, 1999). The value of SNCS in the diagnosis of demyelinating neuropathy is somewhat limited and less helpful than motor conduction tests (Simmons et al., 1993), as although significant slowing can be diagnostic of demyelination, the assessment of conduction block is difficult because of the normal drop in amplitude that occurs in even healthy persons with proximal stimulation (Horowitz and Krarup, 1992; Wilbourn, 1994). Also, a decreased SNAP amplitude or its absence occur in both demyelinating and axonal neuropathies. As discussed previously, NNNRT helps in the diagnosis of large fiber involvement in patients with apparent small fiber neuropathy by the detection of the dispersion phenomenon with very slow components that indicate demyelination (Oh, 2003). SNCS are more sensitive than motor studies in the diagnosis of entrapment neuropathies, such as carpal tunnel syndrome (Buchthal et al., 1974) and tarsal tunnel syndrome (Oh, 2002), and measurements of the

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conduction velocity of the mixed ulnar nerve can be used to diagnose ulnar compression at the elbow (Raynor et al., 1994) and peroneal nerve entrapments (Singh et al., 1974). Inching techniques are also helpful in the diagonosis of carpal tunnel syndrome and ulnar neuropathy (Kimura, 1979; 1984), while the NNNRT is useful in diagnosing tarsal tunnel syndrome and plantar entrapments (Oh et al., 1985, 1999). Finally, measurements of the cutaneous silent period (Aurora et al., 1998) and the refractory periods appear to be more sensitive than routine SNCS in the diagnosis of carpal tunnel syndrome (Gilliatt and Meer, 1990; Pallyath and Holden, 1990). Acknowledgements I would like to thank Dr Pushpa Narayanaswami and Dr William Owens for reviewing the manuscript and their valuable comments, Ms Mary Lucas for her editorial help, Sue Maccarino for technical assistance, and Julie Carter and Cindy Culver for secretarial help. References Aprile, I, Stalberg, E, Tonali, P and Padua, L (2003) Double peak sensory responses at submaximal stimulation. Clin. Neurophysiol., 114: 256–262. Ashworth, NL, Marshall, SC and Satkunam, LE (1998) The effect of temperature on nerve conduction parameters in carpal tunnel syndrome. Muscle Nerve, 21: 1089–1091. Aurora, S, Ahmad, B and Aurora, T (1998) Silent period abnormalities in carpal tunnel syndrome. Muscle Nerve, 21: 1213–1215. Baba, M, Watahiki, Y, Matsunaga, M and Takebe, K (1988) Sympathetic skin response in healthy man. Electromyogr. Clin. Neurophysiol., 28: 277–283. Baysal, AI, Chang, CW and Oh, SJ (1993) Temperature effects on nerve conduction studies in patients with carpal tunnel syndrome. Acta Neurol. Scand., 88: 231–216. Behse, F and Buchthal, F (1971) Normal sensory conduction in the nerves of the leg in man. J. Neurol. Neurosurg. Psychiatry, 34: 404–414. Behse, F and Buchthal, F (1978) Sensory action potentials and biopsy of the sural nerve in neuropathy. Brain, 101: 473–493. Behse, F (1990) Morphometric studies on the human sural nerve. Acta Neurol. Scand. Suppl., 132: 1–38.

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CHAPTER 8

Testing for focal processes: inching in short increments* Mark E. Landaua, * and William W. Campbellb a Walter Reed Army Medical Center, Washington DC and Uniformed Services University of the Health Sciences, MD, USA

b

8.1. Introduction 8.1.1. The role of experimental error Nerve conduction velocity (NCV) determinations vary because of both biological and technical factors. The experimental error inherent in the technique occurs primarily as a result of variability in measuring the latency and the distance. Understanding experimental error is critical when describing inching techniques due to the relatively short distances that are used. Other critical variables in NCV determination include temperature, age, height, and the effects of volume conduction. Maynard and Stolov (1972) examined the experimental error in NCV determinations using sophisticated mathematics, and demonstrated the important relationship between distance and experimental error. The details of this influential paper are relevant to any discussion of segment length in NCV determinations. Their subject electromyographers used a metal compass divider to measure latency on a polaroid photograph of the sweep from a TECA model B-2 electromyograph. Latency measurements accounted for almost 90% of the experimental error. Of the relatively small contribution of distance measurement error to total experimental error, most was due to skin movement errors rather than tape measure reading errors. Skin movement is more of a problem over long distances than over short distances. The authors’ primary aim was to construct families of error curves relating the calculated experimental error in msec as a function of conduction velocity (CV) and distance. These curves convincingly demonstrate that the percentage of calculated experimental error

* Correspondence to: Mark E. Landau, MD, Asst. Chief, Clinical Neurophysiology, 6900 Georgia Ave., Walter Reed Army Medical Center, Washington DC 20307, USA. E-mail address: [email protected] Tel.: (202)782-8654; fax: (202)782-2295.

falls as the distance over which the NCV is determined increases. As an example, the authors stated that the experimental error is unacceptably high, greater than 25%, when distances of less than 100 mm are used for all velocities of 40 msec or more. But the main conclusion was that there was a desperate need for improved methodology for latency determinations. Following Maynard and Stolov, most investigators and clinicians adopted a minimum 10 cm distance for studying the ulnar nerve at the elbow and the peroneal nerve at the fibular head. The 10 cm rule was born. 8.1.2. False negative errors Modern electrodiagnostic medicine practice commonly requires the detection of focal nerve pathology, and even in some instances precise localization of the pathology. Unfortunately, as increasing conduction distance lowers experimental error and avoids overdiagnosis, it also decreases the likelihood of detecting a focal lesion. The normally conducting segments mask any abnormality. When nerve conduction studies are performed over longer distances, normal segments of the nerve are included in the latency and velocity determinations. With increasing length, the effect of the abnormal segment on the final calculation decreases. The normally conducting segments dilute the conduction delay of the abnormal segment, and the sensitivity for detecting a focal lesion is lessened. Consider a normal nerve conduction at 50 msec. The conduction time for a one-centimeter segment is 0.2 ms. Presume a lesion that triples the conduction time to 0.6 ms. If nerve conduction studies are performed using a distance of 8 cm, including the

*

The opinions or assertions contained herein are the private views of the authors and are not to be construed as official or as reflecting the views of the department of the army or the department of defense.

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lesioned segment, then the NCV will be 40 msec (seven 1-cm segments with conduction time of 0.2 msec and one 1-cm segment with conduction time of 0.6 ms). If a distance of 14 cm is used, the NCV will be 44 msec. With longer distances, the CV approaches the normal CV for the nerve, despite the lesioned segment. Experimental error and sensitivity for lesion detection are reciprocally related. So, pari passu with low experimental error and increased specificity comes underdiagnosis and decreased sensitivity. Many conventional and commonly used techniques employ distances shorter than 10 cm. For example, most investigators who have examined the issue have concluded that 6–8 cm across palm sensory studies are more sensitive than 13–14 cm studies between wrist and digit in the diagnosis of carpal tunnel syndrome (Carroll, 1987; DeLean, 1988; Cioni et al., 1989; Jackson and Clifford, 1989; Kuntzer, 1994; Padua et al., 1996, 1997; DiGuglielmo et al., 1997; Scelsa et al., 1998). 8.1.3. History Very short nerve segments have been studied reliably and successfully, both percutaneously and intraoperatively. As early as 1963, Kaeser analyzed segments as short as 20 mm. Kimura advocated short segment studies to localize conduction abnormalities in the carpal tunnel syndrome, commenting that “a large perunit increase in latency more than compensates for the inherent measurement error associated with multiple stimulation in short increments” (Kimura, 1979, 1984). Brown et al. (1976) showed that studying median, ulnar and peroneal segments as short as 5 mm intraoperatively was possible and practical, and that abnormalities were often restricted to segments of 5–10 mm. Brown and Yates (1982) also demonstrated that percutaneous short segment studies were feasible. Kanakamedala et al. (1988) and Kanakamedala and Hong (1989) used 20 mm segments to study controls and patients with ulnar neuropathy at the elbow (UNE) and peroneal neuropathy at the knee and found marked abnormalities across discrete segments. Campbell et al. (1988, 1992) found excellent correlation between percutaneous short segment studies and abnormalities demonstrated by direct epineural stimulation intraoperatively in patients with UNE. With digitally acquired waveforms, storage methods and automated latency measurements, latency error now accounts for considerably less of the total experimental error in NCV determinations than in 1972 (Landau et al., 2002, 2003a, 2003b). A repeat of

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Maynard and Stolov’s original work (1972) with the current generation of electrodiagnostic equipment showed that much shorter distances could be used (Landau et al., 2002). The error at a distance of 5–6 cm is comparable to that found at 10 cm in the previous study. Despite the improved equipment, conduction studies at distances of one or two centimeters still have relatively high experimental error as compared to studies done over more conventional distances. 8.1.4. Types of short segment studies The American Association of Electrodiagnostic Medicine (2001) defines inching as a nerve conduction study technique wherein stimulus applications at multiple short distance increments along the course of a nerve are performed to localize an area of focal slowing or conduction block (CB). The term “inching” was introduced by Miller (1979) to describe the technique of moving the stimulator along the nerve in several sequential steps in search of points of abrupt change in compound muscle action potential (CMAP) amplitude or configuration indicative of CB or differential slowing due to underlying focal demyelination. This type of inching technique did not include measurement of latency changes over consecutive segments, or precise measurement of segment lengths. The point where the CMAP changed identified the location of the abnormality. The other type of inching, sometimes called “centimetering”, includes measuring both latency and amplitude changes over precisely measured segments of 5–20 mm (Brown and Yates, 1982; Kanakamedala et al., 1988; Kanakamedala and Hong, 1989; Campbell et al., 1992). This latter, more quantitative type of inching study, is sometimes referred to as a short segment study (SSS) or short segment incremental study (SSIS). We prefer the term SSIS. Percutaneous short segment techniques have been described for the median nerve at the wrist, ulnar nerve at the elbow and wrist, and peroneal at the fibular head (Kimura, 1979; Brown and Yates, 1982; Kanakamedala et al., 1988, Kanakamedala and Hong , 1989; Campbell et al., 1992; Imaoka et al., 1992; McIntosh et al., 1998; Seror, 1998; Hermann et al., 2001; Seror, 2001; Azrieli et al., 2003). 8.1.5. Pathophysiology: conduction block and focal slowing Pathological and electrophysiological data suggest that focal mononeuropathies occur over a 5–20 mm

TESTING FOR FOCAL PROCESSES: INCHING IN SHORT INCREMENTS

segment of the nerve (Brown et al., 1976; Kimura, 1979; Brown and Yates, 1982). When CB or differential slowing is present, the localization of a focal neuropathy by ordinary inching is simple and straightforward. The precise site of the pathologic process is betrayed by a loss of amplitude or change in waveform configuration, or both, between two stimulation sites (Fig. 8.1). Complete CB involving all fibers is rare. More often, loss of amplitude or change in configuration occurs because CB or conduction slowing involves only some of the larger, fast conducting fibers. Slowing involving only some fibers, but without CB, causes temporal dispersion of the CMAP, producing loss of amplitude with prolongation of the CMAP duration, or differential slowing. When slowing involves all fibers uniformly, there is no temporal dispersion and no change in CMAP shape across the lesion, but the conduction time is increased. CB results from the failure of impulse transmission in severely demyelinated fibers. The healing process of remyelination, pathologically accompanied by shorter, more slowly conducting internodes may relieve CB but leave persistent focal slowing. Inching technique that evaluates only amplitude and configuration changes is useful only when CB or significant differential slowing is present. A major degree of CB occurs in only a minority of chronically compressed or

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entrapped nerves. CB, partial or complete, was present in only about a third of ulnar nerves studied intraoperatively (Brown et al., 1976; Campbell et al., 1988). In a chronic compressive neuropathy, the most likely focal abnormality is restricted slowing across the lesion, unaccompanied by significant loss of amplitude or major changes in waveform configuration (Fig. 8.2). Remyelination with short internodes may be adequate to eliminate major CB, but leave a “footprint” of focal slowing to mark the site of injury. When the predominant pathophysiology is uniform focal slowing, conventional inching without precise measurement of segment length will fail to detect the lesion. Unfortunately, methods of localization that depend only on changes in waveform amplitude or configuration will fail in the majority of cases. When compared head to head, SSIS was found to be superior to conventional inching in the evaluation of UNE (Campbell et al., 1992). 8.1.6. Upper limits of normal As with more routine electrodiagnostic tests, specificity and sensitivity for short segment studies relate to the chosen upper limit of normal (ULN) conduction time (see Table 1.1). This determination is even more crucial in short segment studies because of the higher

Fig. 8.1 Conduction block in a patient with multifocal motor neuropathy: decreased CMAP area and amplitude when median nerve stimulated over elbow compared to wrist.

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5 ms

A 1 A 2 A 3

53.3 ma 2 mv

40.0 ma 2 mv

36.9 ma 2 mv

A

28.2 ma

4

2 mv

A

66.7 ma 2 mv

5 A

94.1 ma

6

2 mv

A

22.8 ma 2 mv

7 A 8

19.6 ma 2 mv

Fig. 8.2 SSIS of ulnar nerve in elbow region. Latency between A6 and A7 = 5.0 ms. All other remaining segments < 0.4 ms. No conduction block. The abnormal segment corresponds to the retroepicondylar region.

experimental error. Opting for 3 standard deviations (SD) from the mean ensures relatively high specificity while sacrificing sensitivity (more false negative studies). With lower SDs, the sensitivity is increased and specificity is decreased (more false positive studies). Choice of the ULN depends on the ultimate purpose of the study. If invasive management decisions depend on secure diagnosis and precise localization of the abnormality, then it would be better to choose higher specificity. In relatively mild cases that may be managed conservatively, and with otherwise normal routine electrodiagnostic tests, lower ULNs may be more useful. The standard deviations for ulnar motor CV measurements at various distances were derived using a mathematical model (Landau et al., 2002). For a nerve with a CV of 50 msec (conduction time of 0.2 ms

over 1 cm), considering experimental errors in distance and latency measurements, the conduction time over a 1-cm segment would be 0.2 ± 0.08 ms. Using a two SD cutpoint, the ULN is 0.36 ms; if 3 SD, then 0.44 ms. There are five published reports that derived reference values for 1-cm segments clinically (Kimura, 1979; Nathan et al., 1988; Campbell et al., 1992; McIntosh et al., 1998; Seror, 1998). Kimura (1979) developed a short segment study of median sensory fibers at the carpal tunnel. Normal patients had 1-cm segment conduction times between 0.16 and 0.22 ± 0.1 ms. If two standard deviations were used as a cutoff of the normal, then 0.42 ms would be the upper limit of normal, corresponding closely to the value derived from the mathematical model. In Kimura’s study, focal slowing was arbitrarily defined as conduction time exceeding 0.5 ms over 1-cm, i.e. approximately three standard deviations, ensuring high specificity. Using the Kimura technique, Nathan et al. (1988) derived the nonparametric range of normal between 0.1 and 0.3 ms. The sensitivity for CTS improved from 54 to 81% when using an upper limit of normal of 0.4 ms instead of 0.5 ms. As expected, the specificity declined from 97 to 81% with an upper limit of 0.4 ms. Campbell et al. (1992) described a short segment technique for ulnar motor fibers at the elbow. Reference values were determined from segments proximal to lesions studied intraoperatively. The mean latency change over 52 such 1-cm segments was 0.23 ± 0.1 ms. This method suggests a ULN of 0.43 ms for a 1 cm segment (mean + 2 SD). This value may be spuriously long due to the operative setting and inability to regulate temperature. Nonetheless, it is nearly identical to Kimura’s data derived percutaneously. 8.1.7. Clinical utility One question regarding short segment studies is when to use them in electrodiagnostic practice (Campbell, 1998; Geiringer, 1998). In general, a screening tool should have high sensitivity, and be practical, cost effective and time efficient; specificity is less important. If the highly sensitive tool suggests an abnormality, a test with higher specificity can be used. Short segment studies are not as practical as screening studies and are relegated to a secondary role, following more time-efficient tests. Unfortunately, routine electrodiagnostic tests may be less sensitive because of the longer distances and dilution effect. If clinical suspicion is high and routine studies are normal, SSIS may disclose an abnormality. Making the diagnosis of a

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Table 8.1 Short segment techniques with derived reference valuesa ULN (ms) Study

Nerve

Distance (cm)

Normal (ms)

SD (ms)

2 SD

Kimura (1979)

Peroneal (M)

0.16–0.22 0.19–0.25 0.184 0.39 0.45 0.29 0.23 0.43 0.36 * 0.2 0.4 0.42

0.4

Kanakamedala (1989)

1.0 1.0 1.0 1.5 2.0 1.0 1.0 2.0 2.0 1.0 1.0 2.0 2.0

0.08–0.10

Seror (2001) Imaoka (1992) Seror (1998) Nathan (1988) Campbell (1992) Kanakamedala (1988) Azrieli (2003) McIntosh (1998) Landau (2002)

Median (M) Median (S) Median (S) Median (S) Median (S) Median (S) Ulnar (M) Ulnar (M) Ulnar (M) Ulnar (Wrist) Ulnar (M)

* 0.057 0.04 0.08 0.1 0.1 0.08 * 0.08 0.11 0.1

* 0.51 0.49 0.45 0.43 0.63 0.5. 0.43 0.36 0.62 0.62

3 SD

0.36 0.57 0.53 0.53 0.53 0.73 0.58 0.44 0.73 0.72

a

The normal values are the mean or range of values published. M, Motor; S, Sensory; cm, centimeters; ms: milliseconds; SD, Standard deviation; *, unstated.

subtle demyelinating nerve lesion may not change management, but mere diagnosis may avoid obtaining other unnecessary diagnostic tests. Short segment studies can precisely localize a focal abnormality. Management decisions rarely exploit this knowledge. Ulnar neuropathy at the elbow (UNE) can occur at different sites (see below), most are in the retroepicondylar groove or at the humeroulnar arcade (HUA). Short segment studies can distinguish between the two (Campbell et al., 1992). It has been suggested that HUA lesions be managed by surgical release of the aponeurosis, and lesions of the retroepicondylar segment by ulnar transposition (Campbell et al., 1992). Further data is necessary to support this contention. If the surgical technique is chosen regardless of the pathoanatomy, which is usually the case, then one can argue against the need for precise localization (Geiringer, 1998). Non-surgical management is the same, regardless of localization. 8.2. Specific nerves 8.2.1. Median nerve Short segment techniques for evaluating carpal tunnel syndrome (CTS), have used increments of 1.0, 1.5, and 2.0 cm (Kimura, 1979; Imaoka et al., 1992; Seror, 1998, 2001).

8.2.1.1. Motor and antidromic sensory using 1-cm increments In 1979, Kimura described a percutaneous short segment technique for localizing the exact site of median nerve abnormality in CTS (12). The technique evaluated both motor and sensory fibers (Fig. 8.3). The median nerve was stimulated percutaneously at 12 sites in 1-cm increments, between the midpalm and distal forearm. The zero level represented the proximal edge of the transverse carpal ligament (TCL) and was at the distal wrist crease. To determine the origin of the thenar motor branch, stimulating electrodes, with the anode distal, were moved proximally in small increments from the distal palm until a thumb twitch appeared. The most typical location for the thenar branch origin was 3–4 cm distal to the zero level, approximately the distal edge of the TCL. Electrodes were placed over the abductor pollicus brevis and its tendon to record the CMAP and around the second digit to record an antidromic sensory nerve action potential (SNAP). Two trials were given at each stimulus site to confirm consistency. Care was taken to use just maximal shock intensity at each stimulus site to avoid spread of the stimulating current. A bandwidth of 20 Hz to 32 kHz for CMAP recordings, and 20 Hz to 2 kHz for SNAP recordings, was used. For the sensory studies, the onset latency was used when possible. A specially designed amplifier with

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In Kimura’s patients with CTS, most of the sensory conduction delays occurred 2–4 cm distal to the distal crease of the wrist. Most of the motor delays were 1–2 cm more proximally. Sensory and motor axons were also not delayed to the same degree. The discrepancy may be due to the fascicular organization of the median nerve. Of the 172 symptomatic hands, 52% had a sharply localized slowing. The remainder possessed abnormalities evenly distributed across the carpal tunnel. The same patients had sensory and motor studies comparing palm-digit or palm-APB latencies to wrist-palm latencies. These studies had a 92% sensitivity for detecting CTS. The aim of the study was not to compare palmar segment techniques to 1-cm incremental techniques and, therefore, cannot make conclusions as to which was more sensitive. The short segment study disclosed abnormalities in four patients who would have otherwise had a false negative palmar study.

Fig. 8.3 Motor and sensory 1-cm inching technique for the median nerve. Recording electrodes over the APB and the second digit for evaluation of the motor and sensory studies, respectively. From Kimura (1979) with permission from Oxford University Press.

short blocking time and low noise was used in order to determine onset latencies accurately. If the onset was not clear, then peak latencies were used. There was no significant latency difference using peak or onset latency. Reference values were obtained from patients with complaints unrelated to CTS. The 1-cm segments had mean latencies ranging from 0.16 ± 0.08 to 0.22 ± 0.10 ms. Focal slowing was arbitrarily defined as conduction time exceeding 0.5 ms/cm, or more than twice that of the other 1-cm segments. For the motor studies, the latencies ranged from 0.19 to 0.25 ms/cm. A drawback of the motor studies was difficulty testing serially over the wrist-to-palm segment. The latency changes per cm in this region were inconsistent even in the control population, presumably due to inadvertent stimulation of the recurrent thenar branch near the motor point rather than the median nerve proper.

8.2.1.2. Orthodromic sensory 1-cm technique Ring electrodes are used to stimulate, with the cathode placed around the proximal interphalangeal joint of the middle finger and the anode distal (Seror, 1998). A pediatric ground electrode is placed over the proximal finger, between the ring electrode and the metacarpophalangeal joint. SNAPs are recorded with a bar electrode at 1 cm intervals from a point 4 cm proximal to the wrist crease up to a point 6 cm distal to it. It is sometimes necessary to average when recording from the segment of nerve beneath the transverse carpal ligament. Peak latencies are compared over successive segments. Seror determined that the mean and maximum conduction delay between 1-cm segments in normal patients was 0.184 and 0.36 ms, respectively. All patients with CTS had at least one segment with a conduction delay of 0.34 ms or greater. 8.2.1.3. Antidromic 1.5-cm technique A modified inching method for CTS diagnosis and prognosis was published in 1992 (Imaoka et al., 1992). Nine recording electrodes, separated by 1.5 cm, were placed along the median nerve, beginning 3 cm proximal to the wrist crease and extending in a straight line towards the third digit. The median nerve was stimulated at the antecubital fossa to avoid stimulus artifact, while an 8-channel electromyograph recorded from the neighboring paired electrodes in a bipolar montage (Fig. 8.4). To avoid motor contamination, the stimuli were not necessarily supramaximal. Twenty to fifty stimuli were averaged.

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distal wrist crease

183 Fig. 8.4 Schema of recording electrodes over the median nerve. 8-channel device with recording electrodes separated by 1.5 cm. Stimulation over the median nerve in the antecubital fossa. From Imaoka et al. (1992) with permission from John Wiley and Sons, Inc.

8ch 7ch 6ch 5ch 4ch 3ch 2ch 1ch

12 11 10 9 8 7 6 5 4 3 2 1 0

The peak latencies were plotted against distance. A line connecting the proximal four latencies was extended to derive theoretical latencies for the distal four latencies. The actual latencies for these points were then compared to the theoretical ones. The mean latency for the proximal 4 segments was 0.39 ms/(1.5 cm). Abnormality was defined as a segment exceeding 3 SD from the norms, which approximated 0.61 ms. Of the 68 symptomatic hands, testing was abnormal in 66 (97% sensitivity). Recall that Kimura used a 1-cm segment latency > 0.5 ms as the criterion for abnormality, with a sensitivity of 62%. 8.2.1.4. Orthodromic 2-cm technique A prospective study comparing mild CTS with controls using an orthodromic median sensory nerve technique was published in 2001 (Seror, 2001). The median nerve was stimulated over the third digit by ring electrodes with the cathode over the proximal interphalangeal joint. Five recording electrodes were placed 2 cm apart over the median nerve at the wrist to record sensory nerve action potentials. The conduction time delay was the time between the negative peaks of two consecutive SNAPs. The control population had conduction times of 0.45 ± 0.04 ms over 2 cm. The authors chose 2.5 SD as the upper limit of normal and derived a sensitivity and specificity for CTS of 94 and 98%, respectively. Note the relatively short conduction time for the control population compared to Kimura’s study at 1-cm. Using a relatively shorter upper limit of normal, the

sensitivity of the study increased dramatically. The technique was directly compared to the orthodromic median-ulnar latency difference to the fourth digit (64% sensitivity). Short segment techniques had a vital role in defining the pathophysiology of CTS. Kimura (1979) established that abnormal conduction of the median nerve occurred 2 to 4 cm distal to the distal wrist crease, and that conduction abnormalities were restricted to 1 to 2 cm of the nerve segment. Rarely, however, are these techniques necessary in the routine evaluation of patients with suspected CTS. Relatively simpler techniques, with good sensitivity and specificity are sufficient (Robinson et al., 1998). We consider using short segment studies for patients with high clinical suspicion of CTS, negative combined sensory index, and no other etiological explanation for the symptoms. 8.2.2. Ulnar nerve at the elbow Ulnar neuropathy at the elbow (UNE) is the secondmost common mononeuropathy. At least three distinct compression neuropathies may affect the ulnar nerve in the general region of the elbow. Lesions occur in the retroepicondylar groove, at the humeroulnar arcade (HUA) or “cubital tunnel,” and at the aponeurosis investing the deep surface of the flexor carpi ulnaris, 4–7 cm distal to the medial epicondyle (deep flexorpronator aponeurosis). Lesions greater than 3 cm proximal to the medial epicondyle (ME) have also

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been reported (Hermann et al., 2001). Miller first used the inching technique in 1979 to confirm that the UNE due to HUA compression, the “cubital tunnel syndrome”, is distinct from the more proximal lesions in the ulnar groove (at the time often referred to as tardy ulnar palsy). All patients had partial CB when stimulated above the elbow. In 9 of 15 ulnar neuropathies, inching disclosed CB 1.5–3.5 cm distal to the medial epicondyle. The findings were confirmed intraoperatively in seven patients. In contrast, Brown and Yates (1982) encountered no instances of cubital tunnel compression in 30 cases of UNE studied by short segment stimulation intraoperatively. Conduction abnormalities were all centered around the ME. There is no clear explanation for the differences in their observations. A recent large surgical series using intraoperative nerve action potential recordings again found the majority of ulnar nerve lesions occur in the ulnar groove (Kim et al., 2003). In 1988, Campbell et al., performed intraoperative electroneurography during 19 ulnar nerve explorations. Normative data was derived from segments proximal to the lesion sites. In no proximal segment was the conduction time greater than 0.4 ms over 1 cm. The mean ± SD of the uninvolved segments was 0.23 ± 0.1 ms. Only two patients had CB. Of the 19, using direct epineural SSIS, conduction abnormality was noted in the ulnar groove in nine, HUA in four, both in three, and deep within the flexor carpi ulnaris in one. The surgical procedure done depended on the electrodiagnostic information, with anterior (subcutaneous) transpositions performed for patients with retroepicondylar abnormalities and simple release for

Fig. 8.5 Short segment incremental study of the ulnar nerve. Stimulations are separated by 1-cm. The “zero” point represents the point at which a line connecting the medial epicondyle to the olecranon crosses the ulnar nerve. Recording over the ADM.

MARK E. LANDAU AND W.W. CAMPBELL

those with HUA compression. This unique approach to surgical intervention for UNE has not been replicated. This is seemingly the most rational approach to the surgical management of UNE. As there are different pathophysiologies for UNE, so should there be different management decisions. Unfortunately, there is not yet any outcomes data to support this view. This approach would encourage more widespread use of SSIS. In 1992, percutaneous SSIS was directly compared to inching and intraoperative short segment studies in patients with UNE (Campbell et al., 1992). Of the 16 abnormal ulnar nerves studied by ordinary, nonquantitative inching, clear localizing changes occurred in only 3 (19%). Of the 26 nerves studied by SSIS, localizing abnormalities occurred in 21 (81%). These methods were compared to the intraoperative results. Percutaneous SSIS accurately predicted the localization in 17 of 21. The authors concluded that SSIS identifies compression levels more accurately than standard inching, and that percutaneous SSIS and intraoperative electroneurography correlate highly but not perfectly. 8.2.2.1. Short segment incremental study: one-centimeter technique The elbow is flexed at 70–90˚ with the shoulder abducted and externally rotated. The course of the ulnar nerve is mapped with subthreshold stimuli (Fig. 8.5). The stimulator is moved perpendicular to the course of the nerve until the maximum CMAP amplitude for a given subthreshold stimulus is obtained, and the point is marked. This process is

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185

repeated at intervals proximal and distal to the ME. The points are then connected to outline the precise course of the nerve. A perpendicular line is drawn from the ME to the olecranon. This is the zero point. One-centimeter intervals are measured along the course of the nerve. The proximal points are labeled −1, −2, etc., and the distal ones +1, +2, etc. Stimulation with intensities just above maximal is applied at each point. Excessive stimulation must be avoided, as distal migration of the effective cathode can produce erroneous latency determinations. Although not done in the original study, it may be wise to use calipers with a defined distance of 1-cm for measurements to diminish measurement errors. Latencies between two points of stimulation greater than 0.45 ms are considered suspicious. At least two trials are done, with the starting position offset at least by one position in order to maintain objectivity. Only if there is anatomic concurrence on both trials is a definite diagnosis made. The location of the lesion is reported anatomically in relation to the distance in cm from the ME. 8.2.2.2. Two-centimeter techniques Two short segment studies of the ulnar nerve using 2-cm increments have also been described (Kanakamedala et al., 1988; Azrieli et al., 2003). In one study, the control group had a mean conduction time of 0.43 ± 0.10 ms, and the other 0.36 ± 0.08 ms (Fig. 8.6). From a practical standpoint, the electrodiagnostic medicine consultant must decide what increments to use for short segment studies and what ULN to use in determination of abnormality. The 2-cm increments are less time consuming. For the ulnar nerve, studies have indicated a 2 SD ULN of 0.63 or 0.52 ms for 2-cm segments and 0.43 ms for 1-cm segments. The variance is very likely due to differences in control population, technique, equipment, and temperature. In our opinion, the potential risk of overdiagnosis resulting in an unnecessary surgical intervention is greater than missing the diagnosis of mild UNE. Therefore, we would tend to use relatively longer latencies for diagnostic purposes: 0.5 ms for 1 cm intervals and 0.7 ms for 2 cm intervals. There are no studies comparing the distances to determine which is more sensitive, although each of the studies reported approximately 80% sensitivity of diagnosis. 8.2.2.3. Technique following anterior transposition Short segment stimulation studies are useful for evaluating the ulnar nerve after surgical treatment for UNE

Fig. 8.6 Short segment study of ulnar nerve, with stimulations separated by 2-cm. Stimulation points (D6, D4, D2, P, P2, P4, and P6) denote distal or proximal to medial epicondyle and the number of centimeters. Splint is used to maintain elbow flexion at 90 degrees. Reprinted from Kanakamedala et al. (1988) with permission from American Congress of Rehabilitation Medicine and the American Academy of Physical Medicine and Rehabilitation.

with a poor outcome. Paternostro-Sluga et al. (2001) studied 19 patients with 21 anterior transpositions. The exact position of the nerve was localized by submaximal stimulations. Six stimulation points separated by 2.5 cm were measured along the course of the nerve. Although supramaximal stimulations were performed over each point, care was taken to avoid excessive stimulation and subsequent distal migration of the effective cathode. The upper limit of normal was 0.98 ms for the 2.5 cm segment. Of the 21 ulnar nerves, 16 were abnormal. The authors compared the electrophysiological findings with a clinical scale, and found that focal conduction slowing did not correlate with postoperative clinical findings.

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8.2.2.4. Summary Routine electrodiagnostic techniques currently used in the evaluation of UNE have limitations of both sensitivity and specificity, and are associated with high experimental error due to distance measurement errors (Landau et al., 2002). We use SSIS in the following situations.

an abnormality. Only two patients were studied and each had evidence of CB. SSIS precisely localized the lesion to 1 cm proximal to the distal wrist crease in one patient. In the other patient, there was a more diffuse lesion affecting the ulnar nerve over several centimeters from the takeoff of the superficial branch to the deep palmar branch.

(1) Patients with signs and symptoms suspicious for UNE and an isolated abnormality of motor conduction across the elbow, without confirmatory abnormalities of the sensory study or needle examination. SSIS is used to confirm or disprove the finding. (2) Patients with signs and symptoms suspicious for UNE in whom routine motor conduction studies fail to demonstrate an abnormality. (3) Patients with evidence of ulnar neuropathy in whom motor conduction studies do not clearly localize the lesion to the elbow. Many patients with “pure axon loss” UNE will have focal slowing demonstrable by SSIS. A completely normal SSIS across the elbow raises the possibility of a lesion in another location, such as the forearm. (4) Patients with UNE who are potential surgical candidates. SSIS is used to localize the conduction delay and assist in appropriate surgical management. In cases of UNE due to lesions at the HUA, we favor avoiding transposition of the nerve, and limiting the intervention to simple release of the aponeurosis. (5) Patients with persistent symptoms following ulnar nerve transposition. A modified short segment study is performed. As with SSIS, submaximal stimulations are used to define the new nerve course. Then stimulations separated by 1-cm are used to determine if any persistent, or new, conduction delays exist.

8.2.4. Peroneal neuropathy at the knee

8.2.3. Ulnar nerve at the wrist A short segment incremental study has also been used to evaluate ulnar neuropathy at the wrist (McIntosh et al., 1998). In ten normal subjects, while recording from the first dorsal interosseous, stimulations separated by 1-cm from 3 cm proximal to the distal wrist crease to 4 cm distal to the distal wrist crease were applied to the ulnar nerve. The maximal change in latency over each of the seven 1-cm segments was 0.4 ms/cm, and the mean +2 SD was 0.43 ms. The authors chose a latency of 0.5 ms between segments to denote

Kanakamedala and Hong (1989) described a short segment technique for peroneal neuropathy at the knee, using 2-cm segments distal and proximal to the fibular head (FH). In normal subjects, the mean conduction times ranged from 0.38 ± 0.08 to 0.43 ± 0.17 ms, and the mean CMAP amplitude change ranged from 0.3 ± 1.7 to 0.9 ± 6.1 mv. In 18 patients, most abnormalities were just proximal to the FH, but abnormalities were detected anywhere between 6 cm proximal and 4 cm distal to the FH. Using this method, 15 patients had a statistically significant amplitude decrement, compared to only seven patients when using conventional methods (50% decrement in CMAP amplitude directly comparing the amplitudes resulting from stimulations 6 cm proximal to FH and 4 cm distal to FH). Amplitude decrements as little as 6.9% over a 2-cm segment were considered abnormal. Three had prolonged conduction time without any amplitude change. 8.2.5. Other Short segment studies can be performed on many other nerve segments, but standardized methodologies do not exist. In nerve segments that are relatively deep, subcutaneous needle stimulations may be given at sequential intervals to ensure adequate stimulus intensity. This is particularly useful to distinguish the physiological from abnormal temporal dispersion when a decreased CMAP amplitude is detected across a long segment of nerve. A 30% decrease in amplitude between two sites of stimulation separated by 20–25 cm may not be significant. That same decrease noted between 1 or 2 cm could be considered highly significant. For example, a smaller amplitude ulnar CMAP is noted with below elbow stimulation compared to wrist stimulation. Possible explanations for this occurrence include a Martin-Gruber anastamosis (MGA), physiological temporal dispersion, or true conduction block in the forearm segment of the ulnar nerve, assuming good electrodiagnostic techniques. Sequential subcutaneous stimulations of the ulnar nerve, beginning

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distally and progressing proximally, can determine whether an amplitude decrement occurred over a very short segment (1–2 cm) as would occur with CB or a MGA. Decreased CMAP amplitude that occurs with normal temporal dispersion would not occur over such a short segment. This could be particularly useful in cases of multifocal motor neuropathy, peripheral nerve tumors, and focal hypertrophic mononeuropathies, among others. Other methods do exist for detecting MGAs, but an MGA would show an amplitude decrement 8–10 cm proximal to the wrist crease where the crossover fibers usually join the ulnar nerve. 8.3. Conclusion Short segment techniques have been used percutaneously, subcutaneously as well as intraoperatively to refine the pathophysiology and localization of focal mononeuropathies. These techniques can be used clinically to improve the sensitivity for detecting abnormalities, but are generally more time consuming than routine electrodiagnostic studies. When conduction block is absent, clinicians must judiciously choose the upper limit of normal for latencies between different sites, respecting the problems that may occur with a false positive or false negative result. In UNE, where there are multiple possible sites of compression, short segment techniques have the most promise for routine use. Controlled studies comparing the outcome measures of different surgical interventions determined from the electrodiagnostically determined pathoanatomy are needed. References AAEM (2001) Glossary of terms in electrodiagnostic medicine. Muscle Nerve, Suppl. 10: S1–S49. Azrieli, Y, Weimer, L, Lovelace, R and Gooch, C (2003) The utility of segmental nerve conduction studies in ulnar mononeuropathy at the elbow. Muscle Nerve, 27: 46–50. Brown, WF, Ferguson, GG, Jones, MW and Yates, SK (1976) The localization of conduction abnormalities in human entrapment neuropathies. Can. J. Neurol. Sci., 3: 111–122. Brown, WF and Yates, SK (1982) Percutaneous localization of conduction abnormalities in human entrapment neuropathies. Can. J. Neurol. Sci., 9: 391–400. Campbell, WW (1998) The value of inching techniques in the diagnosis of focal nerve lesions.

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Inching is a useful technique. Muscle Nerve, 21: 1554–1556; discussion 1561. Campbell, WW, Sahni, SK, Pridgeon, RM, Riaz, G and Leshner, RT (1988) Intraoperative electroneurography: management of ulnar neuropathy at the elbow. Muscle Nerve, 11: 75–81. Campbell, WW, Pridgeon, R and Sahni, KS (1992) Short segment incremental studies in the evaluation of ulnar neuropathy at the elbow. Muscle Nerve, 15: 1050–1054. Carroll, G (1987) Comparison of median and radial nerve sensory latencies in the electrophysiological diagnosis of carpal tunnel syndrome. Electroencephalogr. Clin. Neurophysiol., 68: 101–106. Cioni, R, Passero, S, Paradiso, C, Giannini, F, Battistini, N and Rushworth, G (1989) Diagnostic specificity of sensory and motor nerve conduction variables in early detection of carpal tunnel syndrome. J. Neurol., 235: 208–213. DeLean, J (1988) Transcarpal median sensory conduction: detection of latent abnormalities in mild carpal tunnel syndrome. Can. J. Neurol. Sci., 15: 388–393. DiGuglielmo, G, Torrieri, F, Repaci, M and Uncini, A (1997) Conduction block and segmental velocities in carpal tunnel syndrome. Electroencephalogr. Clin. Neurophysiol., 105: 321–327. Geiringer, SR (1998) The value of inching techniques in the diagnosis of focal nerve lesions. Inching techniques are of limited use. Muscle Nerve, 21: 1557–1559; discussion 1559–1560. Hermann, DN, Preston, DC, McIntosh, KA and Logigian, EL (2001) Localization of ulnar neuropathy with conduction block across the elbow. Muscle Nerve, 24: 698–700. Imaoka, H, Yorifuji, S, Takahashi, M, Nakamura, Y, Kitaguchi, M and Tarui, S (1992) Improved inching method for the diagnosis and prognosis of carpal tunnel syndrome. Muscle Nerve, 15: 318–324. Jackson, D and Clifford, JC (1989) Electrodiagnosis of mild carpal tunnel syndrome. Arch. Phys. Med. Rehabil., 70: 199–204. Kaeser, HE (1963) Conduction Disorders in Ulnar Nerve Paralysis. Dtsch Z Nervenheilkd, 185: 231–243. Kanakamedala, RV and Hong, CZ (1989) Peroneal nerve entrapment at the knee localized by short segment stimulation. Am. J. Phys. Med. Rehabil., 68: 116–122. Kanakamedala, RV, Simons, DG, Porter, RW and Zucker, RS (1988) Ulnar nerve entrapment at the

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elbow localized by short segment stimulation. Arch. Phys. Med. Rehabil., 69: 959–963. Kim, DH, Han, K, Tiel, RL, Murovic, JA and Kline, DG (2003) Surgical outcomes of 654 ulnar nerve lesions. J. Neurosurg., 98: 993–1004. Kimura, J (1979) The carpal tunnel syndrome: localization of conduction abnormalities within the distal segment of the median nerve. Brain, 102: 619–635. Kimura, J (1984) Principles and pitfalls of nerve conduction studies. Ann. Neurol., 16: 415–429. Kuntzer, T (1994) Carpal tunnel syndrome in 100 patients: sensitivity, specificity of multineurophysiological procedures and estimation of axonal loss of motor, sensory and sympathetic median nerve fibers. J. Neurol. Sci., 127: 221–229. Landau, ME, Barner, KC and Campbell, WW (2003a) Optimal screening distance for ulnar neuropathy at the elbow. Muscle Nerve, 27: 570–574. Landau, ME, Diaz, MI, Barner, KC and Campbell, WW (2002) Changes in nerve conduction velocity across the elbow due to experimental error. Muscle Nerve, 26: 838–840. Landau, ME, Diaz, MI, Barner, KC and Campbell, WW (2003b) Optimal distance for segmental nerve conduction studies revisited. Muscle Nerve, 27: 367–369. Maynard, FM and Stolov, WC (1972) Experimental error in determination of nerve conduction velocity. Arch. Phys. Med. Rehabil., 53: 362–372. McIntosh, KA, Preston, DC and Logigian, EL (1998) Short-segment incremental studies to localize ulnar nerve entrapment at the wrist. Neurology, 50: 303–306.

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Miller, RG (1979) The cubital tunnel syndrome: diagnosis and precise localization. Ann. Neurol., 6: 56–59. Nathan, PA, Meadows, KD and Doyle (1988) Sensory segmental latency values of the median nerve for a population of normal individuals. Arch. Phys. Med. Rehabil., 69: 499–501. Padua, L, LoMonaco, M, Valente, EM and Tonali, PA (1996) A useful electrophysiological parameter for diagnosis of carpal tunnel syndrome. Muscle Nerve, 19: 48–53. Padua, L, LoMonaco, M, Gregori, B, Valente, EM, Padua, R and Tonali, PA (1997) Neurophysiological classification and sensitivity in 500 carpal tunnel syndrome hands. Acta Neurol. Scand., 96: 211–217. Paternostro-Sluga, T, Ciovika, R, Turkof, E, ZundrDungl, A, Posch, M, and Fialka-Moser, V (2001) Short segment stimulation of the anterior transposed ulnar nerve at the elbow. Arch. Phys. Med. Rehabil., 82: 1171–1175. Robinson, LR, Micklesen, PJ and Wang, L (1998) Strategies for analyzing nerve conduction data: superiority of a summary index over single tests. Muscle Nerve, 21: 1166–1171. Scelsa, SN, Herskovitz, S, Bieri, P and Berger, AR (1998) Median mixed and sensory nerve conduction studies in carpal tunnel syndrome. Electroencephalogr. Clin. Neurophysiol., 109: 268–273. Seror, P (1998) Orthodromic inching test in mild carpal tunnel syndrome. Muscle Nerve, 21: 1206–1208. Seror, P (2001) Sensory segmental latency values of the median nerve for a population of normal nerves. Muscle Nerve, 24; 1595–1600.

Peripheral Nerve Diseases Handbook of Clinical Neurophysiology, Vol. 7 J. Kimura (Ed.) © 2006 Elsevier B.V. All rights reserved

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CHAPTER 9

Late responses (F- and A-waves): methodology and techniques Roberto J. Guiloff * Neuromuscular Unit, West London Neurosciences Centre,Charing Cross Hospital, UK

9.1. Introduction Late muscle responses occur over a variable period of time after the direct or orthodromic M-response when a mixed peripheral nerve is stimulated. They include Fwaves (Magladery and McDougal, 1950) and the Hreflex (Hoffman, 1922) at comparable delays after the M-response. Waveforms with latencies intermediate between the M-response and the F-wave and Hresponse include motor axon reflexes (Fullerton and Gilliatt, 1965), indirect double discharges (Roth, 1974, 1985), peripheral late waves (Tomasulo, 1982), myoaxonal ephaptic responses (Roth, 1993) and motor axon loop responses (Roth and Egloff-Baer, 1984). The term “A-wave” has been used to denote just motor axon reflexes, just indirect double discharges or peripheral late waves, all of them and, more recently, also constant and stable waveforms within the F-wave or with more prolonged latencies than the F-waves. F-waves are considered late recurrent muscle responses generated by groups of motor neurons stimulated antidromically, originally with supra-maximal stimuli, though they can also be obtained with submaximal stimuli. H-waves are obtained by submaximal, near threshold, stimulation of mixed nerves with excitation of Ia afferents and monosynaptic motor neuron reflex responses, though they can also occur with supra-maximal stimuli. The different late responses included in the term “A-wave” may have different origins and mechanisms in the peripheral motor axons and their clinical applications are still being defined. This chapter will consider F- and A-waves.

* Correspondence to: Roberto J. Guiloff MD, FRCP, Director, Neuromuscular Unit, West London Neurosciences Centre, Charing Cross Hospital, London W6 8RF, UK. E-mail address: [email protected] Tel.: +44-208-746-8319; fax: +44-208-746-8420.

9.2. F-waves 9.2.1. Introduction F-waves are considered to be late waveforms of muscle action potentials, produced largely by recurrent responses of motor neurons to antidromic supramaximal stimulation. They occur after the M-wave or direct (orthodromic) muscle response to the stimulation of a peripheral nerve. F-waves are indirect responses, their latency is decreased by more proximal stimulation, since this shortens the afferent (antidromic) part of the loop, and increased by more distal stimulation, which lengthens the afferent part of the loop. In contrast, the latency of M-waves, and of any of their late components if they are dispersed, increases with more proximal stimulation and decreases with more distal stimulation. Considerable progress has been made in refining the methodology and techniques and in defining the clinical applications of F-waves since their description in humans (Magladery and McDougal, 1950). The initial reports of the clinical use of F-wave measurements (Kimura, 1974; Kimura et al., 1974, 1975; Conrad et al., 1975; King and Ashby, 1976; Panayiotopoulos and Scarpalezos, 1977; Panayiotopoulos et al., 1977; Eisen et al., 1977a, 1977b) were followed by a number of criticisms regarding, in particular, the variability of the responses and the assumptions made in the calculations of conduction velocities (Bradley, 1978). Since those initial controversies, and with further experience and work in this area, the technique has gradually become widely used in departments of clinical neurophysiology throughout the world, and is considered by most to be a routine procedure in nerve conduction studies. It is now accepted to have a particular value in assessing motor conduction in proximal segments of peripheral nerves, including plexii and nerve roots. It seems also to be the most sensitive technique to detect motor conduction abnormalities in peripheral nerve diseases because of the length of the

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peripheral motor pathways tested (Kimura et al., 1979; Andersen et al., 1997; Kohara et al., 2000). The latency variability, once thought to be a major limitation for clinical use (Kimura, 1983b), can be measured and has been called chronodispersion (Panayiotopoulos, 1979). This latency variability of F-waves can be used to look at the spectrum of conduction velocities of motor axons of a population of motor units in a muscle (Guiloff and Modarres-Sadeghi, 1991), which has been called tacheodispersion (Chroni and Panayiotopoulos, 1993a, 1993b). Latency variability seems to be a very sensitive parameter of disease in peripheral nerves (Weber, 1998), as predicted initially (Panayiotopoulos, 1979). 9.2.2. Historical background F-waves were described in humans by Magladery and McDougal (1950). They recorded from eight young adults with surface electrodes, or unipolar needles insulated to the tips, from the hypothenar eminence, short flexor muscles of the foot, or from antero-lateral or calf leg muscles. They stimulated with monopolar cathodal shocks of about 0.5 ms duration, at not less than 1 Hz, six to twenty times. On stimulating the ulnar nerve they found that, in addition to the M-wave, a smaller potential change appeared later, at about 30 ms latency. This potential tended to increase in amplitude with increasing strength of the stimulus up to supra-maximal stimulus intensities for the M-wave. This distinguished it from the H-wave, described by Hoffman (Hoffman, 1922), which decreased in amplitude as the stimulation strength increased. This Fwave, as Magladery and McDougal called it, could have a large amplitude or could be apparent only at relatively high magnification. In one of the examples shown in their paper the F-wave has a peak to peak amplitude of about 0.1 mV (their Fig. 9.5). It is believed that the name F-wave was invented by Magladery and McDougal because these late responses were easily recordable from the foot muscles (Kimura, 1989). Magladery and McDougal remarked that the Fwave varied in prominence in different subjects, that in any one subject they were remarkably constant in their time relationships, and that as opposed to the direct muscle response, its latency increased as the stimulation point was closer to the muscle being recorded, indicating that it was an indirect response with an afferent and an efferent arm. They observed that they were absent if the nerve was stimulated distal to a complete procaine nerve block at the elbow and

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concluded that the F-waves resulted from impulses discharged from the central nervous system. They thought that F-waves were more likely to represent a true reflex response rather than motor neuron discharges caused by antidromic stimulation of the motor axons (Magladery and McDougal, 1950). Previous observers had also noted, but not named, these late responses, which had been either disregarded or considered reflex in origin (Hoffman, 1922; Renshaw, 1941; Kugelberg, 1946). Renshaw (1941) had shown in the cat that a centrifugal response in the nerve to the semimembranosus could be elicited by antidromic stimulation, when the dorsal roots had been severed and the caudal cord transected. He showed that the centrifugal response was nearly abolished by a preceding shock applied to the dorsal columns, probably the first demonstration of inhibition of this late response by a change in the excitability of motorneurons. Further, Renshaw calculated that the delayed efferent volley occupied only 2–3% of the number of fibers involved in the antidromic volley that evoked it. This figure is remarkably close to those obtained with calculations of the proportion of the motor neuron pool producing each F-wave in humans (1.2–5%), made much later, using the ratio of amplitudes of the F-wave to the associated M-wave (F/M ratio) (see, for example, Kimura et al., 1984; Fisher, 1992; Fisher et al., 1994). Dawson and Merton first suggested that F waves represented motor neuron discharges produced by antidromic volleys as they found no difference between the afferent and efferent conduction velocities of the F-response over long segments of the ulnar nerve (Dawson and Merton, 1956). Hagbarth reported that during post-tetanic stimulation in normal human subjects, F-waves in the small muscles of the hands are replaced by characteristic H-waves (Hagbarth, 1962). Gassel and Wiesendanger (1965) showed in the cat that the early component of the late F-response was antidromically induced as it was not abolished by deafferentation, whilst the late component was a polysynaptic reflex. At low stimulation intensities, or following repetitive stimulation, a monosynaptic reflex could replace or superimpose itself to the antidromic induced recurrent discharge. Thorne used single motor unit recordings with bipolar electrodes in humans and showed that single unit M- and F-responses were identical, indicating that centrifugal impulses generated centrally occupied only the axons that were antidromically stimulated. However, in spastic patients, the F-response showed a

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variable late polyphasic component with features of a reflex response superimposed on the recurrent one (Thorne, 1965). McLeod and Mayer showed that F-waves were present after dorsal rhizotomy in baboons and man respectively (McLeod and Wray, 1966; Mayer and Feldman, 1967). Miglietta showed the disappearance of H-waves and persistence of F-waves in the foot in four patients undergoing dorsolateral myelotomy, a procedure that transects the zone immediately below the dorsal grey matter preventing the inputs to the motor neurons coming from the dorsal horns (Miglietta, 1973). Using single fiber electromyography Trontelj showed by way of threshold stimulation of the human median nerve that a single muscle fiber F-response only appeared if there was a preceding single muscle fiber M-response, that is, if there was direct activation of the motor axon. He also showed that the latency variation (jitter) of the M single fiber response was only slightly less (by 10–30 μs) than that of the F single fiber responses that followed those M-responses, thus excluding the possibility that synaptic transmission is involved in the generation of the F-wave. In contrast, the difference in latency variation was much larger in single muscle fiber H-waves than in Mresponses. However, he also found typical reflex responses, with features resembling those of single muscle fiber H-waves, in muscles of the hand and arm, especially on voluntary contraction. He concluded that neither the F, or the H-waves can be regarded as pure recurrent or pure reflex responses (Trontelj, 1973). Late responses with features of Hwaves were also seen in the hand of spastic, but not normal, patients studied with single muscle fiber recordings (Schiller and Stalberg, 1978). The F- and H-responses of a single motor unit have the same shape and a double stimulation method, which cancels the F-waves, is needed to distinguish them (Roth and Soichot, 1995). Observations on the changes in F-wave parameters in patients with various conditions, including dorsal rhizotomy and peripheral neuropathy were made early (Mayer and Feldman, 1967). The clinical use of Fwaves was first reported in the literature in 1974 (Kimura, 1974; Kimura et al., 1974) (Fig. 9.1). 9.2.3. Generation of F-waves An understanding of the basic physiology of F-wave generation is important when devising the techniques to be used clinically, the parameters to be measured

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and their significance in interpreting the results obtained. 9.2.3.1. Motor neuron discharges Animal studies indicate that recurrent discharges of motor neurons when stimulated antidromically are generated in the non-myelinated proximal segment of the axon (hillock) from where they spread distally. For such recurrent discharges to occur, there must be enough delay of transmission of the antidromically induced depolarization across the motor neuron axonsoma junction to allow repolarization of the axon hillock (Brock et al., 1953; Eccles, 1955). Suprasegmental and segmental excitatory and inhibitory influences contribute to the resting excitability state of motor neurons. F-waves are more readily elicitable when the muscle is voluntarily contracted or with the Jendrasik manouver (Fraser and Olney, 1992; Doherty et al., 1994). The mean size and persistence of F-waves increases in spastic patients (Eisen and Odusote, 1979; Milanov, 1992a, 1992b). Subthreshold transcranial magnetic conditioning stimulation, appropriately timed, in normal human subjects resulted in increased amplitude, persistence and duration of F-waves in abductor pollicis brevis and extensor digitorum brevis muscles (Mercuri et al., 1996). Reductions in F-wave persistence, amplitude and small increases in minimal latency have been described in the abductor pollicis brevis muscle of normal human subjects during sleep (Ichikawa and Yokota, 1994). During cataplexy F-waves are absent (Yokota et al., 1992). F-wave size in the abductor pollicis brevis of 10 patients with spasticity was significantly reduced by stimulation, or lesioning, of the dentate nucleus (Fox and Hitchcock, 1982). These studies suggest that supra-segmental influences can both increase and decrease the resting excitability state of human spinal motor neurons. When a mixed nerve is stimulated antidromically a variety of segmental excitatory and inhibitory postsynaptic potentials are generated in each motor neuron. These include post-synaptic potentials mediated by sensory afferents (Eccles et al., 1957; Fetz et al., 1979), Renshaw cells (Renshaw, 1941; Willis, 1971) and direct connections between motor neurons (Gogan et al., 1977). The range of conduction velocities in a mixed nerve is wide in both motor (Borg et al., 1978) and sensory fibers (Burke et al., 1983). Therefore, the different excitatory and inhibitory impulses arrive at different times to a variety of motor neurons so that the relative balance of excitatory and

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Normal Study Site of stimulation

Left Median Nerve

C.J. Aug 23,1973 Left Ulnar Nerve F

F

Wrist M M

F F

Elbow

M

M Axilla

M

M

F(Axilla)

F(Axilla)

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Fig. 9.1 Normal F-responses in human thenar muscles and abductor digiti minimi with stimulation at wrist, elbow and axilla. Normal M-response (horizontal brackets) and F-wave (small arrows) recorded from the thenar muscles (left half of the figure) and from abductor digiti minimi (right half) through surface electrodes. Sites of supra-maximal stimulus to the median and ulnar nerves are shown. Three consecutive traces are superimposed for each tracing. As the stimulus was moved from the wrist to the elbow, the latency of the M-response increased whereas that of the F-wave decreased. With stimulation at the axilla, the F-wave occurred before the completion of the M-response and was not clearly visible. In the bottom tracings, this difficulty was avoided by delivering a simultaneous second stimulus at the wrist, which blocked the M-response elicited by axillary stimulation. The F-wave, F (axilla), elicited by axillary stimulation could now be detected because it was clearly separate from the M-response, M (wrist), elicited by wrist stimulation. Reproduced from Kimura, J. et al. (1974) Neurology 24: 539–546, with permission.

inhibitory post-synaptic potentials is continuously changing in time and space and, accordingly, the excitability of individual motor neurons will also be changing continuously during the arrival of the antidromic volley. Whether a motor neuron generates a recurrent response or not will depend on a critical range of membrane depolarization when the antidromically induced depolarization arrives at the axon-soma junction (Eccles, 1955). Experimentally, recurrent responses can be prevented by excessive hyperpolarization, which leads to soma-axon blockage, by excessive depolarization that results in somaaxon delay, and by collision of the antidromic impulse

with an orthodromic discharge (Lloyd, 1943; Eccles, 1955). All these mechanisms may operate in humans, each accounting, probably in different degrees, for the observed fact that only a small proportion of the motor neuron pool (1–5%) generates F-waves (Kimura et al., 1984). For example, there is evidence in humans that F-wave latency increases, and amplitude and duration decreases, with preceding conditioning sensory stimuli probably activating A delta afferent fibers. This suggests that antidromic activation of motor neurons is inhibited by such sensory stimuli with an associated decrease in the excitability of the motor neuron pool (Fisher, 1991).

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9.2.3.2. What is in a conventional surface recorded F-wave? 9.2.3.2.1. Only recurrent motor neuron discharges? It is accepted that F-waves in man largely represent recurrent discharges of motor neurons that have been stimulated antidromically, as was discussed above. However, the matter is not straightforward on current evidence. It remains likely that some, or part, of the responses classified as F-waves in a clinical surface recording are, or contain, reflex induced superimposed discharges of motor neurons to the ascending volley activating both motor and sensory fibers in a mixed peripheral nerve. In animals reflex motor neuron responses are diminished, but not abolished, by a preceding antidromic response (Lloyd, 1943; Brooks et al., 1950) because although some motor neurons invaded antidromically become refractory to afferent volleys, others whose antidromic impulses have been blocked by collision may become more excitable synaptically and are activated reflexly. Afferent sensory stimuli may evoke orthodromic responses as early as 0.1–0.7 ms after an antidromic volley reaches the axon soma junction, but these potentials may have a low chance of occurrence because the size of the excitatory postsynaptic potentials mediated by slower conducting sensory fibers in the cat is much smaller (Mendell and Henneman, 1971; Scott and Mendell, 1976). Following acute deafferentation in the cat, the 57% decrease in the amplitude of F-waves observed was attributed to abolished reflex discharges (Fox and Hitchcock, 1987). The antidromic discharges and the reflex components of the late responses may be mediated by fibers with different conduction velocities (Burke et al., 1989). The latency of such superimposed reflex discharges would be within the range of observed F-wave latencies (Thorne, 1965; Gassel and Wiesendanger, 1965; Mayer and Feldman, 1967; Trontelj, 1973; Guiloff and Modarres-Sadeghi, 1991). Some evidence that reflex discharges may prevent F-waves in low threshold motorneurons has recently been presented (Espiritu et al., 2003). It would be expected that if the excitability of the motor neuron pool increases, such reflex components of the F-wave would also increase. Indeed, such has been shown to be the case with upper motor neuron lesions both in animals (Gassel and Wiesendanger, 1965) and in man (Thorne, 1965; Schiller and Stalberg, 1978). Further, F-waves in the small muscles of the hand in normal human subjects were substituted by H-waves during post-tetanic stimulation (Hagbarth, 1962).

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A method to cancel single unit F-waves with double stimulation may clarify the nature of single motor unit spinal responses that have the same shape whether elicited with single stimuli above threshold or with maximal intensity, such as heteronymous H- reflexes (Roth and Soichot, 1995). It has been proposed that constant component waves within the F-waveforms are A-waves (Bischoff et al., 1996). This is considered in the section on A-waves (see below). 9.2.3.2.2. How many motor units are contained in a surface recorded F-wave? It is clear that most surface recorded F-waves in normal subjects represent the recurrent discharges of two or more motor neurons. Thorne first identified upto 3 single motor units in an F-wave in ulnar innervated muscles using a bipolar needle electrode by comparing them with the single motor units seen in the direct M-response (Thorne, 1965). Simultaneous bipolar needle and surface recordings in the human abductor digiti minimi muscle of normal subjects have been helpful (Guiloff and Modarres-Sadeghi, 1991) (Fig. 9.2). It has been possible to measure, in the same 8 subjects, single motor unit F-responses to near threshold stimuli at the elbow, both intramuscularly and with surface electrodes, as well as the conventional surface recorded F-waves to supra-maximal stimulation. Single motor units, recruited voluntarily, were also recorded simultaneously with a bipolar needle and surface electrodes in the same muscle of another 8 subjects who also had conventional F-wave recordings. The average number of motor units per F-wave turned out to be similar using two different methods to calculate it. Dividing the mean amplitude of the conventional surface recorded F-wave by the mean amplitude of the surface recorded voluntary single motor units gave a mean number of 2.8 (SD2.2), with a range of 1.3 to 7.9, motor units per F-wave. If the conventional surface F-wave amplitude, as a proportion of the compound muscle action potential (cMAP), is divided by either the surface amplitude of the F, or of the voluntary single units, both also as a proportion of the cMAP, the average number of motor units per F-wave was the same, 2.3. A close positive correlation between the estimated number of motor units contributing to the cMAP and the mean number of motor units per surface recorded F-wave was found in the abductor digiti minimi of the 8 normal subjects (Guiloff and Modarres-Sadeghi, 1991).

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M

A

F

M

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1 2 3 4 5 6

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1

0.1 mV 5 ms 2

Fig. 9.2 Intramuscular and surface recorded single motor unit F-responses. Examples of surface recorded single motor units with identical orthodromic (M-SUs) and F-responses (F-SUs). M = orthodromic response; F = Fresponse. Stimulation of the ulnar nerve at the elbow; recordings from the abductor digiti minimi with a bipolar needle (BNE, left) and surface (SE, right) electrodes. (A) Single motor unit with (4, 5, 6) and without (1, 3) its F-wave in a 23-year-old normal female. Both BNE and SE recorded responses appeared in an all-or-none manner (1, 2). The size and shape of the SE recorded M-response varied (1, 3, 4, 5, 6); it was identical to its F-response in record 6. The stimulus intensity was constant at 8 mA for this unit. (B) Single motor unit in a 54-year-old normal male with (1) and without (2) its F-response. In this example, SE recorded M- and F-responses were identical but did not occur at the same time. Reproduced from Guiloff, R., Modarres-Sadeghi (1991) Brain, 114: 1771–1801, with permission from Oxford University Press.

9.2.3.3. What do F-wave conduction velocities measure? The population of F-waves can be seen as a sample of the population of motor neurons innervating a muscle and their axons. Thus, the estimated conduction velocities of all the individual F-waves in a representative sample of the population of F-waves should be a sample of the conduction velocities of the population of

motor axons in a nerve. The latencies of surface recorded F-wave are measured to the onset of each F-wave. Thus, the fastest of the individual component motor units forming an individual surface recorded F-wave (on average 2–3, see above) is measured. The first problem is whether the maximal conduction velocity, estimated by the minimum F-wave latency, represents conduction in the fastest motor axon. Such seems to be the case on current evidence. Kimura showed that the antidromic maximal conduction velocity of the F-wave is the same as the orthodromic conduction velocity of the M-potential in a given nerve segment (Kimura, 1974). The conduction velocity of the F-wave, as measured with the shortest F-wave latency, is similar to the maximal motor conduction velocity (Panayiotopoulos et al., 1977; Barwick and Fawcett, 1988; Guiloff and ModarresSadeghi, 1991; Nobrega et al., 2000). It has been assumed that the motor unit indexing the latency to the onset of the compound muscle action potential is often the same as, or has a similar CV to the fastest F-wave obtained with 50 or more supra-maximal stimuli (Burke et al., 1989). Single unit studies support this assumption; the fastest recorded orthodromic units had a similar latency to that of the CMAP and, in the same subjects, the latency of the associated surface recorded single unit F-response was only 0.4 ms shorter than the shortest conventional F-wave (Guiloff and Modarres-Sadeghi, 1991). The second problem is whether the spectrum of conduction velocities of F-waves, obtained by using the latency to the onset of each F-wave to calculate it, represents the spectrum of conduction velocities of the motor axons of a peripheral nerve (Schiller et al., 1979). Using a collision technique, Kimura first showed that if the faster motor fibers are blocked, F-waves are also produced by the slower ones (Kimura et al., 1984) (Fig. 9.3). The range of conduction velocities obtained with the range of F-wave latencies in abductor digiti minimi (Schiller and Stalberg, 1978; Guiloff and Modarres-Sadeghi, 1991) and abductor pollicis brevis (Burke et al., 1989; Felice, 1998), of normal individual subjects is similar to the range of conduction velocities of motor fibers calculated by some authors (Thomas et al., 1959; Leifer et al., 1977) and less (Lee et al., 1975), or greater (Hopf, 1962; Ingram et al., 1987) to that calculated by other authors. In general, the spectrum of velocities of the pooled F-wave data agrees well with the spectrum of velocities calculated by other methods, including collision (Hopf, 1962; Dorfman, 1984; Ingram et al., 1987),

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M(E)

1.6

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M(W)

S

F(W) S(E) S(W)

F(E)

M(W)

1.4 2.2 F(W)0

200 μV 5 ms

200 μV 5 ms

Fig. 9.3 Slow and fast motor fibers mediating F-waves. Supra-maximal stimulation, S (E), at the elbow (top), S (W), at the wrist singly (middle), and S (E) and S (W) combined bottom). The orthodromic impulse of S(E) and the antidromic impulse of S(W) collided in the slow (S) but not fast (F) conducting fibers, allowing differentiation between F(E) and F(W), representing S-and F-fibers, respectively. Reproduced from Kimura, J., et al. (1984) Muscle Nerve, 7: 392–399. (With permission from John Wiley and Sons, Inc.)

deconvolution of cMAP (Dorfman, 1984), reconstruction of cMAP (Lee et al., 1975), and single unit studies (Juul-Jensen and Mayer, 1966; Dalpozzo et al., 2002). It seems clear that when at least 60 supra-maximal stimuli are delivered to a motor or mixed nerve in normal subjects the evoked surface F-waves recorded have a wide spectrum of conduction velocities that roughly samples the spectrum of conduction velocities of the motor fibers of a peripheral nerve. This is one basis for looking at chronodispersion and tacheodispersion (see later) as parameters of interest. 9.2.3.4. Populations of motor neuron generating F-waves in humans The ratio of amplitudes between the M-response and the mean F-response indicates that only 1–5% of the motor neuron pool discharges recurrently to a supramaximal antidromic volley that stimulates all the motor and sensory axons of a peripheral nerve. If this is the case, why does only a small proportion of motor neurons discharge recurrently with each stimulus? And why is it that, with such small proportion discharging recurrently each time, it requires very few antidromic stimuli (10–20), in such

a large proportion of subjects tested, to obtain an F-wave equally fast, or very similar, to that obtained with 100 or more antidromic stimuli or to the maximal velocity obtained with the M-response (Kimura, 1974; Marra, 1987; Guiloff and Modarres-Sadeghi, 1991; Taniguchi et al., 1993; Fisher et al., 1994; Chroni et al., 1994, 1996; Nobrega et al., 2001). Is it that the 1–5% of all motor neurons stimulated antidromically is always the same pool of motor neurons generating the recurrent responses or can all motor neurons produce them if a large enough number of antidromic stimuli are delivered? If all motor neurons can produce recurrent discharges, do certain groups of motor neurons generate them more frequently than others? Given the way the F-responses are elicited in clinical practice, do all motor neurons generating recurrent responses have an equal chance of conducting the response through their axons so that they can be finally recorded in the muscle? Are the F-waves a representative sample of the population of motor neurons and their axons innervating a muscle, or do they represent all motor neurons but in a “biased” way, i.e., certain groups being represented more frequently than others?

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The above problems have been addressed both with conventional surface and single motor unit recordings of F-waves. By plotting F-wave latencies as a difference with the fastest in each of 5 control subjects it was shown that a large proportion (26%) of the F-waves had latencies within 1 ms of the fastest F-wave. The group and individual histograms did not have a normal distribution and it was concluded early that “there was a tendency for fast F-waves to be elicited more easily than slow F-waves” (Panayiotopoulos, 1979). Kimura et al. (1984) showed that using supra-maximal and sub-maximal stimuli did not change the minimal latency or duration of the surface recorded F-waves but there were significant reductions in amplitude and persistence. Using a collision technique (Fig. 9.3) that allowed a separation between low and higher stimulation threshold motor fibers they showed that if the faster motor fibers are blocked, F-waves are also produced by the slower ones. Further, there was no significant difference in amplitude or persistence of F-waves in both groups of fibers. Interestingly, there was no consistent change in the F-wave minimum latency in progressively slower conducting fractions of the motor neuron pool. The results indicated that large populations of motor neurons produce F-waves and that slow and fast fractions of the motor neuron pool were equally susceptible to recurrent discharges (Kimura et al., 1984). The results did not answer the question as to whether recurrent responses of faster conducting fibers were recorded more frequently than those of slower conducting axons. However, the constancy of the F-wave minimal latency in high- and low-threshold fibers, and within the low threshold group, are suggestive of more frequent generation of F-waves in the faster conducting pools. Single unit studies are also consistent with the notion that F-waves are generated by all motor neurons. With these techniques motor units with lowstimulation threshold and faster conduction velocity are usually preferentially sampled, though in one study smaller motor neurons were preferentially sampled (Dengler et al., 1992). One study with bipolar needle electrodes in abductor digiti minimi and threshold stimulation of the ulnar nerve showed that single unit F-waves were also produced by motor neurons with conduction velocities of up to 15 m/s less than the maximal conduction velocity for such individual. (Guiloff and Modarres-Sadeghi, 1991). No relationship between the twitch force of motor units and their capability to produce F-waves was found

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using intramuscular micro-stimulation of single axons and surface recordings in the first dorsal interosseous muscle nerve (Dengler et al., 1992). Twitch force is closely related to motor neuron size. No difference in conduction velocities was found between 7 thenar units that produced F-waves and 37 that did not in a study using intra-neural axonal stimulation of the median nerve (Thomas et al., 2002). Burke and co-workers studied the spread of F-wave latencies in abductor pollicis brevis, tibialis anterior and soleus muscles using at least 50 supramaximal stimuli. They suggested that, while F-waves may occur in all motor neurons in many motor neuron pools, they appear preferentially in high threshold motor neurons and rarely occur in slowly conducting motor units (Burke et al., 1983). The question of preferential (i.e. more frequent) generation of recurrent responses by certain groups of motor neurons has been systematically addressed in one study (Guiloff and Modarres-Sadeghi, 1991) with surface conventional and single unit intramuscular and surface recordings. It was found that if the F-wave velocities were expressed in each individual as a difference from the maximal F-wave conduction velocity in that individual, the frequency distribution of conduction velocities of the F-waves in a group of 33 normal subjects was shifted towards faster values than what would be expected from studies of the distribution of conduction velocities in single peripheral nerves (Fig. 9.4). Such way of expressing the frequency distribution of the minimal latency of F-waves had been introduced when chronodispersion was described (Panayiotopoulos et al., 1977; Panayiotopoulos, 1979). Panayiotopoulos had already found in this way that the frequency distribution of F-wave latencies was not normal in individuals, nor in the grouped normal subjects or patients, and had already pointed out that faster F-waves appeared to be elicited more frequently (26% of the F-waves were within 1 ms of the fastest F-wave in the control subjects). It was also reported subsequently that the distribution of the latencies of F-waves of 17 individual normal subjects was not normal in the majority (Petajan, 1985). In a study of 11 normal subjects, much later, it was reported that the F-wave latencies seemed to be distributed normally (Fisher et al., 1994), supporting the use of the mean F-wave latency as a clinical parameter by this author. The reason for the difference between this last study and the previous three may lie on the fact that plotting the frequency distribution of absolute F-wave latencies in a population, without

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59

65

Fig. 9.4 Frequency distribution of estimated conduction velocities (CVs) in surface recorded F-waves. (A) Distribution of the proportion of estimated F-wave CVs (n = 1692) with values within 10% of that of their maximum CV in 33 normal subjects. For example, in each of 8 patients 80–89% of their F-wave CVs, evoked by 60 stimuli, are within 10% of their respective maximum F-wave CV. (B) Distribution of the difference in F-wave CVs from the maximum F-wave CV (standardized as 0 in each subject) in 33 subjects. For example, each of the 98% F waves out of a total of 1692 had CVs within 10 ms of their subject’s maximum F-wave CV. (C) Relative frequency distribution of estimated F-wave CVs (n = 1692) within the subject range of F-wave CVs (33 subjects). The group means and 95% confidence intervals are given. The number of subjects who had F-waves with CVs within each 2 ms bin is also shown. Reproduced from Guiloff, R., and Modarres-Sadeghi (1991) Brain, 114: 1771–1801, with permission from Oxford University Press.

normalizing the values in each individual to the fastest F-wave in that individual, will hide the true relative distribution of latencies in that population because of the wide spectrum of fastest motor velocities in normal subjects. There is no firm evidence to suggest that there is a much larger number of axons with faster conduction velocities in a peripheral nerve. It can be calculated from the data of Dorfman et al. (1982) that the fastest conducting motor fibers, defined as those within 10% of the maximal conduction velocity constitute about 20% of the total number of fibers of a peripheral nerve. Using such data, and assuming an equal chance for each motor axon to mediate an F-response and three motor units per F-wave, it can be calculated that the expected, theoretical, proportion of F-waves velocities within 10% of the maximum conduction velocity should be 49%. However, the mean from the individual data in 33 normal subjects was much higher at 65%. The pooled F-wave data in the same study

(Guiloff and Modarres-Sadeghi, 1991) also showed that the proportion of F-waves conducted by faster fibers was 2–3 times higher than expected from the proportion of faster fibers calculated for peripheral nerves using a collision or deconvolution technique (Dorfman, 1984). Comparisons of the maximum and minimum conduction velocities of the fibers that generate the M-wave with the F-waves elicited by ulnar nerve stimulation also led to the conclusion that the F-waves registered were preferentially generated by the fastest conducting axons (Nobrega et al., 2000). Further, single unit studies showed that, within the population of motor units sampled by threshold stimulation, there was a positive correlation between the conduction velocity of single motor units and the frequency with which they produced F-responses to 200 antidromic stimuli (Guiloff and Modarres-Sadeghi, 1991). Dengler and his co-workers did not study the frequency of F-responses in each of their 35 units in 15 subjects that produced F-responses out of 209 units

198

studied in 21 subjects, and only gave 20–40 stimuli to each unit. Their units had a narrow range of latencies, between 30 and 35 ms (Dengler et al., 1992) and they remarked that three out of four very large motor neurons produced F-waves. Interestingly, of the 7 units with F-wave responses out of 44 thenar units reported by Thomas et al. (2002) five had conduction velocities above, and two below, 60 m/s. These last authors did not report the frequency of F-responses generated by each of these units and used stimulation frequencies of up to near 30 Hz. The following mechanisms have been postulated to explain the more frequent occurrence of F-waves and single unit F-responses produced by faster motor units (Guiloff and Modarres-Sadeghi, 1991). (1) Greater chance of axon-soma block in smaller motor neurons. Recurrent inhibition would affect more smaller than larger motor neurons because by the time the antidromic impulse arrives to the smaller motor neurons the Renshaw cell recurrent inhibition, triggered by the discharge of larger motor neurons following the earlier arrival of antidromic impulses mediated by fast fibers, would have already taken place. (2) Greater chance of collision in smaller motor neurons. The early arrival of sensory (Ia) excitatory impulses with antidromic stimulation and the balance of other excitatory and inhibitory influences on the small motor neurons will determine whether they discharge before the arrival of the motor antidromic volley. If they do then the orthodromic discharge would cancel the slow incoming antidromic impulse and no F-wave would be generated. (3) Greater chance of shortening of axon-soma delay in smaller motor neurons. There is some evidence that shortening of the axon-soma delay is more likely to occur in smaller motor neurons (Barakan et al., 1949) and this would reduce their chance to discharge after invasion by an antidromic stimulus. The three mechanisms would affect all motor neurons and contribute to limit to about 1% of the motor neuron pool, those producing recurrent responses, when measured either by amplitude criteria in conventional F-wave recordings (Kimura, 1984, 531/id), or as a proportion of single unit F-responses per threshold stimuli (Guiloff and Modarres-Sadeghi, 1991). However, these mechanisms would affect more the smaller motor neurons resulting in more

R.J. GUILOFF

frequent occurrence of F-responses in faster motor neurons. The important finding of a similar (1%) proportion of the motor neuron pool generating F-responses when fast and slow fibers were tested with a collision technique (Kimura et al., 1984) (Fig. 9.3) suggests that the above mechanisms result in a constant fraction of the motor neuron pool being recorded as recurrent responses, but preferential (more frequent) occurrence of F-responses in the faster motor neurons within any motor neuron pool would still occur. The following facts are all consistent with this view: (1) The fastest F-wave is often recorded with as little as 10–20 stimuli. (2) Shift to faster velocities in the frequency distribution of F-wave conduction velocities. (3) Positive correlation between F-response frequency and conduction velocity in low threshold single units. (4) Little variation in mean F-wave latency when the slower fibers are progressively blocked (Kimura et al., 1984; Fisher, 1985). (5) Similar mean minimal latency of F-waves with supra-maximal and sub-maximal stimulation. (Kimura et al., 1984). 9.2.3.5. Repeaters (Figure 9.5) The use of the term “repeater F-wave” needs to be standardized. It has been used for surface recorded F-waves with a shape that appears more than once when a number of stimuli are given. These F-waves can have different shapes, each of which can repeat more than once. Therefore, the proportion of repeaters in an F-wave study can be expressed as percentage of repeater F-wave shapes or percentage of repeater F-waves (Fig. 9.5) (Peioglou-Harmoussi et al., 1985a). The mean percentage ±SD of repeater F-waves in normal subjects using 100 stimuli was 17.3 ± 16 for abductor pollicis brevis (Macleod, 1987). For abductor digiti minimi using 60 stimuli it was 19.5 ± 8.1% and that of repeater shapes was 9.9 ± 4.4% (Guiloff and Modarres-Sadeghi, 1991), whilst in another study using 200 stimuli, the mean percentage of repeater shapes in the hypothenar muscle was 3.4 ± 1.1% (Peioglou-Harmoussi et al., 1985a). The term repeater F-wave has also been used for frequent occurrences of the same single motor unit F-wave in single motor unit needle recordings of M- and F-waves of identical shape (Roth and Soichot, 1995). Single motor unit F-wave recordings with threshold stimulation or just above, should not be equated to the surface recorded F-wave repeater waves to supra-maximal stimulation described by a number of authors (Peioglou-Harmoussi et al., 1985a; Macleod, 1987; Guiloff and Modarres-Sadeghi, 1991).

LATE RESPONSES (F- AND A-WAVES): METHODOLOGY AND TECHNIQUES

A M

B

C

199

D

F a

c d

a c

b b

d b 50 μV a 5 mV 0.2 mV 5 mV

5 ms

5 ms

Fig. 9.5 F-waves evoked by 60 stimuli in normal subjects. (A–C), F-waves recorded over the abductor digiti minimi following 60 consecutive stimuli at 1 Hz at the wrist in a healthy male subject aged 46. Four repeater F-wave shapes are shown (a, b, c, d). A total of 10 repeater F-waves are seen. In this example: measurable F-waves = 58; F-wave frequency = 96.6%; total F wave shapes = 52; percentage repeater F-wave shapes = 7.7%; percentage repeater F-waves = 17.2%. (D) All the F-waves with an amplitude <40 μV, evoked by 60 stimuli in a male subject aged 31 are shown. Larger F-waves are not shown. The compound muscle action potentials are shown at the bottom. M = M-wave. Reproduced from Guiloff, R., Modarres-Sadeghi (1991) Brain, 114: 1771–1801, with permission from Oxford University Press.

The frequency of occurrence of F-waves in single motor unit can be high, with reports of a mean 4.5% (up to 21% in one unit) in one study using 200 stimuli (Guiloff and Modarres-Sadeghi, 1991), and a mean of 10.6% (up to 49%) in a study with variable but not stated number of stimuli (Roth and Soichot, 1995). The very high frequency of F-waves in this last study was acknowledged to be due to selection bias. A mean frequency of 23% (up to 45% in one subject) was quoted in another study using 100 stimuli of variable intensity, and a different definition of unit F-responses, which included surface recorded responses that were assumed to contain only one MUP (Yates and Brown, 1979), When these last authors considered single F units

obtained with stimulation at or just above threshold, they obtained a maximum F frequency of 10% in control units. In contrast, the frequency of surface recorded F-wave repeater shapes is much less, with a mean of 3.4% (up to 5.5% in 2 of 21 normal subjects who had 11 repetitions of the same shape after 200 stimuli) in one study (Peioglou-Harmoussi et al., 1985a). Only a mean of 0.4% ± 2.2 of 138 different repeater shapes out of a total of 1556 F-waves in 30 normal subjects repeated 5 or more times in another study using 60 stimuli (Guiloff and Modarres-Sadeghi, 1991). Further, there is evidence to suggest that surface recorded repeater F-waves are composed of several motor units (Guiloff and Modarres-Sadeghi, 1991) (see below).

200

A comparison of surface recorded repeater (n = 304) with non-repeater F-waves (n = 1252) obtained in the abductor digiti minimi of 30 normal subjects with 60 stimuli showed no significant differences in mean minimum F-wave latency, mean maximal conduction velocity and mean duration between the two groups. The distribution of conduction velocities was also similar for repeater and non-repeater F-waves. However, both amplitude and area of repeater F-waves were significantly larger than in the non-repeater F-wave group (Guiloff and Modarres-Sadeghi, 1991). These results were independently confirmed in a later study in the abductor pollicis brevis of 11 normal subjects where mean latencies of repeaters and non-repeaters were similar but the amplitude of repeaters was significantly larger (Fisher et al., 1994). Thus, it is clear from the latency, conduction velocity, distribution of conduction velocities and duration results of surface recordings, that the range of motor units represented in the repeater F-wave group is as wide as those in the non-repeater group. It has been suggested that a specific group of motor neurons that are more prone to recurrent discharges generate repeaters (Kimura et al., 1984). The greater amplitude, but similar latency and CV of repeaters compared with non-repeaters is a strong argument against repeaters being preferentially generated by very large single motor neurons. If so, their latency should be shorter and their velocity faster. If anything, the greater amplitude suggests that repeaters may, in fact, contain more motor units than the average F-wave. The latency of repeaters also excludes them being mediated by peripheral axon reflexes (Fullerton and Gilliatt, 1965). The available data support the hypothesis that repeaters are generated by groups of motor neurons that backfire together repeatedly. Assuming that each of the average 3 motor neurons generating an F-wave has an equal chance of being part of the 1% of the pool generating F-waves with each antidromic stimulus, the possibility that 3 or more motor neurons backfire together repeatedly, by chance, is far less than the 3.4–9.8% repeater shapes and 17–19% of repeater waves actually found in the studies quoted above. A biological reason to account for these frequent timed locked associated motor neuron responses to antidromic volleys needs therefore to be invoked (Guiloff and Modarres-Sadeghi, 1991). Close proximity with particular cytoarchitectonic (connections) and functional arrangements between motor neurons could be one of the mechanisms postulated. There are

R.J. GUILOFF

no data in humans but, in animals, small groups of motor neurons can lie in very close proximity (Rexed, 1954), some motor neurons connect to others through axon collaterals (Cullheim et al., 1977) and a small proportion can have a direct excitatory effect on close neighboring ones (Gogan et al., 1977). Such groups of motor neurons might be prone to discharge together following antidromic stimulation. The question as to whether some F-wave repeater shapes may be, in fact, A-waves has not been researched systematically. It has become relevant since constant shape components of F-waves have been considered to be A-waves (Bischoff et al., 1996; Puksa et al., 2003a) (See below, section on A-waves). The analysis of repeater F-waves and the interpretation of their mechanisms may be different under pathological conditions disrupting both the normal anatomy and physiology of motor units and will not be considered in detail here. For example, in conditions with loss of motor units such as amyotrophic lateral sclerosis, and in other conditions causing chronic partial denervation, an increase in the proportion and size of F-wave repeaters may be seen (Petajan, 1985; Macleod, 1987). If only a few reinnervated motor units remain in a muscle they will be the only ones to backfire, will do so more frequently, specially if hyperexcitable in the setting of pyramidal tract dysfunction, and they will have large amplitudes because of their increased territory. These frequent repeater F-waves in neurogenic disorders need to be distinguished from A-waves (see section on A-waves). Detailed systematic studies looking at repeaters in different pathological conditions are needed. 9.2.4. Methodology and techniques 9.2.4.1. Stimulation Electrical stimulation is applied to peripheral nerves supplying the muscles from where F-waves are recorded. 9.2.4.1.1. Stimulator orientation. A number of experts advise that, when eliciting F-waves, the cathode should be placed proximally to the anode to avoid anodal block of antidromic impulses (Kimura, 1989; Fisher, 1992; Liveson and Ma, 1992; Fowler, 1993; Kimura, 1993; Oh, 1993; Delisa et al., 1994; Daube, 1996). However, others point out that in routine clinical practice F-waves are elicited with cathode distal stimulation (Young and Shahani, 1978).

LATE RESPONSES (F- AND A-WAVES): METHODOLOGY AND TECHNIQUES

Anodal block is an unlikely event during routine nerve conduction studies (Dreyer et al., 1993). A study of distal versus proximal cathode stimulation in 207 median, 204 ulnar, 136 tibial, and 110 peroneal nerves in 225 subjects looked at F-wave minimum latency and persistence and stimulus intensity required (Young and Triggs, 1998). The stimulus orientation did not affect the classification of results as normal or pathological. Significant differences were found only in the median nerve with a slightly lower mean persistence (80.6% versus 82.5%) and slightly lower mean minimal latency (28.8 versus 29.3 ms). The stimulus intensity required was significantly higher for the median and tibial nerves and lower for the ulnar nerve. Amplitude of F-waves was not measured either, though incomplete anodal block could result in decreased amplitude of F-waves since surface recorded F-waves contain several motor units. For example, the average number of motor units in a normal surface recorded F-wave in the human abductor digiti minimi is about 3 (Guiloff and ModarresSadeghi, 1991). However, in another study of the median nerve in 30 healthy volunteers, minimum, maximum and mean F-wave latencies and amplitudes were unaffected by the proximal or distal positioning of the anode (Kirshblum et al., 1998) but persistence was slightly lower with distal anodal stimulation. The first of these two studies used 10 stimuli, which is not an optimum sample size for persistence (Guiloff and Modarres-Sadeghi, 1991). The second study used 20 stimuli, which is a more adequate sample size for this parameter. The current evidence suggests that for routine clinical recordings the orientation of the stimulating electrode will not be of clinical significance. It is advisable to standardize and record the stimulus orientation used. 9.2.4.1.2. Intensity. Most authors use stimuli intensities 25% above supra-maximal. Using 50% above supra-maximal did not change the mean latency, mean median amplitude or mean persistence of the F-waves obtained with sixteen stimuli in 25 healthy volunteers (Clinchot et al., 1994). Recently, lowcurrent stimulation (10–15 mA) at 0.2 ms duration and supra-maximal stimulation were compared in the median nerves of the same 30 subjects. F-wave latencies, chronodispersion and persistence were not significantly different with both techniques but F-wave amplitude was significantly larger with supramaximal stimulation (DiBenedetto et al., 2003). If

201

these results are confirmed the technique would avoid the discomfort of supra-maximal stimulation, which could be used only when amplitude measurements of F-waves were required. However, Kimura et al. (1984) had previously described that latency and duration did not change significantly with sub-maximal stimulation but persistence and amplitude decreased significantly compared with supra-maximal stimulation. Thus, the data of these two authors is concordant for minimum latency and amplitude, but not on persistence. Kimura et al. did not report on chronodispersion. Thus, on current evidence, it appears that the use of sub-maximal stimuli to calculate minimal latency and conduction velocity is safe, though proof that this is also the case in different patient populations is not available. 9.2.4.1.3. Duration. The stimulus duration is usually set at 0.2 ms. Using 0.05, 0.1 or 0.2 ms did not significantly change F-wave mean latency, mean median amplitude or mean persistence (Clinchot et al., 1994). 9.2.4.1.4. Number of stimuli. The number of stimuli used varies widely amongst different workers, from 5 or 10 to 100. In terms of reliability, i.e., test-retest reproducibility, as little as 10 responses were required to get an intra-class correlation coefficients for the mean F-wave latency of 0.99 in 30 normal subjects and of 0.96 in nine patients with neuropathy (Taniguchi et al., 1993). However, the main issue is that the optimum number of stimuli depends on the sample size required to obtain values for F-wave parameters that are accurate and representative of the motor neuron pool supplying a particular muscle. This will be discussed under sample size. The number of stimuli used also depends on patient’s tolerance, which may vary in different populations and ethnic or cultural groups. 9.2.4.1.5. Rate. There is agreement that the rate should be no faster than 0.5 Hz to avoid the effects of an earlier stimulus on a subsequent response. With paired stimuli to the median nerve in nine normal subjects there was a decrease in persistence and amplitude of F-waves during the first 5 ms of interstimulus separation. This was followed by a period of increase in these parameters between 5 and 10 ms, a further depression for up to 60–80 ms in males and 30 ms in females, and a subsequent facilitation for up to 400 ms of inter-stimulus separation (Mastaglia and Carroll, 1985). These authors concluded that the

202

recovery curve of the F-waves was similar to that of H-waves. Increasing the rate of stimulation from 0.5 to 3 Hz resulted in a decrease in mean latency and in an increase in mean median amplitude and mean persistence of F-waves in abductor pollicis brevis in a group of 25 healthy controls (Clinchot et al., 1994). It has been hypothesized that increasing the frequency of stimulation may increase “central facilitation” and result in more and faster motorneurons being activated antidromically (Clinchot et al., 1994). 9.2.4.2. Recording 9.2.4.2.1. Surface. There is agreement that the recording cathode should be placed over the motor point, or the belly, of the muscle and the recording anode over the tendon. The ground electrode is usually placed between the stimulating and the recording electrode. The display should be set with a gain of 0.2 mV per division. Usually F-waves are positively identified if larger than 10–20 μV in peak to peak amplitude. F-waves of less than 40 μV of amplitude are better demonstrated with gains of 0.1 or 0.05 mV/division (Guiloff and Modarres-Sadeghi, 1991) (Fig. 9.5). Thus, persistence depends on the gain used. The filter settings are usually similar to those used in motor nerve conduction studies, 20 Hz–2KHz. Some recommend that a better baseline is obtained using a high pass (low frequency) filter set at 100 Hz (instead of 20 Hz), making it easier to measure latencies of individual F-waves. The sweep speed is normally set to 5 ms per division for the upper limbs and 10 ms/division for the lower limbs. When chronodispersion is greatly increased, or F-waves are apparently not seen with the standard sweep speeds, such as in demyelinating peripheral neuropathies, 10 ms/division or more, and 20 ms/division or more, may be required for upper and lower limbs respectively. F-waves are recorded in clinical practice with surface electrodes over a number of muscles, including abductor digiti minimi for the ulnar nerve and abductor pollicis brevis for the median nerve in the upper limb (Tables 9.1, 9.2). In the lower limbs extensor digitorum brevis for the common peroneal nerve and abductor hallucis for the posterior tibial nerve are commonly used (Tables 9.3, 9.4). F-waves are less frequently recorded from the triangularis (Zappia et al., 1993) and nasalis muscles (Wedekind et al., 2001) for the facial nerve, extensor digitorum communis (Fisher, 1978; Zappia et al., 1993) and extensor

R.J. GUILOFF

indicis (Papathanasiou et al., 2001) for the radial nerve, deltoid for the axillary nerve (Zappia et al., 1993) and vastus medialis (Zappia et al., 1993) and vastus lateralis (Wochnik-Dyjas et al., 1996) for the femoral nerve (Table 9.5). The F-wave may be obscured by the M-wave, or is part of it, when recording from proximal muscles, such as biceps and deltoid or with very proximal stimulation (i.e., Erb’s point). A collision technique may then be used with a distal stimulus triggering an antidromic volley that blocks the descending M volley produced by proximal stimulation, but not its F-wave (Kimura, 1974). 9.2.4.2.2. Intramuscular. F-waves can also be recorded intramuscularly using single fiber electromyography (Trontelj and Trontelj, 1973; Schiller and Stalberg, 1978) and concentric or bipolar needle electrodes (Thorne, 1965; Yates and Brown, 1979; Guiloff and Modarres-Sadeghi, 1991). The sweep speed and gain used with these techniques are within the same range as those used for surface recordings. For single fiber electromyography, the filters are set to 500 Hz to 10 kHz. For concentric needle and bipolar electrodes recordings, the filter settings are usually set to 2–5 Hz to 10 kHz. With these techniques, F-waves produced by single motor neurons can be identified because the morphology of the single muscle fiber or single motor unit M-response and that of the ensuing F-response are identical. By doing simultaneous intramuscular and surface recordings the morphology of surface recorded single motor unit F-waves can also be shown to be identical to that of surface recorded single motor unit M-responses (Guiloff and Modarres-Sadeghi, 1991) (Fig. 9.2). 9.2.4.3. Clinical measurement The parameters usually measured in F-wave studies are considered below. F-wave measurements like other electrophysiological parameters will depend on temperature and on age (Fisher, 1982; Peioglou-Harmoussi et al., 1985b). All latencies are measured from the stimulus artefact to the onset of the F-wave waveform. Amplitudes of F-waves (and M-waves) are measured peak to peak. Supra-maximal stimuli can be delivered at many points along the course of a peripheral nerve. Recordings are usually carried out in distal muscles of the hand and foot but, as can be seen in Table 9.5 with normative values, recordings can be made from many other muscles, including the facial muscles, quadriceps, deltoid and extensor digitorum communis.

LATE RESPONSES (F- AND A-WAVES): METHODOLOGY AND TECHNIQUES

The measurement of F-wave latencies and velocities can be made for the whole nerve from a point of distal stimulation, such as wrist or ankle, to the spinal cord and back, or for different segments of a nerve. In the upper limbs the segments axilla-spinal cord, axillaelbow, elbow-spinal cord and elbow-wrist can be studied. In the lower limbs the segments knee-spinal cord and knee-ankle can be studied. Measuring F-wave conduction in the axilla-spinal cord segment, or axillaelbow segment requires the use of a collision technique, as the F-wave is buried in the M-wave when stimulating very proximally. A second distal stimulus collides and cancels the orthodromic (but not the antidromic) volley elicited by proximal stimulation. Therefore, there is no M-wave from this proximal site but the M-wave from the distal stimulation site and the ensuing F-wave from the proximal one can then be recorded (Kimura, 1974) (Fig. 9.1). In routine clinical practice, distal stimulation is normally used and Fwave conduction over nearly the entire course of the nerve, to the spinal cord and back, is determined. 9.2.4.3.1. F-wave shortest or minimum latency. The shortest or minimum F-wave latency indexes the fastest conducting motor fibers of a peripheral nerve (see section on F-wave conduction velocities above). This latency includes the time for the impulse to travel antidromically through the motor axons from the point of stimulation to the anterior horn, the “turn around time” taken to re-excite a motor neuron so that it backfires (Renshaw, 1941; Lloyd, 1943) (see below), and the time for the orthodromic impulse thus generated to travel back to the muscle (Kimura, 1974). Latency is heavily dependent on the distance (Panayiotopoulos et al., 1977), which in turn, is directly proportional to height (Russo, Jr., 1985). Thus, authors that do not use F-wave conduction velocities may use height to establish the range of normality (Lachman et al., 1980; Tonzola et al., 1981; Russo, Jr., 1985; PeioglouHarmoussi et al., 1985b; Buschbacher, 1999a; Puksa et al., 2003b). For clinical purposes, but also for the mean values in a group of subjects, minimal F-wave latency can be reasonably estimated with 20 consecutive supra-maximal stimuli (see below discussion on sample sizes). 9.2.4.3.2. F-wave maximum conduction velocity. This parameter was described in detail when F-waves were introduced for clinical use (Kimura, 1974; Kimura et al., 1975). The shortest F-wave latency is used in the calculations.

203

Distance measured. For the upper limbs the distance measured was from the stimulation point to the spinous process of C7 with the limb extended and abducted 90˚, pronated for measurement of the ulnar nerve and supinated for the median nerve. The distance from axilla to the C7 spinous process can be measured with an obstetric caliper. For the lower limb the distance between stimulation points at the ankle and knee to the spinous process of T12 was made. Others have used L1 (Panayiotopoulos et al., 1977). The accuracy of surface clinical distance measurements for the lower limb to T12 was good as ascertained by measurements of the surface and actual length of the nerve in five cadavers (Kimura et al., 1975). Time measured. The time used in the velocity calculation is the minimal latency of the F-wave less the M wave latency from the same stimulation point. To this time 1 ms is substracted assuming it to be the “turn around time” for the stimulus to excite the motor neuron and for the latter to backfire. The time is then divided by 2 since it represents both antidromic conduction time to the spinal cord and orthodromic (backfiring) conduction time from the cord to the stimulation point. The turn around time of 1 ms is based on animal studies (Renshaw, 1941; Lloyd, 1943) and on the fact that the absolute refractory period of the fastest human axons is about 1 ms (Kimura, 1976). There is, however no direct evidence that this is the turn around time in humans or that it is the same for all human motor neurons. Formula. The following formula described by Kimura (1974, 1978b) can be used to calculate the conduction velocity of the F-wave with the shortest latency (FwCVmax) as well as the conduction velocity of any F-wave in the limbs (FwCV). FwCV =

Distance to C7 or T1 × 2(mm) F latency–M latency–1(ms)

Controversies. The perceived inaccuracies in the measurement of distance and the assumptions made in the estimation of the central latency led some to criticise the use of F-wave conduction velocities and to prefer nomograms relating F-wave latency measurements to arm length, or height (Young and Shahani, 1978). The counter argument was that it was preferable to have some reasonable measurement of central latency than none at all (Kimura, 1978a), that errors in such measurement would have a negligible effect in the calculation of conduction velocities

Zappia et al., 931 (10) Parano et al., 93 (?)

PeiogluHarmoussi 85 (20) Males Females Burke 89 (min 50) Misra et al., 89 (10)

Russo 85 (10)

WRIST Kimura 74 (10) Eisen et al., 77 (10) Fisher et al., 78 (min 10) Right Left Lachman et al., 80 (min 10) Kimura 83 (16) Kwast 84 (20–100)1,7

Stimuli (number)

42 44.2

39.8

28

35 41.5

17.4 15.6

mean SD

7 d–14

7 d–1 m 1–6 m

20 23

1 d–28 d 9.7 1–12 m 4.7m 2–12 6.2 19–70 36.4 15.7

155

20 20 20 51

60

21–43

2 m–12 0–1 1–3 3–5 5–7 7–12 20–64

65 54 20 14 23 13 20 30

32 32 10

12–81

18 17 124

12–81

22–39

18

64

22–58 11–74

range

33 60

n

Age yrs*

27 25.2 25.9

15.4 15.3 17 18.2 19.5 24.8 27.6 29.6

26.6

26

29.1 26.6

mean

46–56 (47.8)3 16.95 14.93 61–65 (60)3 66–130 (98.2)3 16.26 25

175.5 (7.8)3 162.4 (6.1)3 157–195

150–162 163–175 176–190

142–188

Height cm

28.4

31

22.3–30

limit

1.3 1.5 1.9 2.4 32.2

1.9 1.3

0.9 0.7 1.1 1 1.1 2.1 1.6 2.1

2.2

1.9

2.3 2.2

SD

Min Latency ms

0.95

mean

0.67

SD

2.3

< 2.0

limit

Side diff-Min Lat ms

26.9

27.1

25.9 25.9

mean

limit

2.4

30.1

1.6 22.4–28.2 1.9 22.8–28.4

SD

Mean Lat ms

16.12 16.89

58.9

59.2

65.3

59.2 62.7

mean

1.5 1.7

5.3

4.7

3.9 3.9

SD

55.7

56

limit

Max Velocity m/s

F-waves. Normal values. Median nerve (surface electrodes over abductor pollicis brevis, except where indicated)

Table 9.1

69.8

81 84

mean

23.2

0.8

1.06 0.45 0.24 0.14 0.22 0.14 720 500

2402

1.6

129 98

0.1– 0.62

limit

Amplitude μV mean SD

60–100 350 60–100 354

limit

Persistence %

1.89 2.01 2.47 4.8

8.8

3.64

3 2.9 3.6 4.1 3.8

1 0.9 1.1 4.3

1.2

1.1 1.1 1.4 1.4 1.2

mean SD

10.7

Limit

Chronodispersion ms

Weber 982(10) Buschbauer 99 (10)

Doko-Guina 97 (6)

Cai & Zhang 97 (>10)

Stimuli (number)

Continued

Table 9.1

< 160 > 160, < 180 > 180 < 160 > 160, < 180 > 180

19–79

19–49

195

50–79

< 160, > 180

0–3 m 4–6 m 7–9 m 10–12 m 13–18 m 19–24 m 25–36 m 37–48 m 49–60 m 61–72 m 30

8 10 8 10 10 10 7 6 7 10 31

Height cm

24–72 h 0–3 m 4–6 m 7–12 m 1–3 4–6 7–14 18–30 0–72 m

6–12 1–2 2–4 4–6 6–14 24 h–30

25 24 22 20 21 168

mean SD

25 20 20 20 20 21 22 19 86

range

n

Age yrs*

23.5 26.2 29.2 25 28.1 30.4

26.8

19.56 17.62 17.54 16.86 16.41 17.62 20.18 26.14

mean

limit

1.3 2 1.8 1.9 1.4 1.7

26.3 31.1 32.9 28.4 30.8 32.4

2.4 32.9

2.4 1.4 2 1.5 1.1 1.6 1.6 3

SD

Min Latency ms

0.2

1.03

mean

1.2

0.73

SD

2.6

2.5

limit

Side diff-Min Lat ms

24.3 27.2 30.3 25.6 28.8 31.3

27.7

mean

0.6 2 1.9 1.9 1.6 1.9

2.5

SD

limit

Mean Lat ms

26.15 33.32 36.75 41.69 51.86 55.72 59.68 63.83

17.31 17.44 17.91 19.44 23.23

mean

5 3.9 4.2 4.1 6.5 3.8 4.5 4.2

1.8 1.3 1.1 1.5 2.6

SD

limit

Max Velocity m/s

>50 in 97%**

87.5 60 63 60 30 30 85.7 66.7 100 80 90

66

mean

30

limit

Persistence % mean SD

limit

Amplitude μV

4.48

mean SD

8.92

Limit

Chronodispersion ms

12 11 12 14 14 17 152 51 101

80

n

168 25 20 20 20 20 21 22 19

35

22–58

35

24 h–30 24–72 h 0–3 m 4–6 m 7–12 m 1–3 4–6 7–14 18–30

20–64

22–58

18.26

mean SD

<1m 1–6 m 6–12 m 12–24 m 24–48 m 48–72 m 15–83 43.1

< 1–72 m

range

Age yrs*

150–162 163–175 176–190

Height cm

21.7 20.3

16.51 15.39 15.41 14.37 14.21 15.81 17.34 22.84

24.8 22.8 21.2 23.3 24.9

18.17 15.91 15.67 15.64 16.36 18 22.6

mean

limit

2.8 1.6

1.7 1.5 1.6 1.2 0.8 1.2 1.5 1.3

2 1.9 1.9 1.6 1.8

**

27

2.2 1.2 0.9 1.1 1.5 1.3 1.7 28.5

SD

Min Latency ms

Except where indicated, h = hours, d = days, m = months In 97% of the cases 1 Recording with concentric needle electrode 2 Medians instead of means given 3 Mean (SD) 4 Calculated on 61 left median nerves Median F/M ratio 1.8 (range 0.5–5.5) 5 Latencies given have had the distal motor latency substracted 6 Used in multiple regression equations 7 Number of nerves in each of the age groups

*

AXILLA Kimura 74 (10) 33 Kimura 83 (16)

Cai & Zhang 97 (>10)

Puksa 03 (20) 5 Males Females ELBOW Kimura 74 (10) 33 Kimura 83 (16) 65 Russo 85 (10) 30

Garcia et al., 00 (10)

Stimuli (number)

Continued

Table 9.1

0.73

0.92

0.76

0.25

mean

0.54

0.7

0.56

0.79

SD

2.1

2.3

1.9

limit

Side diff-Min Lat ms

23.9

mean

limit

1.7 30.2

SD

Mean Lat ms

64.3

27.37 34.8 37.9 44.7 52.91 56.47 51.5 64.58

62.2 67.8

mean

6.4

4.6 4.1 6 6.1 8.7 5.4 6.8 4.7

5.2 5.8

SD

56

limit

Max Velocity m/s

73.5

mean

15

limit

Persistence % mean SD

limit

Amplitude μV

2.2

0.9

mean SD

5.5

Limit

Chronodispersion ms

30–66

12–81

42 17.4 44.2 15.6 20–81 51

21

64

32 32 33

PeiogluHarmoussi 85 (200) PeiogluHarmoussi 85 (20) Males Females Guiloff 91 (60)

26.1 28.1 29.2

19.1

17.6

1.7 1.6 1.4

1

0.9

0.8

0.7 0.4

2.2 32

2.34 1.8 31.4

2

3 2.5

26–32

1

0.83 2.7

0.38 0.31

limit

Side diff-Latency ms

limit mean SD

175.5 (7.8) 27.3 1.8 162.4 (6.1) 24.3 1.3 27.9 2.5 32

150–162 163–175 176–190

7–12

15

20–64

5–7

15

30

3–5

25

15.9

14.6 14.7

26.02 26.2

0–1 1–3

139–183

19 12

9

27

30.5 29.9

mean SD

27.6

63

Height cm

Min Latency ms

65 54

38 21*

11–74 41.5

60

mean SD

28–58 35

range

33 17

n

Russo 85 (10)

WRIST Mayer 67 (min 10) Kimura 74 (10) Kimura et al. 74 (10) Eisen et al. 77(10) Weber 78 (20) Argyropoulos 79 (100) Kimura 83 (16) Kwast 84 (20-100)**7

Stimuli (number)

Age yrs^

65.3

61.7

56.7 56.6

58.9

55

45–60

57.2 4.7

4.8

3.5

2.9 3.1

2.6

limit

Max Velocity m/s

mean SD mean SD

Mean Lat ms

47.8 83.6

79.1

mean SD

14.7

10

2.4

limit

Persistence %

0.183 0.1 0.64 0.24

0.21

1.1

SD

%

range

F/M ratio amp

0.4–2.2

SD1.1

5.7

1.1

mean SD

3.7–10.8 9.8

3.3

1.5

1.2

1

0.9 1.3

range

5.32 2.7–11.3 1.8 1 0.7–5.9

5.5

3.6

3.6

3.1

2.5 3.6

mean

Chrondispersion Repeaters ms %

3.4

range

area

1.71 0.8–4.0

%

0.56 0.24 1.473 0.6–2.5 1.1

100–770

50–300 300

range max

Amplitude μV mean SD

F-waves. Normal values. Ulnar nerve (surface electrodes over abductor digiti minimi except where indicated)

Table 9.2

n

65 8 7 11 12 11 16 23

Nobrega et al. 01 (64) Puksa 03 (20)5 196 Males 73 Females 123 ELBOW Mayer 67 10 Kimura 74 (10) 33 Kimura et al. 74 (10) below elbow 17 above elbow 17 Eisen et al. 77 60

Garcia et al. 00 (10)

Zappia et al. 51 93** (10) Cai & Zhang 168 97 (>10) 25 20 20 20 20 21 22 19 Buschbacher 193 99 (10)

Stimuli (number)

Table 9.2 Continued

mean SD

1.8 32.9 1.9 28.4 1.4 30.8 1.7 32.4

29.2 25 28.1

30.4

1.8

2 2 1.6

26

26 23.5 23

28–58 35

11–74 41.5

20–27

1.9 28.9

23.7

14–94 44.1 19.16

1.6 1.6 1.37 0.76 1.41 1.48 2.2

1.3 26.3 2 31.1

2.74 1.39 1.24 1.45 1.88 1.74 1.92 2.14 2.5 32.9

2.4 32.8

0.1

0.2

0.81

1.1

3.1

0.94 0.69 2.3

limit

Side diff-Latency ms

limit mean SD

23.5 26.2

19.67 17.65 16.99 17.02 16.63 18.51 20.66 27.03 26.5

25

mean SD

9.4

< 160 > 180 < 160 > 160 > 180 > 180 < 160 > 160 < 180 > 180

Height cm

Min Latency ms

18.63 15.71 15.45 15.67 16 18.25 25.5

< 1–72 m < 1m 1–6 m 6–12 m 12–24 m 24–48 m 48–72 m 17–59 32.9

50–79

19–49

24 h–30 24–72 h 0–3 m 4–6 m 7–12 m 1–3 4–6 7–14 18–30 19–79

19–70 36.4 15.7

range

Age yrs^

24.8

27

31.3

30.3 25.6 28.8

24.3 27.2

27.7

27

1.9

2.2

1.9

1.9 1.9 1.6

0.6 2

2.5

58.2 61.1 62.3

59.4

27.12 33.26 38.19 42.43 51.89 55.36 61.31 62.97

2.2 57.2

3 5.4 5

4.7

4.54 3.52 3.94 4.42 7.07 5.17 4.27 4.77

5.1

limit

Max Velocity m/s

mean SD mean SD

Mean Lat ms

87.5 16.6

67.4 27.4

mean SD

15

0.57

limit

Persistence %

0.31

mean SD

range

Amplitude μV max SD

%

range

F/M ratio amp % 4.2

range

area

2.3

4.1

3.2

mean

SD0.8

SD1.3

range

mean SD

Chrondispersion Repeaters ms %

1.9 1.8 1.6 24

1.3 1.8 1.28 0.88 1.35 1.48 1.46 1.9

1

Recording with concentric needle electrodes Population median of individual median amplitudes given 2 Population median of individual maximal F/M ratios given 3 Population mean of individual median amplitudes or F/M ratios 4 Population mean of individual maximal amplitudes 5 Latencies given have had the distal motor latency substracted 6 Used in multiple regression equations 7 Number of nerves for each of the age groups

**

0.85 0.61 2.1

0.88 0.64 2.2

0.68 0.48 1.6

limit

Side diff-Latency ms

limit mean SD

1.7 27 1.5 1.3 1.2

^ Age in years except where indicated, h = hours, d = days, m = months * sides

21.9 21.5 20.3

23.1 22.6 24 24.8

mean SD

28–58

150–162 163–175 176–190

Height cm

Min Latency ms

16.64 14.93 14.91 14.41 14.69 16.53 18.14 23.42

mean SD

Age yrs^

24–72 h 0–3 m 4–6 m 7–12 m 1–3 4–6 7–14 18–30

24 h–30

20–64

Kimura 83 (16) above elbow 65 Russo 85 (10) 30

Cai & Zhang 168 97 (>10) 25 20 20 20 20 21 22 19 AXILLA Kimura 74 (10) 33 Kimura 74 (10) 17 Kimura 83 (16) 65

range

n

Stimuli (number)

Table 9.2 Continued

63.1 64.4

26.14 33.34 37.9 42.78 50.36 55.29 59.81 61.18

65.7

5.9 6.5

3.87 4.93 4.25 4.7 7.43 5.92 5.75 6.29

5.3

55

limit

Max Velocity m/s

mean SD mean SD

Mean Lat ms mean SD

limit

Persistence % mean SD

range

Amplitude μV max SD

%

range

F/M ratio amp %

range

area mean

range

mean SD

Chrondispersion Repeaters ms %

Stimuli

22–39

18 10

Doko-Guina 97 (6)

Cai & Zhang 97 (>10)

7 d–14 1–6 m 6–12 m 1–2 2–4 4–6 6–14 24 h–30

24–72 h 0–3 m 4–6 m 7–12 m 1–3 4–6 7–14 18–30 0–72 m

0–3 m 4–6 m

20 23 25 24 22 20 21 168

25 20 20 20 20 21 22 19 86

8 10

27.56 26.14 25.18 25.54 26.73 30.57 38.16 49.63

42.7

7 d–14

15.7

155

36.4

19–70

51

3.82 2.84 4.37 2.04 2.87 3.82 4.43 7.74

5.2

4.6

50.7

28–57

17

3.6

4.7

SD

65

44.6

51.3

mean

3.2 3.8 4.2 4

Height cm

147–160 46.3 163–175 49.3 178–193 52.8 48.4

8.7

SD

3.7

60

28

41.5

26

mean

56

limit

Min Latency ms

43.2

Kimura 83 (16) Petajan 85 (20–60+) Zappia et al., 93***(10) Parano et al., 93 (?)

11–74

60

7 21*

21–47

range

66

n

Left Argyropoulos et al., 78 Tonzola 81

ANKLE Kimura et al., 75 (10) Eisen at al., 77 (10) Fisher 78 (10) Right

(number)

Age yrs^

1.19

1.42

mean

1.17

1.04

SD

3.5

3.5

limit

Side diff-Latency ms

45.4

45.4

46.7

mean

5.1

5.5

4.4

SD

limit

40– 52.1 40.3 –52.7

Mean Lat ms

3.6

3.3

3.8

SD

22.63 28.18 33.83 38.76 41.37 46.91 48.48 49.74

22.07 23.11 25.86 25.98 29.52 29.98 34.27

4.56 5.42 6.19 3.18 3.43 5.24 3.08 5.52

1.46 1.89 1.35 1.95 2.15 2.68 4.29

47.7**** 6.6

49.8

59

53.3

mean

43

limit

Max Velocity m/s

12.5 40

46

37.2

73.1

35

38

Limit

25.3

4

0–753

0–803

SD

Persistence % mean

F-waves. Normal values. Peroneal nerve (recording over extensor digitorum brevis except where indicated)

Table 9.3

0.64

mean

0.6

SD

max

Amplitude μV SD

5.3

mean

7.1

SD

1.9

limit

Chrondispersion ms

0.9

mean SD

Repeaters %

mean

4.5

54.6

46

56.5

53.2 46.8 51.2

45.7 48.8

52

30–58 15–48

39.2 28.6

2.9 3.1 2.6

172–220** 37.6 172–220** 35.5 38.5

3.1

5

3

4.3

4.7

4.1 4.5 4.6

2.9 3.9

5.6

SD

Mean Lat ms mean

29.8

5.1

limit

6

1.58

2.4

SD

30.1

0.15

0.7

mean

22–39

38.2

4

64.2

60.5 58.1 60

47.2 54.6

60.5

limit

18 6

41.5

42.7

4.82 1.78 1.1 1.66 2.21 2.58 4.1

4.1 4.8 4.6

51.5 45.4 49.6

25.2 21.4 20.33 22.82 24.64 29.45 44.5

2.5 3.7

5.5

SD

43.6 47.1

50.2

mean

11–74

< 160 > 180 < 160 > 160 > 170 > 170 < 160 > 160 > 170 > 170

cm

Height

Side diff-Latency ms

60

26

202

SD

Min Latency ms

21–47

< 1–72 m <1m 1–6 m 6–12 m 12–24 m 24–48 m 48–72 m 15–94 47

40–79

19–39

7–9 m 10–12 m 13–18 m 19–24 m 25–36 m 37–48 m 49–60 m 61–72 m 19–79

range

Age yrs^

66

72 10 10 9 13 16 14 121 42 79

8 10 10 10 7 6 7 10 180

n

Panayiotopoulos 20* et al., 77 40* Panayiotopoulos 79 (100) 5

Puksa 03 (20)1 Males Females KNEE Kimura at al., 75 (100) Eisen et al., 77 (10) Fisher 78 (10) Tibialis ant Right Left

Garcia et al., 00 (10)

Buschbacher 99(10)

(number)

Stimuli

Continued

Table 9.3

mean

24.6– 33 22.9–7 34.7

limit

SD

33

32

56.3

5

165 0–100

143

mean

3

0

Limit

0–1003

4.9

2.5

SD

Persistence % mean

37.5 20 50 70 42.9 33.3 85.7 70 > 50 in 55%

limit

Max Velocity m/s

60

98

SD

271

285

max

Amplitude μV SD

6.4

3.3

mean

0.8

1.79

SD

limit

Chrondispersion ms

mean SD

Repeaters %

24 h–30 24–72 h 0–3 m 4–6 m 7–12 m 1–3 4–6 7–14 18–30

168 25 20 20 20 20 21 22 19

mean

3.74 2.76 2.68 3.36 2.46 3.44 4.75 7.63

46 35.5

limit

^ Age in years except where indicated, h = hours, d = days, m = months a Surface electrode in tibialis anterior * sides ** Distance from knee to L1 × 2 *** Recording with concentric needle electrodes **** Calculated using mean and not minimum F-wave latency 1 Latency values have the distal motor latency substracted 2 Considered in multiple regression equations 3 Range given 4 The author gives the mean % of absent responses 26.9% SD 19.6%

24.38 23.46 22.15 21.56 24.3 25.22 31.38 41.23

3.2

SD

Min Latency ms

39.9 157–195 30.5

21–43

65 10

Kimura 83 (16) Burke 89 (50 minimum)a Cai & Zhang 97 (>10)

SD

cm

mean

Height

range

n

(number)

Age yrs^

Stimuli

Continued

Table 9.3

1.24

1.25

mean

1.03

0.92

SD

3.5

3.1

limit

Side diff-Latency ms

33

mean

SD

Mean Lat ms

39.3

limit

22.23 28.46 34.84 39.17 41.67 49.62 49.74 50.85

53.7 59.2

mean

3.81 4.34 5.95 4.28 4.05 7.31 5.34 4.85

4.8

SD 44 55.7

limit

Max Velocity m/s mean

SD

Limit

Persistence % mean

SD

max

Amplitude μV SD

4.1

mean

SD

11

limit

Chrondispersion ms

mean SD

Repeaters %

15.7

41.1

15–67

50.6 55.4 47.7

163–175

178–193

4.7

23.48 29.29 35.03 40.52 43.64 48.43 50.61 50.68

44.4

2.85 2.34 2.63 3.71 2.66 4.01 4.45 2.39

5.4

4.3 44 77.4

34 38 80 80 80

1.01 3.3

49.9

52.6

4–6 m 7–9 m 10–12 m 13–18 m 19–24 m

1.26

1.04 3.5

100 93

limit mean

100

64

1.4

4.7 4.9

2.9

SD

mean

SD

25.4 6002

660

max

604 650 890

Amplitude μV

401 123 62–1001 374 130 236 105

SD

Persistence %

0–3 m

3.27 2.41 1.85 1.83 2.2 3.1 3.72 3.09

6

4.2 5 58

3.7

3.6

48.2 48.9

51.3

limit mean

Max Velocity m/s

76

26.92 28.59 23.93 23.78 25.44 31.07 36.32 48.27

46

47.3

147–160

4.8

3.3

4.3

Mean Lat ms

limit mean SD limit mean SD

Side diffLatency ms

24–72 h 0–3 m 4–6 m 7–12 m 1–3 4–6 7–14 18–30 0–72 m

24 h–30

36.4

28

22–39

18 15 16 42 49.1

47.4

19–70

Min Latency ms mean SD

11–74

Height cm

60

41.5

mean SD

52.3

range

66

n

Kimura 83 65 (16) Zappia et al., 51 93 (10) Cai & Zhang 168 97 (>10) 25 20 20 20 20 21 22 19 Doko-Guina 86 97 (6) Flex hall. 8 brevis 10 8 10 10 10

ANKLE Kimura et al., 75 (10) Eisen at al., 77 (10) Fisher 78 (10) Right Left Eisen et al., 79 (32) Tonzola et al., 81 Flex. hall. brevis

Stimulus (number)

Age yrs^

F-waves. Normal values. Tibial nerve (recording over abductor hallucis, except where indicated)

Table 9.4

299

SD

1.2

%

F/M

0.4

SD

7.8

8.8

SD

Repeaters %

mean SD limit mean

Chrondispersion ms

Panayiotopoulos et al., 80 29* Gastroc Soleus 29* Tonzola et al., 81 Soleus Kimura 65 83 (16)

20–72 20–72

42.6 42.6

28

22–39

15.7 15.7

163–175

147–160 32.6 39.6

29.1 32.8 29.48

Mean Lat ms

61.4 45.4 52.7 59.2 59.8 57.2 59.7 59 67

2.1 4.4 48

1.25

0.6

5.7

0.92 3.1

2.3

53 44.7 49.2 54 55.8 48.6 52.4 56.5 60

53.7

60.3 57.6

54.4

83

86

57 100 100 90

limit mean

4.8 44

517

max

75–1001350 120

SD

700

mean

Amplitude μV

75–1001370 169

SD

Persistence %

4.8 50.5–71.4 4.8 50.8–69.9

3.6

SD

Max Velocity m/s limit mean

2.7 26.2– 35.3 2.3 27– 35.3

5.6 2.4 3.1 4 5.3 4.4 3.4 4.2 5.2

limit mean SD limit mean SD

2.3 33.8 2.7 38.8 2.3

2.95

3.4

43.5 39.7

1.62 1.35 2.05 1.63 2.53 2.52

5.3 2.2 3 4 4.4 4.6 3.5 3.6 5.3

23.92 21.4 22 24.21 25.6 30.12

50.8 43.2 47.2 52 53.1 46.8 50.5 53.9 57.9

mean SD

30.5

16

41.5

< 160, > 180 < 160 > 160, < 170 >170, <180 >180 <160 >160, <170 >170, <180 >180

Height cm

Side diffLatency ms

Left

mean SD

11–74

< 1–72 m <1m 1–6 m 6–12 m 12–24 m 24–48 m 48–72 m

40–79

25–36 m 37–48 m 49–60 m 61–72 m 19–79 19–39

range

Min Latency ms

30.2

n

Age yrs^

7 6 7 10 Buschbacher 159 99 (10) 7 22 15 15 23 32 33 12 Garcia et al., 79 00 (10) 12 10 11 14 15 17 Puksa 03 (03) KNEE Kimura et al., 66 75 (10) Eisen et al., 60 77 (10) Fisher 78 (10) 18 Soleus Right 16

Stimulus (number)

Continued

Table 9.4

SD

%

F/M SD

SD

Repeaters %

mean SD limit mean

Chrondispersion ms

n

21–43 24 h–30 24–72 h 0–3 m 4–6 m 7–12 m 1–3 4–6 7–14 18–30 0–72 m 0–3 m 4–6 m 7–9 m 10–12 m 13–18 m 19–24 m 25–36 m 37–48 m 49–60 m 61–72 m

range

mean SD

Age yrs^

157–195

Height cm

23.51 22.52 20.67 20.92 23.65 25.97 32.78 39.93

29.4 2.45 2.1 2.3 1.59 1.71 2.32 3.89 2.73

mean SD

Min Latency ms

1

Age in years except where indicated, h = hours, d = days, m = months range given 2 Recording with concentric needle electrode

^

Burke 89 (min 50) Soleus 10 Cai & Zhang 168 97 (>10) 25 20 20 20 20 21 22 19 Doko-Guina 86 97 (6) 8 Soleus 10 8 10 10 10 7 6 7 10

Stimulus (number)

Continued

Table 9.4

Mean Lat ms

34 1.18

0.98 3.3

30.8

limit mean SD limit mean SD

Side diffLatency ms

36.1 23.66 29.96 36.89 41.11 43.93 50.32 51.26 51.88

limit mean

3.33 2.61 3.92 4.08 3.34 5.99 4.32 3.58

SD

Max Velocity m/s

45 62.5 30 37.5 60 30 30 28.6 66.6 57.1 50

limit mean

SD

Persistence % mean

SD

Amplitude μV max

SD

%

F/M SD

3.6

5.7

SD

Repeaters %

mean SD limit mean

Chrondispersion ms

30.7

10.7

12

30.7

11.1

30 18–55 36.6

154 –188

167

29.6 6.4

21.9 1.4

22.7 6.7

10.7

12

30.7

20.5 1.9

49

20.4 4.8

17.8 –25.3

24.4

0.4 0.3 0.1 –3.1

0.55 0.5 0–1.7

30.3 8.3

24.3 6.2

20.6 2.2 20.8 4.9

10.7

78 12

11.2 1.11 8.9–13.4

13.9 2.5

Limit

Mean Lat6 ms

limit mean SD limit mean SD

Sides diff-Lat ms

10.6 1.23 8.5–13 0.87 0.7

12.3 1.6

mean mean SD

157–203 176.6

range

Min Latency ms

20.1 1.8

5

30.7

10.7

SD

Height cm

8

42 21–66 34.4

12

n range mean

Age yrs

F Cond. time ms

12.7 0.9 < 14.1

59.5 19.1

limit

3

2

10 12.7

mean SD

24.1

16.6

75

0–100 184

78

35.2

30.2

108 60–520

750

max SD

12–242

2.2

0.8

2.8

3.7

4.1

4.9

0.6

3.2 0.59

2.8

0.9

0.57

4.1

SD

0–3.33

range

Chronodispersion ms

limit mean SD mean

F/M Ratio %

23–273 127 66 41–423 4.6

45 100–210 280

50

31

90

limit

Amplitude6 μV

500 66–271

2350 4070

106

1070

0–100 145

26 0–100 154 17.5 19.5 300

29

0–100

200

limit mean SD

Persistence %

3.86 0.52 2.75–4.8 28.5

mean SD mean SD

Velocity7

Recording with concentric needle Stimulating cathode over zygomatic branch of the nerve, anode distally over presumed course of the nerve (Wedekind) or over mastoid (Zappia) Ground midway between recording electrode and lateral epicondyle. Stimulating electrode midway between ground and the lateral epicondyle, half way between radial and ulnar bones (Papathanasiou) or elbow (Zappia). 4 Stimulating electrode over the inguinal ligament 5 Stimulation at Erb’s point 6 Whether the mean is calculated including or excluding absent responses not stated 7 Calculated using the mean of the individual mean F-wave latencies 8 For amplitude 12 nerves were recorded on each side

1

Facial Nerve2 Zappia et al., 93 (10)1 Triangularis Wedekind et al., 01 (16–60) Nasalis Radial Nerve3 Fisher 78 (10) Ext.digit. comm Right Left Zappia et al., 93 (10)1 Ext.digit. comm Papathanasiou et al., 01 (16) Ext. indicis Femoral nerve4 Zappia et al., 93 (10)1 Vastus medialis WochnikDyjas et al., 96 (10) Vastus lat. Axillary nerve5 Zappia et al., 93 (10)1 (Deltoid)

Muscle

Nerve & Author (number of stimuli)

F-waves. Normal values. Facial, axillary, radial and femoral nerves

Table 9.5

LATE RESPONSES (F- AND A-WAVES): METHODOLOGY AND TECHNIQUES

(Panayiotopoulos, 1978) and that replacing the measurement of the distance of a nerve segment by the length of a limb or by height did not change significantly the error of the distance measurement (Panayiotopoulos, 1978; Kimura, 1978a). The F/M latency ratio (see below) was proposed to bypass the arguments about distance measurement (Eisen et al., 1977b). At present, it seems that whatever method is chosen, the measurements will be clinically useful provided the technique is standardized and applied in the same way to the control subjects to obtain normal values and to the patients. 9.2.4.3.3. F/M latency ratio. This ratio was originally described as useful in the evaluation of proximal entrapment syndromes (Eisen et al., 1977b). It has the advantage that no distance measurements are required. In essence, the F-wave shortest latency to the stimulation point minus the turn around time is divided by the latency of the M-response and this is then divided by 2 to express the ratio for one length of the pathway only (since the F-responses have an antidromic and an orthodromic arm). Thus, the following formula can be used for this ratio (Kimura, 1978b). F/M (F shortest latency – M latency) – 1(ms) latency ratio = M latency × 2(ms) The method provides a comparison of conduction proximal to the stimulation site with that distal to the stimulation site, including conduction in the neuromuscular junction. 9.2.4.3.4. F-wave mean latency. This measurement has been advocated by some (Fisher, 1982, 1992). It needs few stimuli and is reproducible. The problem is that its biological meaning is obscure. Most authors agree that F-wave latencies and conduction velocities are not normally distributed (see Section on Population of motor neurons generating F-waves) so that the mean latency is not a representative parameter of the F-wave population in an individual subject nor in a group of subjects. If anything, the median latency would be more representative (Petajan, 1985; Guiloff and Modarres-Sadeghi, 1991). The mean latency cannot be compared to conventional latencies or motor conduction parameters in peripheral nerves that measure either latency for the fastest motor fibers or maximum motor conduction velocity. The mean latency cannot be compared either with other techniques that

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look at the spectrum of motor conduction velocities, such as collision (HOPF, 1962; Ingram et al., 1987), deconvolution of cMAP (Dorfman, 1984) or reconstruction of cMAP (Lee et al., 1975); these can be compared, however, with the distribution of F-wave latencies or F-wave conduction velocities (chronodispersion and tacheodispersion). 9.2.4.3.5. F-wave maximum latency. The difference between the fastest and the slowest F-wave is used to calculate chronodispersion and tacheodispersion (see below). The measurement is being increasingly used as chronodispersion is proving to be increasingly helpful in detecting abnormalities in motor conduction that may not necessarily involve the fastest conducting fibers. 9.2.4.3.6. Chronodispersion. This parameter was described as the scatter or dispersion of the relative latencies of a statistically significant number of consecutively recorded F-waves (Panayiotopoulos, 1979). One hundred F-waves were used in the original paper and probably not less than 60 stimuli should be used in clinical practice (Guiloff and Modarres-Sadeghi, 1991) (see sample size below). Panayiotopoulos plotted histograms of F-chronodispersion as a latency difference between each F-wave and the fastest F-wave in each individual. Marked differences were shown between five control subjects and five patients with chronic renal failure but without clinical or electrophysiological evidence of peripheral neuropathy. Patients showed significantly increased chronodispersion with a mean of 9.9 ms ± 1.3 difference between the fastest and the slowest F-wave compared to 6.4 ms ± 0.8 in the controls. In control subjects 26% of the F-waves had a latencies within 1 ms of the fastest F-wave whilst in patients only 10% did so. It was predicted that this method might be more sensitive than other F-wave parameters in the detection of motor peripheral nerve abnormalities (Panayiotopoulos, 1979), and there is increasing evidence that this is indeed the case (Weber, 1998). There is a need to increase normative data for chronodispersion. There is insufficient data in patient groups, perhaps because of the need to give large number of supra-maximal stimuli to obtain a representative value for chronodispersion. It is important to realize that to measure the true extent of chronodispersion in pathological conditions, it may be necessary to increase the sweep speed from 5 ms/division in the upper limb and 10 ms/division in the lower limb, to 10 ms/division and 200 ms/division,

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respectively, or more. Frequent checks of sweeps with no stimulation are required during the recording to ascertain that “late responses” are not the result of poor relaxation with voluntary activation of units unrelated to the stimuli, which can be mistakenly taken as F-waves. 9.2.4.3.7. Tacheodispersion. This is a similar parameter to chronodispersion but normalises the data for distance (patient’s height and limb span). Thus, the criticisms and counter-criticisms levelled at estimating conduction velocity with F-waves also apply. The distribution of F-wave conduction velocities was shown to sample adequately the spectrum of conduction velocities of the motor axons of a peripheral nerve (Guiloff and Modarres-Sadeghi, 1991). The parameter is the distribution of conduction velocities of a statistically valid sample of consecutive F-waves as a description of the spectrum of conduction velocities of axons within a peripheral nerve in normal and pathological conditions (Chroni and Panayiotopoulos, 1993a, 1993b). The distance between the spinous process of C7, and of L1, and the point of stimulation, for the upper and lower limb, respectively, were used to calculate the distance. The velocity for the fastest F-wave is calculated by Kimura’s formula: Distance FCVmax = [(FLAT min −M lat −1) / 2 ]. Each F-wave velocity (FCVx) is calculated then by multiplying the FCVmax by the ratio of latencies of the shortest and of each x F wave (minus 1 ms of turnaround time for each latency) as follows: FCVx = FCVmax × (FLAT min −1 / FLATx −1) The distribution of F-wave conduction velocities of the sample can then be plotted. To normalise for different FCVmax in different subjects the individual F-wave CVs can be plotted as a difference from the fastest FCV in each subject (Guiloff and Modarres-Sadeghi, 1991). 9.2.4.3.8. F-wave duration. There is at present little or no hard data on this parameter to warrant its use in a clinical setting. Theoretically, its main determinant in normals is the temporal dispersion of the constituent mean 2–3 motor units of each F-wave. In a pathological situation, not only the abnormalities of motorneurons in the cord but also abnormalities in conduction in peripheral nerves will contribute to this parameter.

R.J. GUILOFF

9.2.4.3.9. F-wave amplitude. Mean or median amplitudes can be used. The mean peak to peak amplitude of the F-waves is measured in absolute values or as a proportion of the M-wave amplitude (F/M ratio) (Eisen and Odusote, 1979). Absent F-responses should be considered as 0 in order to calculate mean or median amplitude and the proportion of the motor neuron pool that generates F-waves (F/M ratio) (Kimura et al., 1984; Guiloff and Modarres-Sadeghi, 1991; Fisher, 1992; Fisher et al., 1994). An increase in mean or median amplitude cannot always be assumed to reflect an increase in the excitability of motor neurons. It may result from at least three mechanisms. First, an increase in the number of motor neurons backfiring with each stimulus; second, an increase in the frequency with which motor neurons backfire leading to an increase in persistence and third, pathology such as an increase in the size of the motor units producing F-waves. An increase in the F/M ratio to more than 5% is said to be common in upper motor neuron disorders (Eisen et al., 1977b; Eisen and Fisher, 1999). A reasonable estimate of mean or median amplitude can be made from a sample obtained with 20 stimuli or less (see below, sample size). 9.2.4.3.10. Persistence or frequency of F-waves. It is defined as the proportion of stimuli followed by detectable F-waves (Eisen et al., 1977a). Since F-waveforms are generally of low amplitude this parameter depends critically on the amplification used and on how noisy or not noisy the background is. Most authors accept minimum amplitudes of 20–40 μV and gains of 200 μV/division. However, increasing the gain to 100 or 50 μV/division can dramatically increase persistence if the baseline is suitably flat (Fig. 9.5). Accordingly, normative values need to specify clearly minimum amplitude and amplification used in the collection of data, and recordings in patients need to be done in exactly the same way to draw valid conclusions. Persistence can be established with relatively small number of stimuli (10–20) (see below, sample size). Changes in persistence may reflect a variety of mechanisms, such as changes in excitability, in motor neuron numbers, function or territory (Guiloff and ModarresSadeghi, 1991), or changes in peripheral motor conduction, in particular demyelination or focal conduction block. The values for persistence vary widely in normal subjects (see normative values in tables 9.1–9.5) and, in the peroneal nerve for example, can be as low as 30% in a population, or 0% in an individual.

LATE RESPONSES (F- AND A-WAVES): METHODOLOGY AND TECHNIQUES

9.2.4.3.11. F-wave repeaters. In clinical practice, with surface recordings and supra-maximal stimulation, repeater F-waves are waveforms with a shape that appears more than once when a number of stimuli are given. They can have different shapes, each of which can repeat more than once. The proportion of repeaters in an F-wave study can be expressed as percentage of repeater F-wave shapes or percentage of repeater F-waves (Fig. 9.5) (Peioglou-Harmoussi et al., 1985a; Macleod, 1987; Guiloff and ModarresSadeghi, 1991). To collect a representative sample of repeater shapes, 100 stimuli are required (see below, sample size). It is thus unlikely to be used as a parameter in routine nerve conduction studies. Further, its clinical utility is at present rather limited. 9.2.4.4. Sample size required In normal subjects each surface recorded F-wave represents the recurrent responses of only a small proportion (less than 5%) of the motor neuron pool supplying a muscle, after a supra-maximal antidromic volley stimulates all motor axons to that muscle. The desired objective for using the technique in clinical practice is to sample adequately the whole motor neuron pool, and their motor axons, supplying a muscle. Each of the parameters used are intended to measure features of such motor neuron pool and axons, be it excitability, maximal conduction velocity, or spectrum of conduction velocities. Therefore, it is important to establish how many stimuli need to be applied to obtain measurements of the F-wave parameters of interest that are not only accurate and reliable but also representative of the whole motor neuron pool of such muscle. This problem has been addressed in normal subjects by comparing the value of such parameters when a large number of stimuli are given with the values obtained when a smaller number of stimuli are applied. The number of stimuli usually taken as the “gold standard” is around 100. Panayiotopulos plotted five successive groups of 20 F-waves and showed that only a very small number of shortest F-waves could be elicited with 20 stimuli; he recommended that large number of stimuli may be needed for correct measurements (Panayiotopoulos et al., 1977). Marra compared minimal and mean latency of 3–5 and 20 F-responses to the average of 150–200 F-responses. He used recordings in himself and in two different control groups; twenty five controls were used to record the minimal latency of 3–5 F-responses, and twenty two subjects were used to

219

record responses to 150–200 stimuli. He found that a reliable estimate of the minimum F-wave latency would require at least 20 trials to approach the value obtained by 150–200 trials (Marra, 1987). The minimum latency F-response in the abductor digiti minimi was studied in100 normal subjects by delivering 100 stimuli to each of them. At least 60 stimuli were needed to be 95% certain that the minimum latency F-response had been obtained; the same minimum latency was obtained with 100 or 80 stimuli in all subjects (Barron et al., 1987). The minimum sample size required to obtain estimates for the various F-wave parameters close to those obtained with a “gold standard” sample size was reported in detail in 1991 (Guiloff and ModarresSadeghi, 1991). The abductor digiti minimi was used in 11 normal subjects who were all given 100 supramaximal antidromic stimuli to the ulnar nerve at the wrist. It was shown first that, if F-wave latency, amplitude, area and duration obtained with 100 stimuli are each considered a time series and an auto-correlation function of lag 1 is used, there is no significant variation over time for any of them, that is, there is no last period time dependency. It was then shown that these parameters, and persistence, were not significantly different for the F-waves evoked by five successive groups of 20 stimuli each. Finally, a number of parameters in randomly obtained samples of 5, 10, 20, 40, 60 stimuli were compared with those obtained with 100 stimuli. With 20 stimuli all 11 subjects showed a less than 1.5 ms difference in minimum latency with that obtained with 100 stimuli. This difference only improved to less than 1 ms with 60 stimuli and increased to less than 2 ms with 10 stimuli. In agreement with a previous study (Barron et al., 1987), with 60 stimuli 10/11 subjects had a minimum latency within 0.5 ms of that obtained with 100 stimuli. For median latency and amplitude, on average, all samples sizes gave values less than 0.3 ms and less than 0.01 mV difference respectively from those obtained with 100 stimuli. For maximum amplitude though, samples with 20 stimuli or less gave, on average, smaller amplitudes by 0.18 mV or more. Chronodispersion decreased, on an average, by 0.4 ms with 60 stimuli, by 1.2 ms with 40 stimuli, by 2.5 ms with 20 stimuli and by 3.1 ms with 10 stimuli. For persistence samples down to 20 stimuli gave 0.8% or less difference with 100 stimuli, but with 10 stimuli the difference increased to 3.3%. The mean % of repeater shapes found decreased dramatically with sample size, as expected, from 11.2% with 100 stimuli to 1.2% with

220

10 stimuli. The closest was 60 stimuli with 9.8%. The conclusion from this study was that for minimum and median latency, median amplitude and persistence samples obtained with 20 stimuli are clinically adequate. For maximum amplitude no less than 40 stimuli should be used. For chronodispersion and repeater shapes, samples of no less than 60 stimuli should be used (Fig. 9.5). With 10 stimuli, still a widely used sample size in clinical practice, only mean median latency and mean median amplitude were close enough to the “gold standard” figures obtained with 100 stimuli. These findings have been largely confirmed by others (Taniguchi et al., 1993; Fisher et al., 1994; Chroni et al., 1994, 1996; Raudino, 1997; Nobrega et al., 2001) though the precise departure from the “gold standard” varies in different studies. In any event, it is clear that for most parameters, the “normal values” depend critically on the number of stimuli used to obtain the sample of F-waves. It has been commented that for certain patient populations 20 stimuli may not be enough to establish a representative value for minimum latency (Chroni et al., 1994, 1996). There is a need to establish, with studies similar to those quoted here, the optimum sample size of F-waves to detect abnormal findings in patient populations with different pathologies. 9.2.4.5. Techniques in children A quiet and darkened room may be helpful and the presence of the parents may be valuable. Sedation may be required and does not alter the electrophysiological results (Wagner and Buchthal, 1972; Miller and Kuntz, 1986). Recordings of motor conduction or of F-waves in children can be made with an 80u stainless steel wire inserted in the muscle (Wagner and Buchthal, 1972), with concentric needle electrodes (Kwast et al., 1984) and, more frequently, with surface electrodes (DokoGuina and Jusic, 1997; Cai and Zhang, 1997; Garcia et al., 2000). Stimulation has been given with needle (Wagner and Buchthal, 1972) and, more frequently, with surface (Doko-Guina and Jusic, 1997; Cai and Zhang, 1997; Garcia et al., 2000) electrodes. The number of stimuli delivered has fluctuated from 10 to 100, usually 10. Most authors have used supramaximal stimuli of 0.1–0.2 ms duration. 9.2.4.6. Normative data Available normative data in adults and children from the literature are summarised in Tables 9.1 (median nerve), 9.2 (ulnar nerve), 9.3 (common peroneal

R.J. GUILOFF

nerve), 9.4 (posterior tibial nerve) and 9.5 (facial, axillary, radial and femoral nerves). The number of stimuli used, to obtain such data, is indicated in the tables. 9.2.5. Summary Surface recorded F-waves are mostly indirect late responses to supra-maximal stimulation of a mixed or motor peripheral nerve, mostly generated by recurrent discharges of, on average, 2–3 motor neurons. About 1–5% of the motor neuron pool responds to the supramaximal antidromic volley. All groups of motor neurons can produce F-waves but they are generated more frequently, or preferentially, by the faster motor neurons. Thus, the spectrum, but not the distribution, of conduction velocities of the motor axons of a peripheral nerve is reflected in the spectrum of latencies (chronodispersion) and conduction velocities (tacheodispersion) of the population of F-waves. Conventional and single motor unit F-wave studies indicate that the fastest or shortest F-wave indexes adequately the fastest motor axon as recorded in conventional motor conduction velocity studies. F-wave repeaters are F-waves with shapes that occur more than once during the recording. They are similar to other F-waves except that their amplitude and area are greater. They appear to represent groups of motor neurons that backfire together repeatedly, more than would be expected by chance, probably because of anatomical and functional arrangements between such motor neurons. The F-wave methodology and technique have been gradually refined over the years. Recordings are made with surface electrodes over the motor point or belly of a muscle. 20–25% supra-maximal stimuli at 1 Hz are used. Gain is usually set at 0.2 mV/division, filters are set a 20Hz–2kHz, sweep speed at 5 ms/division for the upper limb and 10 ms/division for the lower limb, but longer sweeps may be needed in pathological conditions. The position of the stimulator cathode and anode does not seem to be of importance. The number of stimuli delivered needs to ensure that the values obtained are representative of the population of motor units sampled by the method. This will depend on the parameters of interest. For routine recordings, 20 stimuli may be adequate for minimum latency or maximal conduction velocity, median latency, amplitude and persistence. Chrono and tacheodispersion require at least 60 stimuli and repeater studies 100 or more. However further work is required to establish the best sample size in pathological conditions.

LATE RESPONSES (F- AND A-WAVES): METHODOLOGY AND TECHNIQUES

The most commonly used parameters are minimum latency, F-wave maximum conduction velocity, chronodispersion and persistence. All measurements need to be related to age and temperature. All absolute latency measurements need nomograms relating them to limb length or height. Though shortest latency and F-wave maximal conduction velocity are wellestablished parameters, chronodispersion, tacheodispersion and persistence are increasingly being used, as they appear to be more sensitive in a number of pathological conditions. The method allows measuring motor conduction in the whole of the peripheral motor pathway or in defined proximal or distal segments of peripheral nerves. It is particularly suited for documenting abnormalities of motor conduction in proximal segments not testable by conventional motor conduction studies. 9.3. A-waves 9.3.1. Introduction and nomenclature A number of late responses have latencies intermediate between the M-response and the F-wave and Hresponses. They include motor axon reflexes (MARs) elicited with sub-maximal, but not maximal, stimulation (Fullerton and Gilliatt, 1965), indirect double discharges (IDDs) (Roth, 1974, 1985) or peripheral late wave responses (PLWs) (Tomasulo, 1982), elicited with either sub-maximal or, in routine F-wave recordings in humans, maximal stimulation. Ephaptic responses originating at the myo-axonal junction (MAERs) have also been described with the same range of latencies of MARs, IDDs and PLWs. (Roth, 1993). MAERs are recognized using both weak and strong stimuli and double stimulation techniques and also occur at latencies intermediate between the M- and the F-waves (Roth, 1993). Motor axon loop responses (MALRs) (Roth and Egloff-Baer, 1984) are also seen at latencies between those of the M- and F-waves. MALRs appear as late potentials, after the M-wave and before the F-wave, with proximal, but not with distal, nerve stimulation. A wealth of information on late motor responses can be found in Roth’s monography (Roth, 2000). There is no universally agreed definition of an “Awave,” nor a universally agreed methodology to record them. The term “A-wave” has been used to denote just motor axon reflexes (MARs) (American Association of Electromyography and Electrodiagnosis (AAEE), 1987), just indirect double discharges (IDDs) or

221

peripheral late wave responses (PLWs) (Bischoff et al., 1996; Kornhuber et al., 1999) or both (Magistris and Roth, 1992; Rowin and Meriggioli, 2000). IDDs and PLWs may be different names for the same phenomenon looked at with different techniques (Magistris and Roth, 1992). Recently, stable and frequent constant waveforms reported to occur with supra-maximal stimuli within the latencies of F-waves or later have been also considered as “A-waves,” or similar to PLWs (Bischoff et al., 1996; Puksa et al., 2003a). However, the frequently occurring, constant shape and latency, waveforms that are recorded with latencies intermediate between the M- and F-waves during F-wave clinical recordings with surface electrodes and supra-maximal stimulation in patients with peripheral nerve disease, but also less frequently in normal subjects, are not recordings of motor axon reflexes as described originally (Fullerton and Gilliatt, 1965). They are probably, at least partly, recordings of IDDs or PLWs (Magistris and Roth, 1992; Bischoff et al., 1996; Rowin and Meriggioli, 2000). PLWs might also sometimes include MAERs within the waveforms recorded. These different late responses have been considered in detail separately below. 9.3.2. Motor axon reflexes (MARs) Fullerton and Gilliatt first described motor axon reflexes in humans in 1965 (Fullerton and Gilliatt, 1965) (Fig. 9.6). They recorded these all or none responses from 9 out of 25 patients with lower motor neuron lesions affecting the hand using both surface and coaxial intramuscular electrodes in abductor pollicis brevis, first dorsal interosseous or abductor digiti minimi. They used stimulus voltages continuously variable up to 300 V. These authors reported to have never seen responses of this type in normal subjects. MARs were only obtained with sub-maximal stimuli but once recorded, they appeared constantly with every stimulus and, also contrary to the F-waves, always with a constant latency, usually intermediate between the M- and F-waves, but sometimes longer (Magistris and Roth, 1992). By moving the stimulating electrode proximally along the course of the nerve there was a decrease in the latency of the response, whilst the latency of the M-response increased, indicating that conduction for MAR was initially in a proximal direction. The latency was too short for the response to be the result of an impulse travelling antidromically to the spinal cord and then back to the

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R.J. GUILOFF

AR

S M

F

200 μV

0

10

R

20

30

40 msec

S

Fig. 9.6 Motor axon reflexes. Mr J. B. Cervical rib syndrome. Muscle action potentials from abductor digiti minimi recorded through surface electrodes. S = stimulus. In addition to M- and F-waves, a small potential (AR) is present with a latency of 20 ms. Eight successive individual traces are shown. Calibration bar 200 μV. See also text (Reproduced from Fullerton, Gilliatt (1965) J. Neurol. Neurosurg., Psychiat., 28: 1–11, with permission from the BMJ Publishing Group).

muscle. The MARs were always preceded by an Mresponse and always appeared in an all or none fashion. Increase in the intensity of the sub-maximal stimulus did not change the waveform but, at a critical level, resulted in a sudden disappearance of MAR. Needle recordings showed that at this stage there was a sudden jump in latency to a value close to the M-wave. Fullerton and Gilliatt interpreted these results as evidence that the weak stimulus excited antidromically one (afferent) branch of an axon spreading to a point of axonal branching, and passing then to another (efferent) branch of the same axon that would conduct orthodromically and produce the potential recorded as MAR. When the stimulation was increased, both branches of the axon would be stimulated distally and the branch previously producing the MAR suddenly produced its own M-response, which would have the same configuration as the MAR as the same muscle fibers would be activated. With a strong stimulus the MAR could not occur after the M-response of the second (efferent) branch because the antidromic wave generated in the second axon would collide with the MAR generating incoming antidromic impulse

from the first (afferent) axonal branch (Magistris and Roth, 1992). Fullerton and Gilliatt measured the conduction velocity in the afferent and efferent branch of the reflex in two patients. In one the afferent branch was fast conducting (50 m/s) and more excitable than the efferent branch (22 m/s). In the other both branches were slow conducting at 14.3 m/s (afferent) and 21 m/s (efferent), but the slowest conducting was more excitable. Others also accepted that MARs are mediated by motor axonal branching (Stalberg and Trontelj, 1970; Roth, 1978, 1979). The M-response of the first axonal (afferent), branch could not be demonstrated in the initial descriptions. This M-response has been subsequently recorded with a different waveform to the MAR produced by the second (efferent) branch, and within the same territory covered by the concentric needle electrode that was recording the MAR. Characteristically, the latency of this response increased with more proximal stimulation whilst that of the MAR in the other branch was decreased. During voluntary activity, the same two components of the MAR have appeared as coupled discharges with a constant interval between the two orthodromic (M) waveforms, the one in the second (efferent) branch having the same waveform as its MAR response (Roth, 1979; Magistris and Roth, 1992). Changes in latency of single muscle fibers potentials to stimuli of various strengths to nerve trunks and intramuscularly in normal subjects were also thought to be due to motor axonal reflexes (Stalberg and Trontelj, 1970). They found such changes in all four subjects with intramuscular stimulation and in some of the 9 subjects with nerve trunk stimulation. Fullerton and Gilliatt considered and rejected the possibility that MARs represented the responses to transmission of impulses through artificial synapses between two axons (Granit and Skoglund, 1945), because of the constant latency and waveform, the lack of fatigue with rapid rates of stimulation and the persistence with paired stimuli separated by 1 or 2 ms. Finally, they pointed out that axonal branching had been already demonstrated anatomically in normal fibers in man and in animals (Lavarack et al., 1951; Sunderland and Lavarack, 1953) and in cat muscle nerves deprived from their afferent fibers by previous removal of the dorsal root ganglia (Eccles and Sherrington, 1930). The subsequent arguments as to whether the responses described by Fullerton and Gilliatt represent motor axon reflexes or ephaptic transmission will be considered later.

LATE RESPONSES (F- AND A-WAVES): METHODOLOGY AND TECHNIQUES

Serial studies in patients with median and ulnar nerves acutely and totally severed in the arm, forearm or wrist, sutured by first intention, have been reported (Montserrat and Benito, 1990). The authors recorded the thenar or hypothenar muscles with surface electrodes and coaxial needles and stimulated the digital nerves of the corresponding fingers at least three times over periods of up to 11 years from the day of the surgical repair. The earliest response was seen at five months, and the latest at 18 months, after the repair. The reflex responses had constant, often complex, shapes and a greatly variable latency that was shorter in more distal lesions and the longer the time elapsed since re-innervation. The reflex responses were abolished by collision if a second stimulus was applied at the wrist, before the stimulus to the finger, with an interval shorter than the latency of the reflex response. Repetitive stimulation showed that the reflex response behaved like an M-wave. F- and H-waves were not studied but many of the responses had latencies intermediate between those of the M-wave and those expected for the F-response. The authors could not decide, with the data they had, whether the observed muscle responses resulted from axon reflexes or ephaptic transmission. They considered: (a) misdirected re-growth of a motor axon to a sensory endoneural tube mediating a true axon reflex; (b) ephaptic transmission from two re-grown axons, one sensory and one motor: and (c) ephaptic transmission from a motor axon misdirected to a sensory endoneural tube to another motor axon. 9.3.3. Indirect double discharges (IDD) Indirect double discharges were first reported in 1974 (Roth, 1974) and subsequently further studied (Roth, 1985; Magistris and Roth, 1992). These authors state that IDDs represent the same phenomenon as the late peripheral waves described with supra-maximal stimulation by Tomasulo (1982). IDDs are motor units recorded with needle electrodes using submaximal or maximal stimulation. They consist of two all or none identical waveforms elicited above a certain intensity of stimulation, one with a latency within the M-wave range and another with a latency intermediate between the M- and the F-wave (Fig. 9.7). With maximal stimulation only the second waveform is seen as the first becomes buried in the Mresponse. The first waveform of the IDD is the result of orthodromic conduction as its latency increases with

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1

2 3

4

5 6

7 1 mv 5 ms

Fig. 9.7 Example of an indirect double discharge (IDD). A 58-year-old patient with sensory motor polyneuropathy showing signs of demyelination and axon loss related to diabetes mellitus and amyloidosis. Stimulation: ulnar nerve. Recording: abductor digiti minimi (concentric needle electrode): (1) With maximal stimulation above the elbow, an intermediate latency response is recorded; it is sometimes followed by its F-response (not shown). In this case, the response is “constant” (evoked by every stimulus) and has a “stable” latency (no conspicuous variation in latency). (2–3) Graded intensity of stimulation performed above the elbow shows that it is one of the first potentials recruited; 2 all-or-none potentials of identical shape are now visible. (4) When stimulation is moved distally, to the wrist, the first potential has a shorter latency (indirect pathway); they correspond to the direct and indirect components of the IDD. At the wrist, the axon giving rise to the IDD has the lowest threshold to stimulation; the indirect component is never recorded without its direct counterpart. (4, 6) With an odd number of stimuli the indirect response is evoked by the last stimulus. (5, 7) With an even number of stimuli the indirect response is abolished by collision. (5–7) The direct response is evoked by every stimulus of the train. Reproduced from Magistris, MR et al. (1992) EEG. Clin. Neurophysiol., 85: 124–130, with permission from International Federation of Clinical Neurophysiology.

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more proximal stimulation. It is constant as it is present with every stimulus, and it is stable, as its latency is constant. It can be evoked by all the stimuli in a train. The second wave of the IDD is an indirect response requiring an antidromic arm as its latency decreases with more proximal stimulation. It is constant, appearing with every stimulus, in 50% of recordings. Its latency varies between 0.5 and 4 ms in 75% and is less than 0.5 ms in 25% of recordings. Further, the second wave can be abolished by an even number of multiple stimuli in a train from two upwards, but may appear in response to the last stimulus in a train with an uneven number of stimuli. 9.3.4. Peripheral late waves (PLW) Peripheral late waves were first reported in 1982 (Tomasulo, 1982). They were obtained with supramaximal stimulation, recorded with surface electrodes and had latencies intermediate between the M- and Fwave. Their latency decreased with more proximal stimulation, indicating a proximal direction of travel for the stimulus eliciting the response and a source proximal to the stimulation site for the orthodromic volley producing the PLW (Fig. 9.8). The PLW did not change; each different waveform repeated many times, unchanged, with trains of stimuli at a largely constant latency, with amplitudes usually lower than that of the F-waves. Tomasulo recorded from the

sole of the foot (flexor hallucis brevis, abductor hallucis and flexor digiti quinti). He took 39.4 ms as upper limit for the latency of the PLW, measured from the stimulus, calculated as the normal mean Fwave latency of 52.3 ms less three standard deviations (4.4 ms). Due to the characteristic latency of the PLW he concluded that they could not have a spinal origin. Constant waveforms within the F-waveform, or occurring later than F-waves, have been subsequently considered as PLW or A-waves (Bischoff et al., 1996; Puksa et al., 2003a) (Fig. 9.9). Conclusive proof that such constant waveforms within the F-wave or occurring later are not repeater F-waves, or that they are of non spinal origin and similar to the responses with intermediate latencies between the M- and the Fwaves, originally described by Roth and by Tomasulo, is not available. It is also not clear whether waveforms after the F-waves could represent F-waves generated by myo-axonal ephapses (Egloff-Baer and Roth, 1979) (see below). Tomasulo could not correlate the presence of PLW with peripheral nerve diseases. He found PLWs in the muscles of the foot of 21.5% of normals and in 35.7% of symptomatic patients, a statistically not significant difference (Tomasulo, 1982). Using stimulation protocols for F-waves, with 15–20 stimuli a number of authors, have confirmed that PLW can be seen in normal subjects (Raudino, 1989; Bischoff et al., 1996; Rowin and Meriggioli, 2000; Gozke et al., 2003). In the only study specifically addressing this point, they

M 6.5 msec

Stim

10 msec

25.5 msec

5.7 msec 26.5

100uv

Fig. 9.8 Peripheral late wave. Patient age 50 with diabetic neuropathy and burning feet. Recording is from the flexor hallucis brevis muscle, with maximal stimulation of the posterior tibial nerve at two sites in the ankle region separated by 2 cm. Upper trace: Stimulus at proximal site. Lower trace: Distal site. The M-response is earlier by 0.8 ms with distal stimulation, whereas the late response is delayed by 1 ms. The latency of the late response, 26.5 ms, is too short for conduction to the spinal cord and back. The response is probably due to reflection of an impulse from a site of demyelinatiom. Reproduced from Tomasulo,WNL (1982) Neurology, 32: 712–719 with permission.

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peripheral nerve disease than in normal subjects (Raudino, 1989; Bischoff et al., 1996; Rowin and Meriggioli, 2000). Both IDDs (Roth, 1974, 1985) and PLWs (Tomasulo, 1982) were originally postulated to be caused by proximal re-excitation of the motor axon, perhaps by a reflection of the ascending impulse at an area of discontinuity of the myelin sheath (Tomasulo, 1982). The arguments against this hypothesis, namely that IDDs and PLWs are mediated by ephaptic transmission between motor axons of different motor neurons are presented below.

early

9.3.5. Myo-axonal ephaptic responses (MAERs)

late

R Per n 0.5 mV 10ms

L Per n 0.5 mV 10 ms

Fig. 9.9 Early and late A-waves occurring before and after F-waves. Control subjects. Early (right common peroneal nerve) and late (left common peroneal nerve) A-waves before and after F-waves are marked by arrows. Dual gain is used; during the M-wave the gain is 10 mV/division and during the late responses it is 0.5 mV/division. In the examples shown the early A-waves occur following every stimulus, whereas the late A-wave does not occur every time. Not shown here are the A-waves occurring among the F-waves and combined multiple late and early A-waves. From Puksa et al. (2003) Muscle Nerve, 28: 626–629. Reprinted with kind permission from John Wiley and Sons, Inc.

were reported in 2% (median and ulnar nerves) to 14–25% (peroneal and tibial nerve, respectively) of normal subjects (Puksa et al., 2003a). They appear to be found in a higher proportions of patients with

Myo-axonal ephaptic responses are usually unstable motor unit responses with a latency intermediate between M- and F-waves. They might be confused with MARs or IDDs if not assessed appropriately and, therefore, classified as A-waves. Stimulation of peripheral nerve terminations by muscle action potentials was originally described in animals by Lloyd (1942). Physiological ephaptic responses in normal humans have been described as a result of re-excitation of a nerve by its own muscle potential (Epstein and Jackson, 1970; Roth and EgloffBaer, 1979; Egloff-Baer and Roth, 1979). Several motor units appear to be involved in the response that follows closely the M-wave (Roth and Egloff-Baer, 1979; Roth, 1993). MAERs have also been described in patients with chronic neuropathies (Roth and Magistris, 1985; Roth, 1993) as all or none inconstant motor unit action potentials occurring with an unstable latency, well after the M-wave and before the F-waves. All muscles studied had chronic partial denervation. The MAERs were recorded with needle or surface electrodes in the thenar, first dorsal interosseous, abductor digiti minimi and abductor hallucis muscles using variable stimulation intensities at different sites, and sometimes double stimulation techniques (Fig. 9.10). The range of maximum variation in latency in 20 MAERs in these patients was 0.2–1.5 ms, but 2 of the MAERs had a stable latency. They followed the M-wave by 12.5–45.5 ms. Only 6 of the 22 were evoked by every stimulus. They disappeared by stretching the muscle. The MAERs can be followed by F-waves of either the pre-ephaptic or in the post ephaptic motor unit. The cancellation of the post-ephaptic motor unit F-wave, by collision of the descending volley with the ascending one generated at the ephapse, when an MAER is evoked, seemed characteristic of this type of ephapse.

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1mV ER 10ms 1

ER

DR

2

DR DR2

ER ER2

3

Fig. 9.10 Myo-axonal ephaptic responses (ER). A 67-year-old woman with old carpal tunnel syndrome. Recording from the thenar muscles with a concentric needle electrode. One way ephapse between the muscle fibers of a reinnervated motor unit a ( ) and the contiguous post ephaptic axon of another motor unit b ( ). The recorded motor unit potential is that of b. The stimulation of the median nerve at the wrist is above threshold. The pre-ephaptic axon has the highest excitability and therefore the lowest stimulation threshold. There is desynchronization of the M-wave: (1) The evoked ephaptic response (ER) of motor unit b, by stimulating the lower threshold motor unit a, is constant and has an unstable latency. (2) Both motor units are stimulated. The direct response of motor unit b (DR) is constant and has a stable latency; it is followed by the ephaptic response of b evoked by the stimulation of a. (3) With paired stimuli to both axons a and b (interval S1−S2 = 10 ms) two direct responses are seen (DR and DR2), the ER is not cancelled and the second stimulus sometimes evokes a second ephaptic response (ER2). Reproduced from Roth, G (1993) EEG Clin. Neurophysiol., 89: 252–260 with permission from International Federation of Clinical Neurophysiology.

Evidence was presented to suggest that MAERs result from the one way ephaptic excitation of the distal portion of a motor axon or one of its branches by the synchronized potential of a group of re-innervated muscle fibers belonging to another motor unit. That the post-ephaptic structure is axonal was suggested by the close association between a late potential (IDD) and an MAER when they were recorded together, the first always closely preceding the second, suggesting that activation of a re-innervated unit led to activation of another unit producing the MAER. The possibility that the pre-ephaptic structure was axonal rather than

re-innervated muscle fibers could not be completely discarded. The MAER’s latency similar to that of IDDs, and the fact that the electrical signal would be far higher and more likely to excite the post-synaptic structure in a group of re-innervated muscle fibers than in a regenerating axon, favoured re-innervated muscle fibers as the pre-synaptic structure. A distinguishing feature of the MAER from a late potential (IDD) would be that maximal double stimulation always abolishes the second wave of the IDD because of collision between the descending response to the first stimulus and the antidromic volley to the second stimulus in the same motor axon. In the case of the one way ephapse however, the MAER to the first but not to the second stimulus would be abolished by double stimulation. The first post-ephaptic descending volley collides with, and is cancelled by, the second antidromic volley in the same post-ephaptic axon, whilst the second post-ephaptic descending volley can be recorded (Magistris and Roth, 1992). In practice, however, it was described that double stimulation evoking also the direct response had frequently, but not always, an inhibitory effect on the MAER and that “when the second direct response preceded the MEER by less than 15 ms, the MAER always disappeared.” Roth concluded that the inconstancy of the MAER and the variability of its latency related to the excitation of the post-ephaptic axon and that the refractory period of the myo-axonal ephaptic mechanism was much longer than that of axons. Thus, the distinction of MAERs from other late responses requires not only stimulation at different sites with varying intensities but also double stimulation (Roth, 1993). Further, the F-wave generated by the re-excitation of some axons by the ephapse can occur after the F-wave generated by axons stimulated directly. Such F-waves could be erroneously interpreted as late F-waves or A-waves. However, their latency increases with more proximal stimulation since the stimulus travels a longer way to the ephapse within the muscle that will trigger both an orthodromic and an antidromic volley in a neighboring motor axon (Egloff-Baer and Roth, 1979). 9.3.6. The motor axon loop response (MALR) (Fig. 9.11) These waveforms can appear as late responses with latencies intermediate between the M- and F-waves. They might be confused with MARs if not investigated adequately and therefore described as A-waves

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A 1

S

LR

LR

2

B LR 3 BR

LR

4

BR

C BR

BR 6

Fig. 9.11 The motor axon loop response. A 33-year-old woman with a traumatic lesion of the brachial plexus. Recordings from the abductor pollicis brevis muscle. Weak ( ) or strong ( ) stimulation (S) of the median nerve. (1) at the axilla, (2) below the axilla, (3) at mid upper arm, (4) above the elbow, (5) at the elbow, and (6) at the wrist. In (A) only the late response is elicited (LR), even with very strong stimuli. In (B), the LR and the short latency response (BR) are evoked by weaker and stronger stimuli respectively. In C, only the BR is evoked. Its presence in the M wave was ascertained by the collision method (not shown). At this recording site, no response can be elicited by ulnar nerve stimulation at the wrist or elbow. The distances (cm) recorded were: 1–2 = 6.5; 2–3 = 3; 3–4 = 6; 4–5 =2; 5–6 = 22. Calibration bar = 5 mV, 2 ms. Reproduced from Roth, Egloff-Baer, (1984) Muscle Nerve, 7: 294–297, with permission from John Wiley and Sons, Inc.

in needle or surface unit recordings with sub-maximal stimulation. (Fig. 9.11). MALRs were described with concentric needle electrodes, surface stimulating electrodes with variable intensity, and sometimes a collision technique, in seven

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patients with median or ulnar nerve lesions, two with brachial plexopathies and one without clinical evidence of peripheral nerve lesion (Roth and EgloffBaer, 1984). The waveform was detected with proximal stimulation of the nerve trunk and the motor unit showed a stable late direct response, between the M- and the F-wave, with weak stimuli. When the stimulation reached a critical level, there was a sudden jump to a stable direct shorter normal latency. The short response was shown to be part of the M-wave. The latency difference between the two responses remained identical when stimulating the restricted segment of the nerve from where they could both be elicited. Above this segment, only the late response, and below it only the short latency one, were elicited. The authors suggested that the segment had a loop with a descending initial proximal portion kinking upwards with a reversal of the direction of conduction which becomes distal to proximal. The second segment would then kink downwards and this third and final segment would conduct in a proximal to distal direction. The proximal, initial, portion of the loop is the most excitable and proposed to be the origin of the late response. The final portion of the loop is the proposed origin of the shorter latency response. The recurrent section of the axon in the loop is the less excitable one, perhaps because it is furthest away from the stimulating electrode and produces no responses. The proposed kinked axons had normal conduction velocities. The late response was sometimes followed by its own F-wave with a longer latency than that of the F-waves of normal motor units embedded in the M-wave. The F-wave latency of the shorter latency waveform, at the segment where a late response can also be evoked with a weaker stimulus, is the same as the F-wave latency of the late response evoked at the same site. This is because the path for both F-waves includes the loop, for the stimulus evoking the short latency response during the antidromic afferent phase of the recurrent response and for the stimulus evoking the late response during the orthodromic efferent phase of the recurrent response. The authors quote anatomical evidence for the three portions of the loops residing in more than one nerve but propose than the whole loop may also be contained within one nerve. 9.3.7. Ephaptic transmission, axonal reflexes and recurrent re-excitation of motor axons Tomasulo suggested that the techniques used to describe human motor axon reflexes and peripheral

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late waves could not distinguish ephaptic transmission between the axons of two motor units from either the axonal branching proposed for MAR, or the recurrent re-excitation of motor axons at a proximal area of the axon proposed for the PLW (Tomasulo, 1982). The idea was later formally put forward as a hypothesis (Gilchrist, 1988). According to this hypothesis bi-directional ephaptic transmission between the axons of two motor units would result in “motor axon reflexes” and unidirectional ephaptic transmission would result in “peripheral late waves.” Cogent arguments against this hypothesis were presented in 1992 (Magistris and Roth, 1992) by comparing models of ephaptic transmission with the observed facts available for MAR, IDD and PWL. In the case of MARs observed features, not expected with bidirectional ephaptic transmission, were the stable latency of its F-waves, the lack of repetitive firing of MARs, the constant latency with stimulation above the site of reflection, the faster conduction above the site of reflection than in the branches, and the lack of an own firing rhythm in one of the potentials of the coupled discharge. In the case of IDDs and PLWs, observed features, not expected with unidirectional ephaptic transmission, were the constant recording of a direct response preceding the indirect response (unless buried in the M-wave), the cancellation of the indirect response by maximal double stimulation, the similar thresholds of the direct and indirect responses within the IDDs and their lack of F-waves with multiple latencies. 9.3.8. Methodology and techniques There is currently no agreed upon nomenclature for, nor agreed methodology or technique to study, Awaves, or to dissect the responses that may underlie the waveforms seen with an intermediate latency between the M- and F-waves, in a clinical setting. Bischoff and his co-workers suggested the following criteria to classify as A-waves the waveforms seen during 20 supra-maximal stimuli in protocols designed to record F-waves with surface electrodes (see above in the F-wave section): (A) Potentials of stable amplitude and shape; (B) variation in latency of less than 1.5 ms; and (C) occurrence in at least 8 out of 20 stimuli (Bischoff et al., 1996). In another study, criteria B was changed to less than 0.5 ms (Puksa et al., 2003a). With these criteria waves that are part of the F-waveforms or that occur later can be included, but cannot be separated from F-waves repeaters, longer latency F-wave repeaters or other late responses. The frequent occurrence of the waveforms (at least eight out of 20) was considered to be more likely in

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A-waves than in F-wave repeaters (Puksa et al., 2003a). These authors described A-waves intermixed with F-waves in lower, but not upper, limb nerves in 2% of normal subjects. An F-wave shape repeating 11 times out of 200 stimuli was seen in the abductor digiti minimi of two out of 21 (9.9%) normal subjects in one study (PeioglouHarmoussi et al., 1985a). A mean of 9.9% (up to 20% in individual subjects) of repeater shapes out of 1398 F-wave shapes were seen in the abductor digiti minimi of 30 normal subjects following 60 stimuli in another study. Up to nine of these repeater shapes repeated 5 or more times (Guiloff and Modarres-Sadeghi, 1991). Three examples of repeater F-waves in the thenar eminence of patients with carpal tunnel syndrome are illustrated by Macleod (1987). One shows 16 late repeater F-waves out of 39 successive sweeps, another shows just the same F-wave repeater occurring 13 times in 49 sweeps and a further one shows a repeater occurring 13 times in 34 sweeps. Similar data for F-waves in the lower limbs are not available so that a comparison with the data on A-waves available in the lower limbs (Puksa et al., 2003a) is not possible at present. Waveforms occurring near the M-waves, whether IDDs, PWLs or MALRs, may be separated from late components of the M-wave with changes in the stimulation site. More distal stimulation should show an increase in the latency of IDDs and PLWs and disappearance of MALRs. More proximal stimulation should show IDDs and PLWs, but not MALRs, to be closer to the M-wave. Late F-waves associated with myo-axonal ephapses have a longer latency the more proximal the stimulation site (Egloff-Baer and Roth, 1979). Studies with simultaneous needle and surface recordings, double stimulation techniques and variable intensities and sites of stimulation may help in clarifying whether the constant shape and latency waves seen within the F-waveform, or after it, when using F-wave protocols with supra-maximal stimulation are A-waves (i.e., motor axon reflexes, direct double discharges or peripheral late waves), repeater F-waves or other late responses such as late F-waves associated with myo-axonal ephaptic responses. It has been suggested that intermediate latency late responses are best studied looking at all or none potentials with needle recordings. MARs can then, for example, be separated from IDDs when the direct response is hidden in the M-wave, by using the double stimulation technique (Magistris and Roth, 1992). It seems doubtful, however, that such methodology will be widely applied in routine clinical settings.

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9.3.9. Summary The term “A-wave” has been used to denote motor axon reflexes, indirect double discharges and peripheral late waves with intermediate latencies between those of the M- and F-waves or all of them. It has also been used recently for constant and frequent late responses to supra-maximal stimulation within the F-wave waveform or occurring after the F-waves. Other potentials with latencies between those of the M- and F-wave include myo-axonal ephaptic responses and motor axon loop responses. The F-waves of myo-axonal ephapses and some repeater F-waves may also occur after the usual Fwaves. All these responses may be seen in normal subjects. The various late responses can be differentiated using a number of techniques including needle recordings of all or none potentials, stimulation of varying intensities and at various sites, double stimulation and trains of stimuli, all of which are not usually used in routine clinical investigation. The frequent constant waveforms of stable intermediate latency between M- and F-waves, usually termed A-waves, seen during routine F-wave studies with surface recordings and supra-maximal stimulation, are likely to be IDDs or PLWs. They may be caused by the recurrent re-excitation of motor axons at a proximal point of discontinuity of the myelin sheath. References American Association of Electromyography and Electrodiagnosis (AAEE) NC (1987) AAEE glossary of terms in clinical electromyography. Muscle Nerve, 10 (suppl): G5–G6. Andersen, H, Stalberg, E and Falck, B (1997) F-wave latency, the most sensitive nerve conduction parameter in patients with diabetes mellitus. Muscle Nerve, 20: 1296–1302. Argyropoulos, CJ, Panayiotopoulos, CP and Scarpalezos, S (1978) F- and M-wave conduction velocity in amyotrophic lateral sclerosis. Muscle Nerve, 1: 479–485. Argyropoulos, CJ, Panayiotopoulos, CP, Scarpalezos, S and Nastas, PE (1979) F-wave and M-response conduction velocity in diabetes mellitus. Electromyogr. Clin. Neurophysiol., 19: 443–458. Barakan, TH, Downman, CBB and Eccles, JC (1949) Electric potentials generated by antidromic volleys in quadriceps and hamstring motoneurones. J. Neurophysiol., 12: 393–424.

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Peripheral Nerve Diseases Handbook of Clinical Neurophysiology, Vol. 7 J. Kimura (Ed.) © 2006 Elsevier B.V. All rights reserved

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CHAPTER 10

Reflex responses, silent period and long latency reflexes Josep Valls-Solé a, * and Günther Deuschlb a

Unitat d’EMG, Servei de Neurologia, Hospital Clínic, Facultad de Medicina, Institut d’Investigació Biomèdica August Pi i Sunyer (IDIBAPS), Universitat de Barcelona, Spain b Department of Neurology, Christian-Albrechts-Universität Kiel, Germany

10.1. Introduction The integration of sensory inputs into the motor commands for purposeful voluntary or automatic actions (sensorimotor integration) is a very complex function of the central nervous system. With regard to somatosensory inputs, reflex motor responses to external stimuli range from the simplest tendon jerk to quite elaborated behavioral responses. Probably one of the most important accomplishments of human kind has been the development of a sufficient supraspinal control of reflex behavior, in such a way that, to a certain extent, we are able to selectively suppress the unwanted reflex responses, or let them fully manifest with no limitations, if necessary. Typically, simple monosynaptic reflexes are less subjected to control than the complex long latency ones. Many clinical neurophysiological studies can be performed to show how the central nervous system exerts its control over the reflexes and, also, to document the loss of such control in certain neurological diseases. Reflexes involving motor responses can be monosynaptic or polysynaptic. Monosynaptic reflexes follow a segmental reflex arc, and are of relatively short latency. The most widely known monosynaptic reflex is the tendon jerk, which is the muscle response to mechanical activation of muscle stretch receptors. The action potential linked to the reflex muscle response is known as the T-wave. A similar muscle response can be obtained if the Ia fibers are depolarized by relatively weak electrical shocks applied to the supplying nerve. The response obtained in this way is known as

*Correspondence to: Josep Valls-Solé, MD, Unitat d’EMG, Servei de Neurologia, Hospital Clínic. Villarroel, 170 Barcelona, 08036, Spain. E-mail address: [email protected] Tel.: +34 93 2275413; fax: +34 93 2275783.

the H-reflex. Both, mechanical taps and electrical shocks are unnatural stimuli and, consequently, the reflexes induced by these stimuli are likely to be of little functional use. However, they are of great importance for both clinical work in patients with neurological diseases, and research in physiology of sensorimotor circuits. A stimulus activating somatosensory afferents of any type is capable of inducing polysynaptic reflexes. These reflexes may implicate multilevel spinal and supraspinal pathways and are of relatively long latency (long-latency reflexes). As a difference with monosynaptic reflexes that always have a predominant excitatory component, the polysynaptic longlatency reflexes can be either excitatory or inhibitory and, most commonly, they have both effects combined. To make these effects apparent, the examiner requires the use of specific methodological conditions. For instance, the responses to certain stimuli may be so weak that they will only appear if the motor system is already engaged in a voluntary contraction. Maintaining a background contraction allows also for identifying the inhibitory component of the reflex response, as a transient decrease of the tonic level of voluntary contraction. In other instances, a stimulus may not induce a reflex response on its own, but is able to modulate the reflex response to another sensory stimulus in the same reflex circuit or in a distant one. A number of these studies use the H-reflex as a probe for the effects of other stimuli. In the following paragraphs, we shall review the physiology of monosynaptic and polysynaptic reflexes, and describe methods and technical specifications for performing reflex studies and using them in the assessment of peripheral nerve disorders. The reader can also find useful information in the literature devoted to technical recommendations, published elsewhere (Gitter and Stolov, 1995a,b; Bischoff et al., 1999; Deuschl and Eisen, 1999).

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10.2. Monosynaptic reflexes 10.2.1. The T-wave 10.2.1.1. Physiological and technical considerations The T-wave results from monitoring the muscle response elicited by a mechanical tap to the tendon with a hammer containing an electronic device that triggers the oscilloscopic sweep. The tap induces a stretch of muscle spindles, which activate the Ia afferent fibers, sending an afferent volley to the spinal cord. The Ia afferent terminals have monosynaptic excitatory connections with the alpha motoneurons innervating the stretched muscle. If the motoneuronal excitatory postsynaptic potential (EPSP) is synchronized and large enough, several motoneurons will discharge, producing a physically evident movement or muscle contraction. Electrophysiological recording of the T-wave permits to quantify the reflex response observed in clinical practice. The electromyograph should be set for external triggering by the built-in hammer switch. Usually, a time window of 50 ms is long enough to reproduce the whole response in any human muscle, and a gain of 0.5 mV per division is enough to reproduce the response recorded within a bandpass frequency range of 10 to 1000 Hz. The most reliable parameter to measure in the T-wave is the onset latency, while its amplitude is less reproducible. The examiner has to be aware of some methodological constraints, which also apply to the clinical assessment of tendon jerks: A variable

Fig. 10.1 Elicitation of the H-reflex in the soleus muscle. (A) Diagram of the circuits of the H-reflex. S = Stimulus; R = Recording. (B) Electrode placement. (C) Recordings obtained with progressively increasing the stimulus intensity from the top downwards (recruitment curve). The stimulus is applied at the arrowhead (S). (D) Graphic plot showing the changes in the amplitude of the H-reflex and the Mwave, obtained with increasing stimulus intensity, expressed in percentage of the maximum amplitude of the M-wave.

strength of the tap will give rise to a different response latency and size; a variable degree of relaxation of the agonist and antagonist muscles may inhibit or facilitate the reflex response, and a different position of right and left limbs could be responsible for an asymmetry in the size of the response. Therefore, for the T-wave to furnish reliable data, the recording electrodes should be placed symmetrically in the muscles of either side, and reflexes should be tested by maintaining the same position of each limb to avoid differences in the degree of tonic muscle contraction. With these precautions, the strength of the tap seems not to produce significant changes in onset latency (Malcolm, 1951). It was once believed that the excitability of the tendon jerk depended on the existence of a selective drive from gamma motoneurons to intrafusal fibers. The hypothesized constant traffic of impulses in gamma efferents was considered responsible for the possibility to elicit the reflex response in the resting condition in ankle flexor muscles (Landau et al., 1960; Dietrichson, 1971). However, microneurography studies have demonstrated the absence of efferent activity directed to muscle spindles in relaxed subjects (Vallbo et al., 1979). Also, Burke and coworkers demonstrated the absence of changes in the nerve afferent discharges to ankle taps during anaesthetic blockade of the sciatic nerve, assumed to block the gamma drive (Burke et al., 1981a) or during central nervous system manoeuvres of reflex reinforcement

A

B Recording Stimulator

S

A

R

M

C

H

D Size (% of M)

S

M H

Stimulus intensity (mA)

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(Burke et al., 1981b). These experiments demonstrated that the size of the monosynaptic reflex depends on the spinal excitability state rather than on the fusimotor drive. 10.2.1.2. Methods of clinical interest for the study of peripheral nervous system disorders with the T-wave The most important asset of the T-wave is its apparent simplicity. It can be applied to the study of lesions of proximal nerve segments (both, radicular and plexular) or to the study of polyneuropathies. It is likely to be asymmetrically abnormal in patients with unilateral radiculopathies involving the muscles in which it can be easily obtained (soleus and quadriceps in the lower limbs, and triceps, biceps and brachioradialis in the upper limbs. It is usually delayed in disorders causing slowness of conduction velocity in large axons (Kuruoglu and Oh, 1992). Absence of the T-wave, together with a decrease in the amplitude of the sensory nerve action potentials, may be one of the first objective signs of nerve damage in patients with distal axonal neuropathies (Sandler et al., 1969). A particularly interesting muscle for T-wave examination is the masseter muscle. The mandibular reflex is elicited by tapping to the chin. However, it is often difficult to discern whether the reflex is normal in both sides, or even whether it is present at all, by inspection only. Therefore, electromyographic monitoring of the masseter or temporalis muscle response is of paramount importance in the evaluation of suspected brainstem lesions (Hopf et al., 1991; Hopf, 1994). The mandibular reflex circuit is also particular regarding the location of the cell bodies. In contrast to those of all other muscles in the body, the proprioceptive neurons of the jaw muscles lie within the neuraxis, protected by the blood-brain barrier from peripheral circulating agents (Graus et al., 1987). This is actually an important piece of information for the diagnosis of some disorders involving immunological aggressions to sensory neurons of the Gasserian ganglia, such as in some patients with Sjögren’s syndrome. These patients might have abnormalities in multiple cranial nerve reflexes induced by cutaneous stimuli, which are conveyed by sensory neurons lying in the Gasserian ganglia but, since the antibodies do not pass the blood-brain barrier, the mandibular reflex is left intact (Valls-Solé et al., 1990). This observation easily separates patients with sensory neuronopathies from those with axonal neuropathies, which may also be present in patients with connective tissue diseases (Lecky et al., 1987).

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10.2.2. The H-reflex 10.2.2.1. Physiological and technical considerations The H-reflex, named in the honor of Paul Hoffmann, the physiologist who described it for the first time in 1918, is a monosynaptic reflex that results from activation of alpha motoneurons by electrically induced Ia afferent excitatory volleys. The H-reflex is commonly examined in the soleus muscle. However, it is also easily elicitable in other muscles, such as the quadriceps and wrist flexors. It may also be observed in thenar muscles, tibialis anterior, and many other muscles, if a slight facilitation is induced with voluntary contraction (Cowan et al., 1986). Indeed, voluntary contraction can be used safely to study the H-reflex in muscles other than the soleus, since the contractioninduced facilitation allows for distinguishing more clearly the H-reflex from the M-response in proximal muscles (Burke et al., 1989). 10.2.2.1.1. The H-reflex in the soleus muscle. The frequency band pass, gain, and sweep speed of the electromyograph recommended for recording the H-reflex are the same as those used in the study of the soleus T-wave, i.e., a band pass of 10 to 1000 Hz, a recording window of at least 50 ms, and a gain of at least 0.5 mV. The main difference in the set up for the study of the T-wave and the H-reflex is the use of an electrical stimulus, rather than a tendon tap, for elicitation of the H-reflex. In the most conventional procedure, the electrical stimulus is applied to the posterior tibial nerve at the level of the popliteal fossa. Stimuli of relatively low intensity and long duration, typically 0.5 or 1 ms, are the most appropriate for selective depolarization of the Ia afferent terminals (Panizza et al., 1989a). Bipolar stimulation electrodes are perfectly suitable for all purposes in the study of the Hreflex. However, the use of the cathode placed over the nerve and the anode placed over the patella (as shown in Fig. 10.1) may be a safer procedure for selective activation of the Ia fibers in some cases (Hugon, 1973). There are several methods for recording the soleus H-reflex. The most common procedure is to attach the recording electrode over the soleus muscle around the point in which the two gastrocnemius muscles joint the Achilles tendon (slightly distal to the midcalf). The reference electrode is usually placed about 3 cm distally, and the ground electrode is located at a point between the stimulating and the recording electrode. In this way, the H-reflex may show a triphasic form, with a small initial positivity, because the active

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electrode picks up the soleus activity as an approaching vector through the mass of the gastrocnemius muscles. The soleus response may be recorded as an initially negative deflection, without the volume conducted positivity, by placing the active electrode just medial to the tibia, at a midpoint between the tibial tubercule and the medial malleolus (Daube, 1979; Kimura, 2001). The latency of the H-wave should be measured at the point of its initial deflexion, whether positive or negative, since this is the point of onset of reflex activity in the soleus. As with the T-wave, care should be taken with regard to the position of the knee and ankle joints. Both should be slightly flexed to avoid stretching of the gastrocnemius in the knee, which would prevent the stimulating current to reach the nerve, and unnoticed contraction of the tibialis anterior, which would cause reciprocal attenuation of the H-reflex. The soleus H-reflex has a latency of about 30 ms, while the direct M-wave recorded in the same muscle with supramaximal stimulus intensity occurs at a latency of about 5 ms. The latency of the H-reflex is similar to that of the F-wave, which can be elicited in the same muscle by the same stimulating electrode, using larger intensities. Therefore, the examiner must be aware of the characteristics that allow to distinguish relatively easily between the two responses (Table 10.1). The latency of the H-reflex (and, also, that of the F-wave) correlates well with subject’s height (Schimsheimer et al., 1987) and leg length (Braddom and Johnson, 1974). The best position for a careful evaluation of the soleus H-reflex physiological characteristics is with the subject seated, and a soft cushion supporting the knee slightly flexed at about 120 degrees, and the ankle joint passively flexed at about 110 degrees. Relatively sophisticated equipment can be used to stabilize the leg in order to have a more reliable determination of H-reflex amplitude changes (Hultborn et al.,

J. VALLS-SOLÉ AND G. DEUSCHL

1987). Studies involving measurement of the mechanical consequences of the reflex activation require monitoring of the force of the reflex muscle contraction, which may be done through a force transducer placed against the sole of the foot, or the ankle joint movement, which is done through movement transducers placed across the joint. A simple accelerometer attached to the moving segment may also be used to monitor movement if a constant control of the position of the ankle joint is not required. 10.2.2.1.2. The H-reflex in muscles other than the soleus. Apart from the soleus, the H-reflex can be elicited at rest in the flexor carpi radialis (FCR) and in the vastus medialis (VM), although the response is not as consistent as in the soleus. For a response to be accepted as an H-reflex, the response has to fulfill the following criteria: 1. It has to have a latency consistent with the reflex arc. A conduction velocity of 60–80 m/s, characteristic of the large fibers responsible for the H-reflex, would correspond to a latency of 17–21 ms in the FCR, and of 19–26 ms in the VM 2. The action potential amplitude should decrease with increasing the stimulus intensity: The H-reflex should be elicited by stimuli of an intensity below threshold for the alpha motor axons. With increasing the stimulus intensity it should first increase and then decrease together with a progressive increase in the M-wave. 3. It should disappear or diminish significantly with vibration applied to the muscles of the forearm. The stimulus duration and intensity are similar to those used for the soleus H-reflex. For the FCR H-reflex, the recording electrodes are placed over the belly of the flexor carpi radialis, usually at about 1/3 of the distance between the medial epicondyle and the radial styloid (Jabre, 1981). In monopolar recording, the reference electrode may be

Table 10.1 Differences between the H-reflex and the F-wave Observation

H-reflex

F-wave

Stimulus intensity (relative to threshold for the M-wave) Largest amplitude (relative to amplitude of the M-wave) Onset latency Shape of the action potential Effect of a weak tonic muscle contraction Effect of increasing the stimulus intensity Effect of vibration

Subthreshold About 50% Rather constant Smooth and constant Increase in amplitude Decrease in amplitude Inhibition

Suprathreshold About 5% Variable Irregular and variable Increase in persistence Increase in persistence No effect

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placed at a certain distance, away from median nerve innervated muscles. However, good responses can be obtained also with bipolar recording electrodes. The stimulus is applied to the median nerve at the elbow, with a stimulus duration of 0.5 or 1 ms, and allowing enough time between successive stimuli to avoid refractory effects. Even with these precautions, the Hreflex of the forearm muscles cannot be elicited in up to 10% of healthy volunteers. Although the use of surface recording electrodes is generally acceptable (Jabre, 1981), and is usually preferred for assessment of reflex excitability (Day et al., 1984), it should be pointed out that Deschuytère et al. (1976) who first described this reflex, found it in all his patients using needle electrodes. For the VM H-reflex, the recording electrodes are placed over the vastus medialis at the distal 1/3 of a line joining the middle point of the inguinal fold and the internal aspect of the knee. The stimulus is applied to the femoral nerve at the inguinal fold. Due to the depth of the nerve, it may be convenient to exert some pressure with the electrode against the superficial

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tissue, and the help of a sandbag may be considered after careful positioning of the stimulating electrodes (Valls-Solé et al., 1998). 10.2.2.1.3. The afferent volley eliciting the H-reflex. Although the H-reflex is considered to be a monosynaptic reflex, there is no consistent proof for that in humans. Certainly, the onset of the action potential should reflect the motoneuronal excitation induced by the fastest afferent fibers. The size of the motoneuronal EPSP generating a motor response can be calculated in humans as the density of the action potentials forming the predominant spike of the peristimulustime-histogram (Mao et al., 1984). An example of such recordings from the soleus H-reflex is illustrated in Fig. 10.2. In humans, the EPSP generating the soleus H-reflex by the electrical stimulation of the posterior tibial nerve has a duration of 3.5–5.5 ms (Ashby and Labelle, 1977; Burke et al., 1984). During this time, other afferent inputs may reach the same motoneurons after a synaptic delay in spinal interneurons and their action potentials summate to the

A C

50 ms

S

B

cusum

S

ms

Fig. 10.2 Analysis of the soleus H-reflex using the peri-stimulus-time-histogram (psth). A needle is inserted in the soleus muscle and focussed to pick up the activity of a single motor unit action potential. The subject is requested to make a weak contraction to steadily activate that motor unit at a stable frequency. An electrical stimulus at the appropriate intensity is delivered to the posterior tibial nerve in such a way that it elicits the H-reflex repetitively in the middle of a 500 ms epoch window (A). After a predetermined number of stimuli (i.e., after collection of 100 traces), an histogram is made in which each motor unit action potential is represented in the exact time in which it occurred along the 500 ms window (B). The excitatory effect of the stimulus is recognized by the accumulation of MUAPs at the corresponding response time (at about 30 ms in the case of the soleus H-reflex). The steepness and width of the peak of the psth reflect the strength and the duration of the EPSP, respectively. Another way of representing the changes in excitability of the motoneurons affected by the stimulus is the cumulative sum (cusum) of the points gathered in the psth histogram (C). To do that, the running average of a number of consecutive time bins of the psth (i.e., 5 bins) is plotted against time for each ms of the epoch. For instance, the average of the bins 1 to 5 gives rise to the first point, that of the bins 2 to 6 to the second point, and so on. The curve shows a nice response representing the effect of the stimulus on the activated motoneuron pool.

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developing response (Burke et al., 1981b, 1984). Furthermore, inhibitory inputs from Ib or type II fibers activated in the posterior tibial nerve also reach the motoneurons some time later, and generate motoneuronal inhibitory postsynaptic potentials (IPSPs), contributing to the termination of the action potential (Pierrot-Deseilligny et al., 1981). These observations lead to suggest that the size of the H-reflex does not depend only on the size of the excitatory afferent volley, nor on the excitability of the alpha motoneurons, but also on the excitability of some propriospinal interneurons, whose postsynaptic potentials will contribute to the peak and the termination of the action potential (Burke, 1985). 10.2.2.1.4. Presynaptic control of Ia terminals. The number of Ia afferent axons activated by the stimulus is not the only factor determining the size of the EPSP generated in the motoneurons by Ia afferent terminals. Another functional mechanism at play is presynaptic inhibition of those excitatory inputs before they reach the motoneurons (Frank and Fuortes, 1957; Hultborn et al., 1987; Rudomin, 1990). The mechanism of presynaptic inhibition was first demonstrated in the cat motoneurons by showing that the EPSP generated by group Ia fibers was depressed after conditioning stimulation of flexor muscles group I afferents, while there was no change in the size of the antidromic potential. This same behavior can be easily reproduced in humans by using vibration (Delwaide, 1973). Vibratory stimulation at the Achilles tendon that depolarizes Ia afferent fibers, causes inhibition of the soleus H-reflex and, at the same time, tonic activation of motoneurons (the tonic vibration reflex). Even though the H-reflex is inhibited, the motoneurons remain excitable through other pathways like the antidromic volleys, eliciting the F-wave (Hultborn et al., 1987) or the descending volleys generated by transcranial magnetic stimuli, eliciting the motor evoked potential (Nielsen and Petersen, 1994). It is already known that the Ia afferents activate the alpha motoneurons through a monosynaptic connection. However, colaterals from the same afferents terminate in spinal interneurons, which depolarize the Ia axons just before the synaptic terminal. These spinal interneurons, known as “primary afferent depolarization” (PAD) neurons, would establish axo-axonal connections to depolarize the Ia terminals, causing a reduction of transmitter release at the arrival of excitatory inputs. Another mechanism that could be responsible for similar effects would be postsynaptic remote

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dendritic inhibition (Rudomin, 1990). In this scheme, the Ia terminals would establish their excitatory synapse with distal dendrites of alpha motoneurons, giving room for blockade from other synaptic terminals in the same dendrite. The concept of presynaptic inhibition is very important for understanding physiological mechanisms of motor control over reflex responses (Stein, 1995). Neurons in charge of presynaptic inhibition are probably under the control of descending tracts (Rymer et al., 1979; Valls-Solé et al., 1994). Therefore, the CNS can use this mechanism to modulate the amount of Ia afferent terminal excitation desired at any given moment or with any given condition. Combining simultaneous presynaptic inhibition and postsynaptic excitation is like pressing at the same time the clutch and the accelerator to maintain the car ready for a speedy departure. The motoneurons are therefore set in a condition of preparation for very rapid firing. 10.2.2.1.5. The H-reflex recruitment curve. The Hreflex is best elicited with electrical stimuli of low intensity and relatively long duration, typically, 0.5 or 1 ms (Panizza et al., 1989a). A stimulus of an intensity slightly above the threshold for depolarization of the Ia afferents should induce the H-reflex as the only response of the soleus muscle, avoiding concomitant activation of motor axons. Bipolar stimulation electrodes are perfectly suitable for all purposes in the study of the H-reflex. However, the use of the cathode placed over the nerve and the anode placed over the patella may be a safer procedure for separate activation of the Ia fibers in some cases (Hugon, 1973). Increasing the stimulus intensity causes the H-reflex to increase and the direct M-response to stimulation of motor axons to appear. In normal subjects, a stimulus intensity slightly above threshold for activation of motor axons gives rise to a H-reflex of larger amplitude than the M-wave. As the stimulus intensity increases, the H-reflex amplitude continues to increase up to a certain size and then decreases to completely disappear. Progressively increasing the intensity of the stimulus causes depolarization of some motor axons together with the Ia axons. By increasing the stimulus intensity, more Ia afferent fibers will become depolarized, generating a larger afferent volley. Even though the afferent volley will be able to activate a larger number of alpha motoneurons, the size of the H-reflex will decrease rather than increase with increasing stimulus intensity. This may be due

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to collision between orthodromic and antidromic volleys in the motor axons (Magladery and McDougal, 1950) or, most likely, to active inhibition of alpha motoneurons induced by antidromic excitation of recurrent inhibitory (Renshaw) interneurons (Gottlieb and Agarwal, 1976). Renshaw’s cells induce inhibition not only in the parent motoneuron but also in neighboring motoneurons (Renshaw, 1941). Therefore, extinction of the H-wave reflects in part the effects of recurrent inhibition, a subject that will be dealt with in more detail below. Because of its physiological details, the recruitment curve of the H-reflex should be kept in mind in any studies in which the size of the H-reflex is taken as a measure of motoneuronal excitability to sensory stimuli. The following points of reference of the recruitment curve should be considered: The intensity threshold for the elicitation of the H-reflex; the intensity threshold for inducing the M-response, the amplitude of the H-reflex at M-threshold, the maximal amplitude of the H-reflex, the amplitude of the M-wave at maximal H, and the ratio between maximal H and maximal M. Both the amplitude of the H-reflex at M-threshold and the maximal amplitude of the H-reflex are usually described in percentage of the M-wave amplitude. 10.2.2.1.6. The H-reflex excitability recovery curve. Short interval repetitive elicitation of the H-reflex leads to a decrease in the H-reflex size. This is taken as an evidence for the conditioning effects of a stimulus, and has lead to the development of the paired shock technique for the study of the H-reflex excitability recovery curve. Taborikova and Sax (1969) were the first to describe the changes occurring in the H-reflex as a result of a conditioning posterior tibial nerve electrical stimulus. These authors showed an early phase of inhibition, attributed to depletion of neurotransmitter, a phase of relative reduction of inhibition, between 100 and 250 ms, attributed to a long-loop facilitation operating through bulbar and cerebellar centers, and a continuation of the inhibition up to more than 1000 ms. Actually, later studies demonstrated that post activation depression of excitability was not completely recovered until more than 8 seconds after the preceding stimulus (Crone and Nielsen, 1989). The paired (conditioning and test) shock technique for the assessment of the H-reflex excitability recovery has the important drawback of the fact that the test

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H-reflex is not necessarily generated in the same motoneurons that have been activated by the conditioning H-reflex, and other factors modifying the excitability of the motoneurons, such as recurrent inhibition and after-hyperpolarization, affect the results. In an attempt to overcome this problem, Pierrot-Deseilligny and Bussel (1975) proposed a method to study more carefully the early part of the H-reflex excitability recovery curve, and the effects of recurrent inhibition (see below). These authors made sure that the H-reflex elicited by the test stimulus was due to activation of the same motoneuron pool giving rise to the response to the conditioning stimulus by provoking a conduction block in all other motoneurons using a test stimulus of supramaximal intensity. On doing so, Pierrot-Deseilligny and Bussel (1975) obtained evidence for the participation of recurrent inhibition in the early part of the excitability recovery curve. An observation of interest for clinical studies is that the inhibitory effect of repeated stimulation is reduced during voluntary contraction (Burke et al., 1989). Therefore, higher stimulation rates during voluntary contraction can be used for assessment of the H-reflex in muscles in which it is not usually obtained at rest. 10.2.2.1.7. The H-reflex as a measure of the spinal excitability state. Because of the monosynaptic nature of the early part of the response, the H-reflex can be safely used as a measure of segmental motor excitability, and of its control by descending pathways (Schieppati, 1987). The efficacy of afferent inputs in activating the alpha motoneuron depends on the excitability state of the H-reflex arc, including the motoneuron itself, the interneurons mediating presynaptic inhibition of the Ia terminals, and other propriospinal interneurons. Indeed, the excitability of the soleus H-reflex arc is known to change in relation to a large number of conditions (Misiaszek, 2003). Two main questions are, however, largely unsolved: What descending pathways are actually carrying the modulatory effects? and What is the exact target of those modulatory effects? 10.2.2.1.7.1. Modulation of the soleus H-reflex by remote muscle contraction. Various maneuvers cause modulation of the soleus H-reflex in humans. Probably the first to be considered, and the most known, is the Jendrassik maneuver. Remote contraction of

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a muscle induces facilitation of both the tendon jerk and the H-reflex (Delwaide and Toulouse, 1981; Miyahara et al., 1996; Zher and Stein, 1999). The temporal profile of the Jendrassik maneuver was accurately studied by Delwaide and Toulouse (1981). These authors divided the facilitation in three different phases, the first one due to a general preparatory state, before any EMG activity could be seen, the second one related to the descending motor command, that would activate not only the muscle in which the voluntary contraction was intended but also other muscles in the body, and the third due to a long-loop reflex generated in the re-afferentation during contraction. According to Zher and Stein (1999), the increase in the size of the H-reflex related to a remote muscle contraction is in part due to removal of presynaptic inhibition. However, Gregory et al. (2001) were unable to find any change in the amount of presynaptic inhibition in heteronymous afferents from the quadriceps muscle. Many studies have examined the physiology of the H-reflex changes preceding a voluntary movement in reaction time experiments. In these experiments, an imperative signal (IS) gives the subject the instruction to begin moving and, in certain paradigms, the IS is preceded by a warning signal that announces the incoming instruction. Some time after the IS, the size of the H-reflex increases progressively until onset of EMG activity in the agonist muscle, and decreases in the antagonist (Michie et al., 1976; Eichenberger and Rüegg, 1983; Schiepati, 1987). Schieppati et al. (1986) showed that the increase of the H-reflex size preceding muscle contraction, or its decrease preceding the release of an ongoing contraction, occurred independently of changes in the amount of EMG activity, confirming that this modulation takes place at a presynaptic level. Furthermore, premovement enhancement of the H-reflex size was not modified by changing kinesthetic aspects of the movement (Collins et al., 1993), and was found to be timelocked to the movement itself rather than to the IS when changing the complexity of the reaction (Hasbroucq et al., 2000). Thus, these experiments pointed out to the fact that the H-reflex enhancement prior to a reaction is related to the motor commands issued for the actual movement execution rather than to any sensory feedback signals. A different behavior has been reported during forewarning (i.e., the time between the warning signal and the IS). During this time, the H-reflex of the agonist muscle undergoes non-specific changes, which occur regardless whether the intended movement is planned for the

J. VALLS-SOLÉ AND G. DEUSCHL

soleus muscle itself or for distant muscles (Brunia and Vuister, 1979). The presentation of the warning signal induces a small increase of the H-reflex, probably related to an arousal effect. In forewarning periods of 1000 ms, Gerilovsky et al. (1983) found that this increase was followed by a decrease, occurring at about 800 ms, for the H-reflex to increase again together with the arrival of the IS. Such a decrease could be due to a presynaptic effect on Ia afferent terminals since the excitability of the motoneuron itself seems not to change. Many other physical activities lead to soleus Hreflex modulation, probably the most relevant ones being postural changes (Koceja et al., 1993; Trimble, 1998; Goulart et al., 2000) and walking (Dietz et al., 1979; Capaday and Stein, 1986; Faist et al., 1996). Koceja et al. (1993) reported a decrease in the H-reflex size in standing compared to sitting, even though the soleus muscle exhibits tonic EMG activity while standing. This is probably the consequence of an increase in the excitability of the soleus motoneurons, with a concomitant increase in the amount of presynaptic inhibition of the Ia afferent terminals (Goulart et al., 2000). The temporal profile of the changes occurring in the soleus and tibialis anterior H-reflex size during the sit-to-stand movement have been studied by Goulart and Valls-Solé (2001). The H-reflex of the tibialis anterior undergoes an early increase preceding the initial phasic burst seen in this muscle at the beginning of the movement. This is followed by a decrease coinciding with the progressive increase of the soleus H-reflex size. At the end of the maneuver, the soleus H-reflex size decreases again, probably due to the intervention of mechanisms of presynaptic inhibition. During walking, different investigators have reported facilitation of the soleus H-reflex size in the stance phase and inhibition in the swing phase (Dietz et al., 1979; Faist et al., 1996; Capaday and Stein, 1986). Not only physical but also mental activities cause a change in the size of the H-reflex. These include mental rehearsal of a movement (Bonnet et al., 1997), alertness (Hodes, 1967) or focussed attention (Honore et al., 1983). 10.2.2.1.7.2. Modulation of the soleus H-reflex by remote external stimuli. External activation of specific descending tracts causes also modulation of the soleus H-reflex in humans. Vestibulo-spinal tract activation with galvanic stimuli caused facilitation of the H-reflex (Kennedy and Inglis, 2001). External activation of the reticulospinal tract by auditory stimuli has

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also been reported to cause enhancement of the soleus H-reflex by several authors, Rossignol and MelvillJones (1976) examined the sound-related changes of the H-reflex size in volunteers who were hopping to the rhythm of a musical instrument. These authors reported the synchronization of hopping with the strong beats of the musical piece, in such a way that the motor events were timed to make best use of the auditory facilitation of the segmental reflex. Delwaide and Schepens (1995) confirmed the finding of Rossignol and Melvill-Jones (1976), and demonstrated that the audiospinal facilitation was abnormal in patients with reticulospinal tract dysfunction (Delwaide et al., 1993). Finally, activation of the corticospinal tract with cortical stimulation causes also modulation of the soleus H-reflex. Using electrical stimuli, Cowan et al. (1986) reported a short latency, short duration phase of facilitation of the H-reflex of various muscles. However, the soleus H-reflex was inhibited rather than facilitated. This was attributed to the fact that corticospinal projections to the soleus muscle are mainly inhibitory. With transcranial magnetic stimulation (TMS), the soleus Hreflex shows two phases of facilitation (Goulart et al., 2000). The first one occurs at intervals between 5 and 30 ms and is assumed to represent the facilitation of the reflex due to the generation of a soleus motoneuronal EPSP. The second one, occurring between 50 and 100 ms, is presently of unknown origin. However, some results may indicate that it may be mediated by subcortical structures and, more specifically, by the reticulospinal tract, probably activated through corticoreticular connections. In cats, the H-reflex is modulated by direct stimulation of the mesencephalic locomotor region (Gosgnach et al., 2000), an observation that is in line with the changes of the H-reflex size during walking reported in humans. Regarding the target of the effects, it is important to notice that vestibular stimulation in cats induced a change in the presynaptic inhibition of Ia afferent terminals (Rudomin and Schmidt, 1999). Electrical or mechanical stimuli applied anywhere in the body are also capable of inducing changes in the size of the soleus H-reflex. Gassel and Ott (1970) showed that cutaneous stimuli of the dorsal and plantar surfaces of the distal foot in humans induced opposite effects on the soleus H-reflex, with short latency facilitation with stimulation of the dorsum, and inhibition with plantar stimulation. These effects were followed by a prominent late increase of excitability with both stimuli. Later, the same authors demonstrated that such late facilitation occurred with widespread

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cutaneous stimulation (Gassel and Ott, 1973). The short latency effects were considered to be a local sign due to the segmental effects induced by activation of type II cutaneous afferents, while the late facilitatory effects were thought to be the consequence of a widespread, multilevel, spinal effect induced by group III fibers activated by the same cutaneous stimuli. The effect would be transmitted through propriospinal circuits up to the brainstem, according to the concept of the spino-bulbo-spinal reflex, proposed by Shimamura and Livingston (1963). These authors proposed that small myelinated fibers would activate such a spinal long-loop reflex by sending inputs centripetally to the reticular formation where they will produce a bilateral descending efferent ventral root discharge. Even though the existence of spino-bulbo-spinal tracts has not been proven so far, the effects of cutaneous nociceptive stimulation over the H-reflex have continued to be shown in different research studies. The electrical stimuli used to elicit the H-reflex unavoidably activate the skin receptors lying below the cathode. Although this nociceptor afferent discharge will not influence the size of the H-reflex elicited by the same stimulus because of a different conduction velocity, it will likely be partly influencing the excitability of the reflex arc to subsequent stimuli. This possibility was examined by Sabbahi and DeLuca (1981), who showed that the excitability recovery of the soleus H-reflex was enhanced when they applied a topical anaesthetic to the skin of the popliteal fossa. The interaction between pain and muscle afferents has been the subject of more recent studies (Le Pera et al., 2001), demonstrating a late inhibitory effect of tonic muscle pain on the H-reflex. 10.2.2.1.7.3. Propriospinal circuits for inhibitory control of motoneuronal firing. Whatever the source of the modulatory inputs, the last step in the central control of H-reflex excitability is made through spinal interneurons with inhibitory projections to the motoneurons (postsynaptic inhibition) or to the Ia afferent terminals themselves (presynaptic inhibition). These circuits which are summarised in Table 10.2, are available to neurophysiological study using the relatively complex techniques developed by PierrotDeseilligny and coworkers (Pierrot-Deseilligny et al., 1981; Pierrot-Deseilligny and Mazières, 1984a,b). Reciprocal inhibition is mediated by Ia inhibitory interneurons. These interneurons receive excitatory inputs from Ia muscle afferents, and project to the motoneurons of the physiologically antagonistic

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Table 10.2 Propriospinal inhibitory mechanisms Mechanism

Interneurons implicated

Neurotransmitter

Tests

Ia inhibition Ib inhibition Renshaw inhibition Presynaptic inhibition

Ia inhibitory interneuron Ib inhibitory interneuron Renshaw cell PAD* interneurons

Glycine Glycine Glycine GABA

Reciprocal Autogenetic Recurrent Presynaptic

* PAD: Primary afferent depolarization

muscle, in which they induce a disynaptic IPSP (Fig. 10.3A). The Ia interneurons impinging on the extensor muscles, and the alpha and gamma motoneurons innervating flexor muscles, are supposed to receive parallel inputs from descending pathways (“alpha-gamma linkage with reciprocal Ia interneuron coactivation”). The electrophysiological evidence for reciprocal inhibition was first obtained in the leg muscles by Mizuno et al. (1971). These authors attempted to activate the Ia inhibitory interneuron projecting to soleus alpha motoneurons by stimulating the large Ia afferent fibers from the pretibial muscles. However, it turned out that the hypothesized disynaptic decrease of the soleus H-reflex amplitude to peroneal nerve stimuli was present only when the subject maintained a voluntary dorsal flexion. In subjects at rest, these authors reported the observation of two H-reflex suppression phases of long latency, suggesting that other mechanisms were at play (Mizuno et al., 1971). Voluntary contraction would have caused an inhibition of central origin because of the alpha-gamma linkage with reciprocal Ia-interneuron coactivation. Therefore, it was believed that the spinal circuit of Ia reciprocal inhibition could not be demonstrated neurophysiologically because the effect was too week. A few years later, however, Day et al. (1984) reported that the circuit in the forearm muscles was different to that of the leg muscles in such a way that the inhibition of the H-reflex of the forearm muscles by a radial nerve shock was indeed present at rest. Interestingly, Day et al. (1984) reported also two long latency phases of H-reflex suppression following the disynaptic phase, similar to those described by Mizuno et al. (1971). Since then, conditioning of the median nerve H-reflex by electrical stimulation of the radial nerve has been considered the paradigmatic test for reciprocal inhibition. The test is very useful for the evaluation of patients with forearm dystonia (Rothwell et al., 1988; Nakashima et al., 1989; Panizza et al., 1989b;

Deuschl et al., 1992). Appropriate methodological recommendations have been put forward by Fuhr and Hallett (1993). Autogenetic Ib inhibition is mediated by Ib inhibitory interneurons. These interneurons receive excitatory inputs from Ib Golgi tendon organ afferents, and project to the homonymous and synergistic motoneurons, in which they induce a disynaptic IPSP (Fig. 10.3B). It is to be noted, however, that the Ib interneuron receives also inputs from the Ia fibers. During contraction, Golgi tendon organs are very sensitive to muscle stretch. When a stretch occurs (for instance, due to a tendon tap), the stretch receptors are activated and induce the monosynaptic reflex, but both the Ia afferents themselves, and the Ib afferents, will activate the Ib interneuron and inhibit the motoneuron with a delay of 1 synapse. The circuit of Ib inhibition is available to electrophysiological testing with the individual at rest, thanks to the fact that the nerve from the gastrocnemius medialis muscle does not carry Ia afferents for soleus motoneurons (Pierrot-Deseilligny et al., 1979). A low intensity electrical stimulus given to the nerve from the gastrocnemius medialis muscle will, therefore, activate the Ib afferents. The subsequent activation of the Ib inhibitory interneuron will cause disynaptic inhibition on the synergistic soleus motoneurons. This takes place in the form of a small decrease of the amplitude of the soleus H-reflex between 4 and 6 ms after the gastrocnemius medialis nerve stimulus. As in the case of the Ia interneuron, the Ib inhibitory interneuron receives many different inputs from descending tracts. It is to note that the dorsal reticulospinal system originating in the nucleus reticularis gigantocellularis (NRGC) sends facilitatory projections to the Ib interneurons (Takakusaki et al., 1994), and inhibitory projections to the Ia interneuron (Delwaide et al., 2000). Dysfunction of the reticulospinal system may be the cause of an abnormal subcortical state of preparation for the execution of

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a motor action in patients with Parkinson’s disease (Delwaide et al., 2000). Recurrent inhibition is mediated by Renshaw interneurons. These interneurons receive excitatory inputs from the first axonal collateral from alpha motoneurons, and project to the homonymous and

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neighboring motoneurons (Renshaw, 1941), in which they induce a disynaptic IPSP (Fig. 10.3C). PierrotDeseilligny and Bussel (1975) devised an electrophysiological method to assess recurrent inhibition with the soleus H-reflex. The method requires a stimulator capable of delivering two stimuli of different

A

B Supraspinal innervation

SOL IA interneurons

P.A.D.

P.A.D. Test

GM α

α α

Cond.

IA afferents IA afferents

1a

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= excitation = inhibition

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C

D

Vibratory Inhibition

H reflex

40 α Mn

ACh

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Gly α Mn

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0 40 120 200 Volts

Motor axon Renshaw’s cell

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F wave 100 μV 10 msec

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Fig. 10.3 Propriospinal inhibitory circuits available to neurophysiological study with the H-reflex. (A) Reciprocal inhibition. This is mediated through the Ia inhibitory interneuron, which is activated by Ia afferents of one muscle and have inhibitory projections to the motoneurons of the antagonist muscle. There is neurophysiological evidence for three distinct phases of reciprocal inhibition, the first one is suggested to be postsynaptic, following a dysynaptic circuit; the second one is suggested to be presynaptic, occurring at a latency of about 10 ms, and the third is suggested to follow a polysynaptic circuit. (B) Autogenetic (homonymous) inhibition. This is mediated through the Ib inhibitory interneuron, which is activated by axons coming from receptors located at the Golgi tendon organs. The Ib inhibitory interneuron has inhibitory projections to the homonymous motoneurons. The only known evidence for the inhibitory Ib effects is a single phase of weak inhibition. (C) Recurrent inhibition. This is mediated by the Renshaw’s cell, which is innervated by the first collateral stemming from the main axon and has inhibitory projections to the parent motoneuron. Evidence for the effects of Renshaw’s inhibition is obtained with techniques using paired shocks and consists in a long lasting inhibition. (D) Presynaptic inhibition. This is mediated by primary afferent depolarization (PAD) neurons acting over the Ia afferent terminals from the same muscle. Evidence for presynaptic inhibitory effects on the homonymous Ia afferents are obtained with the application of vibratory stimuli, while the effects on heteronymous Ia afferents can be objectivated in the facilitatory effects of quadriceps Ia afferents over the soleus H-reflex.

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intensity through the same pair of electrodes. The first stimulus is of low intensity, capable of inducing a small amplitude H-reflex (H1). The second stimulus, of supramaximal intensity and, therefore, activating massively all efferent and afferent fibers, should be delivered 10 to 15 ms after the first. At that moment, there should be an orthodromic volley descending in the motor axons after the activation of a few motoneurons by the first stimulus. However, such volley will not be able to generate any action potential because it will collide with the antidromic volley induced in all motor axons by the second, supramaximal, stimulus. The collision will abolish all action potentials in those motor axons and, therefore, let the specific population of motoneurons activated by the first stimulus free from any inhibition and ready to fire again when the new excitatory input arrives after activation of the Ia afferents by the second stimulus. This will result in the generation of new H reflex, termed H′. However, H′ is always smaller than H1. If the amplitude of H1 is made to increase by increasing the intensity of the conditioning stimulus, there will be first an increase in the amplitude of H′, which will be followed by a progressive decrease when the intensity reaches a certain level. This decrease is due to the effects of Renshaw cell activation on the motoneuron pool involved in the generation of H1. The smaller the H′ with respect to H1, the larger the recurrent inhibition (PierrotDeseilligny and Mazières, 1985). The beauty of such a method lies actually in the fact that a similar chain of events can be considered partly responsible for silent periods and rebounds induced by stimuli applied to mixed nerves when there is traffic of impulses during voluntary contraction (see the paragraph on Silent Period below). Renshaw cells inhibit the homonymous motoneuron, other neighboring motoneurons, and also the Ia inhibitory interneurons projecting to the antagonist muscle (Houk et al., 1970). Descending central nervous system command may cause either facilitation or inhibition of the excitability of Renshaw cells. Presynaptic inhibition is mediated by axo-axonal synapsis in Ia afferent terminals themselves. The interneurons in charge of such inhibition are known as “primary afferent depolarization” neurons. These interneurons receive excitatory inputs from many sources and project to the terminal, causing its depolarization and the consequent depletion of neurotransmitter for new inputs not to be able to go through. As an important difference with respect to the other spinal mechanisms of motor control discussed so far, presynaptic

J. VALLS-SOLÉ AND G. DEUSCHL

inhibition takes place at a presynaptic level (Fig. 10.3D), therefore, leaving the motoneuron uninfluenced. Methods for electrophysiological testing of presynaptic inhibition involve activation of the Ia afferents of the same muscle by means of vibration (Wong et al., 1977; Pierrot-Deseilligny and Mazières, 1985). A slightly more sophisticated technique has also been developed to test presynaptic inhibition (Hultborn et al., 1987). This is based on the heteronymous facilitation induced by quadriceps Ia afferents on the soleus motoneurons. In their experiment, Hultborn et al. (1987) quantified the effects of vibration on the soleus H-reflex as a percent decrease of the size of the response during vibration. Then made a compensation of the size of the H-reflex by increasing the stimulus intensity until the vibrated Hreflex reached the same size as the control H-reflex. The next step was to induce a facilitation of the soleus H-reflex by activating the quadriceps Ia afferents. It was shown that during vibration, heteronymous facilitation decreased, demonstrating that presynaptic inhibition occurs in heteronymous and homonymous inputs alike and to the same extent. Interneurons responsible for presynaptic inhibition receive inputs from descending motor pathways, even though the exact source of these inputs is largely unknown. As described earlier, presynaptic inhibition is an important strategy of the central nervous system for control of reflexes. 10.2.2.2. Methods of clinical interest for the study of peripheral nervous system disorders with the H-reflex The study of the H-reflex is useful in various clinical conditions. As in all other neurophysiological methods, however, the assessment of the H-reflex on its own does not furnish enough information for reaching a specific diagnostic conclusion. The examiner has to bear in mind that only the combination of various tests is able to bring the evidence for localization of the abnormality or pathophysiologic understanding of lesion mechanisms. Of course, the final interpretation of all results has to be made in the context of the clinical neurological evaluation. 10.2.2.2.1. The H-reflex in the assessment of radiculopathies. The study of the H-reflex and the F-wave is to the assessment of radiculopathies as the study of nerve conduction is to the assessment of nerve entrapment. The electrophysiological approach to the diagnosis of radiculopathy should show the functional damage of a root, which is independent from the structural damage shown by radioimaging techniques (Nardin et al., 1999). The study of the H-reflex should

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be undertaken whenever there is suspicion of a S1 radiculopathy or lumbar canal stenosis. It should be taken into account, however, that the H-reflex is equally affected by processes at the root level (preganglionic) and those at the plexus level (postganglionic). Therefore, the topographic location of the lesion requires the confirmation of normal sensory nerve action potentials. The soleus H-reflex is absent or has an abnormally prolonged latency in 100% of patients with S1 radiculopathy (Braddom and Johnson, 1974; Aiello et al., 1981; Sabbahi and Kahlil, 1990), and in only upto 26% of patients with L5 radiculopathy (Schuchmann, 1978; Rico and Jonkman, 1982). In patients with neurogenic claudication, conventional electrophysiological tests may not show positive signs at rest. Pastor and Valls-Solé (1998) studied the behavior of the soleus H-reflex before and after walking for 30 min, showing that exercise caused a transient increase in the H-reflex threshold relative to that of the M-wave in 10 patients with neurogenic claudication. 10.2.2.2.2. The H-reflex in the assessment of polyneuropathies. In all kinds of polyneuropathies, the H-reflex may be absent, suggesting conduction block in, or functional loss of, large afferent axons. The value of the study of late responses (H-reflex and F-wave) in the diagnosis of polyneuropathies has been compared to that of sensory nerve conduction studies. Different types of polyneuropathies may lead to a different type of abnormality in the H-reflex. Hence, a delayed but still present H-reflex might be consistent with predominantly demyelinating polyneuropathy, while absence of response or abnormally high threshold would be consistent with a predominantly axonal polyneuropathy (Guiheneuc and Bathien, 1976; Albers, 1993). The absence of the H-reflex is commonly paralleled with absence of the Achilles tendon jerk and the soleus T-wave. However, there are instances in which the Hreflex may be present and the Achilles tendon lost. This may occur in predominantly distal axonal polyneuropathies in which axons can be damaged only in their most distal part, preventing mechanical activation of the stretch receptors by the mechanical stimulus, but not electrical activation of the fibers at the popliteal fossa. Presence of a weak Achilles tendon with absence of the H-reflex can also occur. This would be the case in some patients with predominant loss of, or reduced excitability in, large axons. If activation threshold is increased in the Ia fibers, there may be no margin for their isolated or predominant activation. The electrical stimulus

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intended to activate the Ia fibers would always depolarize some alpha motor axons, which antidromic volley may prevent the observation of the H-reflex. The Achilles tendon jerk can still be present if enough Ia fibers are left intact for them to convey the mechanical activation of stretch receptors. The typical pattern of an S1 radiculopathy, i.e., absence of the H-reflex with preservation of the sensory nerve action potential, may also be found in polyradiculoneuropathies at the early stages of the disease. Characteristically, in the Guillain–Barré syndrome, the tendon jerks are lost and the H-reflex is absent at a moment in which there may be a complete nerve conduction block at a radicular level, with preserved excitability in distal nerve segments (Lachman et al., 1980). In a series of diseases with involvement of peripheral and central nervous system, the study of the H-reflex might be helpful for understanding the underlying pathophysiological mechanisms. This is the case, for instance, in spino-cerebellar disorders. In Friedreich’s ataxia, the involvement of dorsal root ganglia should cause the sensory nerve action potential amplitude to decrease and the H-reflex to be absent (Hugues et al., 1968). However, some patients with early onset cerebellar ataxia may have completely normal reflexes even though they show a noticeable sensory loss and absence of response in sensory nerve conduction studies (Harding, 1981). This clinical condition, known as Friedreich’s ataxia with retained reflexes (FARR) has a slightly better prognosis than the classical form. 10.2.2.2.3. The H-reflex in the assessment of diseases modifying the excitability of the motoneurons. The Hreflex is usually of normal latency in patients with motoneuron disease. However, its excitability might be enhanced. When in doubt, the observation of a Hreflex in muscles in which it is not normally observed unless during facilitation by voluntary contraction, may help to rule out chronic polyneuropathy and favor the diagnosis of motoneuron disease. The H-reflex is also useful in the study of the stiff-man syndrome. In this syndrome, patients have painful spasms induced by involuntary activation of spinal motoneurons. Abnormalities in the inhibitory control of alpha motoneurons are supposed to underlie the pathophysiology of this syndrome. In a group of 11 patients with stiff-man syndrome, Floeter et al. (1998a) have consistently found an abnormal decrease in vibratory inhibition of the H-reflex, and more scattered abnormalities in other propriospinal inhibitory circuits.

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10.3. Oligo- and poly-synaptic reflexes elicitable at rest Any electrical stimulus applied over a nerve trunk gives usually rise to a large afferent volley. The effects of that volley in the central nervous system may not be apparent in the majority of nerves when the muscle is at rest. Noteworthy exceptions in healthy human subjects are the orbicularis oculi reflex, the nociceptive withdrawal reflexes, and a few others. Most of these reflexes are known as exteroceptive reflexes. 10.3.1. The blink reflex 10.3.1.1. Physiological and technical considerations The blink reflex consists on a stimulus-triggered eyelid closure involving mainly activation of the orbicularis oculi muscle and relaxation of the levator palpebral muscle. The most commonly used stimulus in clinical practice is an electrical stimulus to the supraorbital nerve (Kimura et al., 1969; Shahani, 1970). An adequate stimulus duration is 0.2 ms, whereas the stimulus intensity is, usually, 2 to 4 times perception threshold. The reflex response of the orbicularis oculi to supraorbital nerve electrical stimulation consists of two separate components: an early ipsilateral R1; and a later bilateral R2 response. R1 is a pontine reflex, while R2 is presumably relayed through a more complex route including the pons and lateral medulla (Kimura and Lyon, 1972; Ongerboer de Visser and Kuypers, 1978). A mechanical tap over the glabella also elicits a blink reflex. Although the stimulus is a gentle tap, this is a cutaneous rather than a stretch reflex, probably relayed via the same polysynaptic reflex pathways as the electrically elicited blink reflex (Shahani, 1970). Response recording is done with surface electrodes. The active recording electrode is placed on the lower aspect of the orbicularis oculi muscle and the reference electrode 2–3 cm lateral. Careful placement of the electrodes may help reduce the stimulus artifact that may sometimes make it difficult to determine the exact onset latency of R1. A low-frequency cutoff of 200 Hz or more may also be helpful. Although it is recommendable that reference values are obtained at each laboratory for their own comparison to patients, normative values have been published by several authors (see, for instance, Kimura, 2001). The upper limit for R1 latency is 13.0 ms, and the latency difference between the two sides in the same subject must be less than 1.2 ms. The upper limit of normal latency for the R2 is 40 ms on the side of stimulation and 41 ms on the contralateral side, and the latency

J. VALLS-SOLÉ AND G. DEUSCHL

difference between the ipsilateral and the contralateral R2 evoked simultaneously by stimulation on one side should not exceed 5 ms. 10.3.1.2. Methods of clinical interest for the study of peripheral nerve disorders with the blink reflex The R1 response is relatively stable with repeated trials and is therefore better suited for assessing nerve conduction through the trigeminal and facial nerves. Analysis of R2, however, is essential in determining whether a lesion involves the afferent or efferent arc of the reflex. With a lesion of the trigeminal nerve, R2 is slowed or diminished bilaterally when the affected side of the face is stimulated (afferent delay), while stimulation of the unimpaired nerve gives rise to normal responses. With a lesion of the facial nerve, R2 is abnormal on the affected side (efferent delay), while the responses are normal in the unaffected side, regardless of the side of stimulation. 10.3.2. Withdrawal reflexes to nociceptive inputs 10.3.2.1. Physiological and technical considerations Fibers of small diameter are not normally accessible to direct electrophysiological tests using conventional methods. The most suitable methods for the study of small fibers are described in Table 10.3. A short train of electrical impulses applied to the sole of the foot, or to the posterior tibial or sural nerves at the ankle causes a reflex response leading to the withdrawal of the leg from the painful stimulus (nociceptive reflex). The response recorded in biceps femoris and tibialis anterior is made of two components, the RII and the RIII. The RII component is usually obtained at a relatively low, nonpainful, stimulus intensity. The RIII is obtained at a high intensity stimulation and is recruited in parallel with an increasingly painful sensation. When recording in the short head of the biceps femoris, the first muscle to be activated after a painful stimulation of the foot, response latency is 50–70 ms for the RII, and 80–130 ms for the RIII (Willer, 1983). The RII response is probably due to activation of large group A afferents, while the RIII response should be due to activation of Aδ and C fibers. The RIII response is largely inconsistent, and poorly comparable among individuals. Withdrawal reflex responses have been also recognized in the upper limb to noxious cutaneous stimuli (Floeter et al., 1998b). Reflex responses to nociceptive stimuli can also be generated in pelvic floor and facial muscles. The bulbocavernosus reflex (Vodusek et al., 1983) is usually obtained by applying electrical stimuli to the dorsal

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Table 10.3 Methods for neurophysiological evaluation of activity in pain and small fiber systems

Sensory pathways Standardized sensory testing Vibrometer Thermotest Algometer Laser stimulation

Touch and pain sensations Vibratory sensation Warm, cold, heat nociception and cold nociception sensations Pressure and pressure nociception sensation Pain sensation

Reflex responses Painful electrical stimulation Sudomotor function Blood pressure Heart rate

Reflex withdrawal motor responses Sudomotor skin response. Postganglionic axonal reflexes Baroreflex functions Cardiovagal functions

nerve of the penis or clitoris. Recording can be done through needle electrodes inserted in the bulbocavernosus muscle or in the anal or vesical sphincter muscles. It is also possible to record the response from the anal sphincter using surface electrodes. Reflex responses of facial muscles are obtained with relatively high intensity electrical stimuli to the median or tibial nerves. The clinical correlate of these responses is the palmomental reflex (Dehen et al., 1975). The reflex circuit of the facial responses to peripheral nerve stimulation is not known, but it may involve transcortical pathways. 10.3.3.2. Methods of clinical interest for the study of peripheral nerve disorders with nociceptive reflexes. The RIII response has been utilized in studies on efficacy of analgesic drugs, but it is not applied to clinical evaluation of patients. Nevertheless, the withdrawal reflexes to nociceptive stimuli may be a valuable tool to examine the function of small fibers. The bulbocavernosus reflex is abnormally large and has a shorter latency in patients with neurogenic bladder due to upper motor neuron disease, and is absent or has a prolonged latency in patients with cauda equina or conus medullaris lesions (Ertekin and Reel, 1976). The analysis of facial responses to peripheral nerve stimuli may help in the assessment of several peripheral and central disorders (Miwa et al., 1996; Valls-Solé et al., 1997). 10.4. Silent period 10.4.1. Physiological and technical considerations The silent period is defined as the transient absence or significant decrement of EMG activity during sustained voluntary muscle contraction. It may be due to actions taking place at the motoneuron cell body itself

or at the motor axons. An example of a silent period taking place initially at the motoneurons is the inhibition induced by postsynaptic inputs, for instance with external activation of descending tracts using transcranial magnetic stimulation (TMS). An example of a silent period due to effects taking place initially at the level of the motor axons is the inhibition caused by electrical stimulation of peripheral nerves. In both conditions, the silent period may be either the actual consequence of an inhibitory input, or the aftereffect of an excitatory input. These two mechanisms lead to a different type of silent period (Ashby, 1995). When it is the consequence of an inhibitory input, the silent period occurs as an initial event, without any facilitatory phase preceeding the decrement of EMG activity. When it is the after effect of an excitatory input, the silent period occurs after an initial EMG burst. 10.4.1.1. The silent period due to actions taking place at the motoneuron cell body Inputs of various sources are able to evoke direct effects on alpha motoneurons. One example of those is TMS, which has actually brought renewed interest for the events taking place during the silent period (Haug et al., 1992). Induction of a silent period with TMS involves discharging the magnetic coil on the scalp over the motor cortex area at a variable intensity during sustained muscle contraction. Low-intensity TMS is able to induce a silent period in hand muscles without preceding motor evoked potential, possibly due to activation of cortico-cortical inhibitory connections (Wassermann et al., 1993). With high intensity TMS, the silent period divides into two parts (Fuhr et al., 1991): the initial part attributable to effects of the descending volley on the spinal interneuronal and motoneuronal machinery, and the final part, due to cortical inhibition. The silent period

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after TMS can be induced in many muscles, including those ipsilateral to the side of stimulation (Wassermann et al., 1991), which physiological particularities are of great interest in the evaluation of interhemispheric inhibitory connections. Once they have been activated, alpha motoneurons are more difficult to reach firing threshold again. This is due to after-hyperpolarization (AHP), a condition that takes place in part because of potassium accumulation in the outer membrane space. The AHP is a means to regulate the rate of repetitive firing of individual neurons receiving a constant excitatory input. This is for instance what determines the firing frequency of alpha motoneurons that are continuously bombarded by inputs from a high frequency depolarizing current. The AHP is also the main feature underlying the EMG silent period occurring after synchronous firing of motoneurons. The length of the AHP may also be modulated by the central nervous system. For instance, this has been the case for shortening of the AHP in rats after stimulation of the raphe-spinal serotoninergic projections (Bayliss et al., 1995). If different motoneurons are activated according to their centrally driven random firing mode, their respective action potentials, and the corresponding AHP that follows, will occur at random intervals from each other. However, the addition of an extra facilitatory or inhibitory impulse reaching the motoneuronal membrane will be capable of synchronizing either the firing or the silence of some motoneurons. Synchronization will be followed by an inevitable series of events. With facilitatory inputs, the synchronous firing will cause an action potential that will be inevitably followed by a silent period because of the synchronization of the AHP period. With inhibitory inputs, the synchronous absence of firing will cause a silent period that will be inevitably followed by a facilitatory phase due to more motoneurons being ready to fire. If the technique of EMG modulation is used, any synchronizing event, like, for instance, the posterior tibial nerve eliciting the H-reflex, will induce a series of facilitatory and inhibitory phases of EMG activity, in a kind of oscillatory pattern (Fig. 10.4). In natural circumstances, the motoneurons receive a continuous bombardment of excitatory and inhibitory inputs. Usually, the effects of the inputs reaching the motoneuron during the AHP period are not evident, except if they do so when the AHP is approaching its end. Their effects can be demonstrated with the construction of the peri-stimulus time histogram of single motor units, recorded with needle electrodes during a weak contraction. If the action potential of a single motor unit is made to fire regularly at a stable fre-

J. VALLS-SOLÉ AND G. DEUSCHL

50 ms 0.2 mV

Fig. 10.4 Modulation of the soleus EMG activity by stimuli inducing the H-reflex. The EMG activity was rectified and 100 epochs of 500 ms duration averaged. The stimulus to the posterior tibial nerve is represented by the thin vertical line and the background level of EMG activity is shown by the double head arrow at the beginning of the trace. Note the small M wave just after the stimulus, the large H-reflex, and the series of clonuslike oscillations induced in the ongoing EMG activity.

quency, and a stimulus is applied at random intervals with respect to the motor unit action potential, the effects observed illustrate the phenomenon of AHP and its relationship with the silent period. If the stimulus finds the motoneuronal membrane excitability rising from the depth of the AHP, it will make that motoneuron fire again but, this time, at a shorter latency (i.e., will be recruited by the stimulus). On the contrary, if the stimulus finds the motoneuronal membrane excitability decreased, the motoneuron will not be activated, and will remain silent until another input becomes effective. The motor units that have been brought to fire will then remain silent for the whole length of the AHP period, and the ones that have not been recruited by the stimulus will fire at their normal pace. As a consequence, there will be a time window in which there will be no motor unit action potentials, the silent period. This will span between the action potentials generated by the motoneurons recruited and those not recruited by the stimulus. With a fixed intensity of the stimulus eliciting the events, the length of the silent period depends on (1) the mean interspike interval of the motor units that were active during contraction; and (2) the proportion of these units that were brought to threshold. 10.4.1.2. The silent period due to actions taking place in the motor axons A different picture arises when the peripheral nerve is activated directly with an external source of depolarization, i.e., when an electrical stimulus is applied to the nerve. This is actually the easiest method to elicit a silent period in routine clinical practice. Several peripheral nerves are accessible to supramaximal electrical stimulation. The stimulus used to induce the peripheral nerve silent period is usually the same as that used to induce a supramaximal compound muscle

REFLEX RESPONSES, SILENT PERIOD AND LONG LATENCY REFLEXES

action potential in the supplied muscle. The time window required to evaluate the whole length of the silent period should be at least of 200 ms. Maintaining a voluntary contraction involves random activation of motoneurons and their axons. The depolarizing volley induced by the external supramaximal activation of motor axons will travel orthodromically to induce the synchronized M-response, and antidromically to induce effects at a motoneuronal level. The first antidromic event will be the collision with the motor axons that happened to be activated at that moment. Note that those axons will be clear of any antidromic or orthodromic impulse for a certain time, which explains the first part of the silent period after the M-wave. In the axons in which the antidromic volley does not find any ongoing activity, the antidromic excitatory volley will reach the motoneuron, where it may induce a reverberating potential to travel again orthodromically and give rise to the F-wave. At the same time, however, the antidromic volley will reach the first axonal collateral and activate the Renshaw’s cell. This will produce inhibition of the homonymous and neighboring motoneurons (Renshaw, 1941), causing a long lasting silent period. In the mean time, descending excitatory inputs might have reached the motoneurons that were freed of any impulse because of the collision between orthodromic and antidromic volleys. These will not have recurrent inhibition on their own, and a number of them would have not undergone the inhibition induced by Renshaw cells in neighboring motoneurons. Therefore, they will give rise to a burst of action potentials, appearing in the middle of the silent period. This burst is known as the voluntary potential (Kimura, 1977), because it grows with the degree of contraction. It also grows with the length of the nerve supplying the muscle under study because of the higher probability of collision. Electrical stimuli to peripheral nerves cannot selectively activate motor axons unless specific preparations are used (Gandevia et al., 1984). Therefore, activation of sensory fibers will also contribute to the last part of the silent period by way of inhibitory inputs impinging on motoneurons. Figure 10.5 shows an example of the silent period elicited in thenar muscles to median nerve stimuli. When the stimulus is applied to purely sensory nerves, the effects on spinal motoneurons will vary depending on various factors such as the nerve stimulated, the muscle recording from, and the action being performed (Uncini et al., 1991). The silent period is of longer latency than the one induced with stimulation of a mixed nerve, and it begins with inhibition. Therefore,

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Silent period

S 20 ms F

V

0.2 mV

S

Fig. 10.5 The silent period induced in thenar muscles by median nerve stimuli. The EMG activity has been rectified and 20 consecutive trials have been averaged. Two traces recorded with the same methodology are superimposed to see consistency of the results. The stimuli are given at the arrow and marked with ‘S’. Two different degrees of intensity of contraction were used, mild for the upper traces and strong for the lower traces. Note the two bursts interrupting the silent period. The first, labelled F, corresponds to a potential generated in the spinal cord at a segmental level. The second, labelled V, corresponds to the ‘voluntary’ potential (see text). Note the increasing size of the V potential with the intensity of contraction.

it is likely due to motoneuronal IPSPs. These may be caused by inputs from inhibitory interneurons, possibly Renshaw’s cells, activated by mildly myelinated type II fibers. The depth and duration of the silent period induced by sensory nerves is directly proportional to the intensity of the stimulus and inversely proportional to the strength of muscle contraction (Caccia et al., 1973; Uncini et al., 1991). After the silent period, there is a rebound of EMG activity, which is the consequence of membrane excitability recovery taking place after the IPSP. The silent period to cutaneous inputs can be interpreted as part of a complex long latency reflex response induced by the sensory stimulus and, as such, its physiological particularities are going to be discussed together with those of the long latency reflexes. 10.4.2. Methods of clinical interest for the evaluation of peripheral nervous system disorders with the silent period 10.4.2.1. The silent period of extremity muscles The silent period has not much use for the evaluation of peripheral nervous system disorders (Ford et al.,

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1995). Possibly the most known method for inducing a silent period is the application of an electrical stimulus over the median nerve at the wrist, while the subject is maintaining a weak muscle contraction of hand muscles by holding an object between thumb and index. Like other neurophysiological events reflecting conduction velocity, the latency of the silent period is going to be delayed in polyneuropathies implying a slow conduction velocity of afferent and efferent fibers. However, since at least part of the silent period is due to the action at the central nervous system, the peripheral delay may be counteracted by the normal central nervous system conduction and, therefore, will be milder in comparison to the results of peripheral nerve conduction studies. Another area in which the silent period may be of interest is in the assessment of diseases causing changes in the inhibitory control of motoneurons (Ford et al., 1995). This is the case with the tetanus infection (Fernández et al., 1988; Poncelet, 2000), in which neurotransmitter mediated inhibition of motoneurons is impaired and, therefore, inhibitory inputs carried by sensory afferents after peripheral nerve stimulation cannot generate IPSPs. The initial part of the silent period elicited by peripheral nerve stimuli that is due to the electronic collision between orthodromic and antidromic volleys, is usually preserved. 10.4.2.2. The silent period of the masseter muscles (masseter inhibitory reflex) Sustained voluntary activity of the masseter muscles is transiently interrupted after stimuli of various kinds. The most common method of examining the masseter inhibitory reflex (MIR) is an electrical stimulus applied to the mentalis nerve at its exit from the mandibular bone at the chin while subjects are asked to clench their teeth. This produces two phases of suppression of EMG activity (exteroceptive suppression [ES], or silent period [SP]). The early phase is called MIR1, ES1, or SP1, and the second phase is called MIR2, ES2, or SP2 (Cruccu and Ongerboer de Visser, 1999). Some subjects with bad dentures are not able to activate their masseter muscles and use the facial muscles instead. In these cases, no suppression of the EMG activity can be elicited either with a tap to the chin or with an electrical stimulus, because there is no silent period in most facial muscles. This should obviously not be mistaken for an abnormal MIR. Quantitative studies of the excitability of the brainstem inhibitory interneurons require measurement of response size (e.g., area of suppression): the level of

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background EMG activity must be kept constant. The signals must either be full-wave rectified and averaged, or examined by superimposing several trials. As with the blink reflex, the MIR provides information on the site of the lesion by analyzing whether the pattern of abnormality is afferent, mixed, or efferent. The efferent pattern of lesion is extremely rare except in conditions such as a purely motor trigeminal neuropathy and hemimasticatory spasm (Cruccu and Deuschl, 2000). ES1 is delayed in patients with demyelinating polyneuropathies and also in patients with severe diabetic polyneuropathy (Cruccu and Deuschl, 2000). ES2 is far less sensitive than ES1 to lesions along the reflex arc. Being mediated by a multisynaptic chain of interneurons, however, it is modulated by suprasegmental influences. 10.5. Long-latency reflexes as a result of modulation of sustained EMG activity Some long latency reflex responses are very small and only elicitable during muscle contraction. The elicitation of long latency reflexes during a sustained contraction allows for the recording of both, excitatory and inhibitory events, which would appear as an increase or a decrease of the amount of EMG activity with respect to the background level of activity recorded during the same muscle contraction without the interference of the stimuli. Therefore, superimposition or averaging of several traces may be required to demonstrate and quantify excitatory and inhibitory reflex responses. The technique of performing this type of examination using an electronically rectified electromyographic recording is sometimes referred as to the “modulation of electromyographic activity”. Using this method, facilitatory and inhibitory reflex phenomena can be quantified with respect to the level of background activity. 10.5.1. Long-latency reflexes of hand muscles Long-latency reflexes in extremity muscles can be elicited with various natural stimuli like muscle stretch (Rothwell, 1990) sudden leg displacements when standing (Dietz, 1993), or cutaneous airpuffstimulation (Deuschl et al., 1995). They consist of a series of reflexes depending on the eliciting stimulus. Their physiological function is to adapt movement in the stimulated extremity according to ongoing sudden changes of the external conditions. Methods to

REFLEX RESPONSES, SILENT PERIOD AND LONG LATENCY REFLEXES

reliably obtain the long latency reflexes of hand muscles without sophisticated equipment have been well standardized (Cruccu and Deuschl, 2000). These involve median nerve or radial superficial nerve stimulation while recording from thenar muscles during maintenance of a contraction of the opponens pollicis done by leaning the thumb at the fifth finger (Fig. 10.6). The reflex responses recorded in this way are termed long-latency reflex I-III (LLR I-III), following median nerve stimulation, and cLLR I-III following radial superficial nerve stimulation (Deuschl and Lücking, 1990). Median nerve stimulation at the wrist, at an intensity just below motor threshold, gives rise to a H-reflex, at a latency of 25–34 ms. This is followed by the LLR I, in about 30% of the normal subjects, the LLR II, in 100% of normal subjects, and the LLR III, in about 20% of normal subjects. The latency of these reflexes depends on the height of the subject. Table 10.4 shows the normal values on latency, amplitude and duration of the hand LLRs. Hand muscle reflexes are mainly used to diagnose alterations within the central nervous system like in myoclonus, multiple sclerosis, parkinsonian syndromes and choreatic syndromes (Deuschl et al., 1988, 1989; Cruccu and Deuschl, 2000). In case of peripheral nerve lesions, delayed motor conduction can be identified on the basis of prolonged nerve conduction leading to a delay of the whole reflex pattern. Desynchronization of the afferent volley, however, A

Primary motor cortex

Primary sensory cortex

255

leads to absence of the reflex pattern. In some cases it may be of interest to see proximal changes of the peripheral nerve conduction that can be assessed with LLR-testing. 10.5.2. Other cutaneo-muscular reflexes Cutaneo-muscular reflexes elicited by stimulation of digital sensory nerves are less standardized and more difficult to obtain (Caccia et al., 1973; Jenner and Stephens, 1982). The subject should wear the surface recording electrodes attached over the 1st interosseous or thenar muscles, while electrical stimuli are given to the digital nerves of the index finger by way of ring electrodes or to any other cutaneous nerve of the hand (Deuschl and Lücking, 1990). The subject is asked to maintain a steady background contraction of the muscle under examination. The EMG activity is rectified and several epochs of, for instance, 500 ms, should be averaged time locked to the electrical stimulus. In this way, the amount of EMG activity will be modulated according to the excitatory or inhibitory effects of the stimulus. Excitatory effects will induce a more synchronous firing of motoneurons, and more motoneurons to fire at a certain time, increasing the size of the averaged EMG activity. Inhibitory effects will cause the opposite. Excitatory effects are labelled E waves, and inhibitory effects are labelled I waves (Fig. 10.7). The E and I waves are numbered B H-reflex I LLR II

LLR – II pathway following muscle and cutaneous stimulation

II LLR III Thalamus

III 0

Nucleus cuneatus I

100

200 (ms)

cLLR II 100 μV

skin

H-reflex pathway

Spinal cord

II cLLR III III 0

100

200 (ms)

muscle

Fig. 10.6 (A) Reflex pathways of the hand muscle reflexes. (B) Normal patterns of reflexes elicited by stimulation of the median nerve (upper set of graphs) and radial superficial nerve (lower set of graphs), in three normal subjects (I-III) (Adapted from Cruccu and Deuschl, 2000 with permission from International Federation of Clinical Neurophysiology.).

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Table 10.4 Normal values of the hand muscle reflexes following median nerve stimulation (HR and LLR) or radial superficial nerve stimulation (cLLR) in 102 normal subjects (age, 18–85 years) Mean

SD

SE

Count

Minimum

Maximum

28.9 40.6 50.3 76.0 37.6 50.2 75.9

2.4 2.5 3.2 4.6 2.6 3.0 3.6

0.2 0.5 0.3 1.0 1.0 0.3 0.7

102 26 102 20 7 100 31

24.1 36.8 43.1 70.3 35.0 43.0 70.0

35.4 47.2 59.3 92.1 43.0 60.0 82.0

B: Amplitude HR LLR I LLR II LLR III cLLR I cLLR II cLLR III

1.9 0.4 1.2 0.8 0.4 1.2 0.8

1.1 0.1 0.6 0.5 0.2 0.5 0.5

0.1 0.0 0.1 0.1 0.1 0.0 0.1

102 25 102 20 7 100 31

0.3 0.2 0.3 0.3 0.1 0.3 0.3

4.8 0.8 3.0 2.3 0.6 2.5 2.4

C: Duration HR LLR I LLR II LLR III cLLR I cLLR II cLLR III

10.4 8.9 22.3 27.6 10.0 24.0 27.7

2.5 7.8 5.4 8.7 3.7 6.6 6.2

0.2 1.5 0.5 2.0 1.4 0.7 1.1

102 25 101 20 7 100 31

6.0 3.0 12.0 9.0 7.0 9.0 15.0

17.0 46.0 40.0 44.0 17.0 50.0 40.0

A: Latency HR LLR I LLR II LLR III cLLR I cLLR II cLLR III

according to their order of appearance (E1, I1, E2, etc.). Some studies have also addressed foot muscle reflexes (Claus and Jakob, 1986). Meinck et al. (1983) reported on the utility of the cutaneo-muscular reflexes in the assessment of disor-

ders causing changes in the excitability of motoneurons. Abnormalities of leg cutaneo-muscular reflexes have been reported in four patients with the stiff-leg syndrome (Brown et al., 1997), in whom cutaneous electrical stimuli induced long-lasting spasms.

Fig. 10.7 The cutaneo-muscular reflex elicited in the 1st interosseous to electrical stimulation of the index finger. Two traces are superimposed to see the consistency of the result. The subject was maintaining the index finger straight against the weight of a small book. Electrical stimuli were applied with ring electrodes to the index finger.

20 ms E2

0.2 mV

E1 L

I1 S

I2

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Valls-Solé, J, Valldeoriola, F, Tolosa, E and Martí, MJ (1997) Distinctive abnormalities of facial reflexes in patients with progressive supranuclear palsy. Brain, 120: 1877–1883. Valls-Solé, J, Hallett, M and Brasil-Neto, JP (1998) Modulation of vastus medialis motoneuronal excitability by sciatic nerve afferents. Muscle Nerve, 21: 936–939. Vodus˘ek, DB, Janko, M and Lokar, J (1983) Direct and reflex responses in perineal muscles on electrical stimulation. J. Neurol. Neurosurg. Psychiatry, 46: 67–71. Wassermann, EM, Fuhr, P, Cohen, LG and Hallett, M (1991) Effects of transcranial magnetic stimulation on ipsilateral muscles. Neurology, 41: 1795–1799. Wassermann, EM, Pascual-Leone, A, Valls-Solé, J, Toro, C, Cohen, LG and Hallett, M (1993) Topography of the inhibitory and excitatory responses to transcranial magnetic stimulation in a hand muscle. Electroenceph. Clin. Neurophysiol., 89: 424–433. Willer, JC (1983) Nociceptive flexion reflexes as a tool for pain research in man. In: JE Desmedt (Ed.), Motor Control Mechanisms in Health and Disease. Raven Press, New York, pp. 809–827. Wong, PKH, Verriere, M and Ashby, P (1977) The effect of vibration on the F-wave in normal man. Electromyogr. Clin. Neurophysiol., 17: 319–329. Zher, EP and Stein, RB (1999) Interaction of the Jendrassik maneuver with segmental presynaptic inhibition. Exp. Brain Res., 124: 474–480.

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CHAPTER 11

Nerve root stimulation Daniel L. Menkes* University of Tennessee Health Sciences Center at Memphis, TN, USA

11.1. Introduction Root stimulation (RS) techniques are capable of activating the proximal portions of the motor and sensory nerve roots resulting in nerve action potentials. While sensory nerve action potentials (SNAPs) are usually recorded directly, motor nerve potentials are usually recorded in the form of their induced compound muscle action potentials (CMAPs). All nerve action potentials are composed of subcomponent waveforms that demonstrate some degree of temporal dispersion (TD) over increasing distances. Given the lower amplitude of the sensory nerve action potentials, normal degrees of physiological TD limit the distance over which sensory nerves can be conducted and recorded, necessitating the use of somatosensory evoked potential techniques (Menkes, 2002a). In contrast, compound muscle action potentials are three orders of magnitude greater than their SNAP counterparts and can evaluate motor nerve conduction studies (NCS) over greater distances despite the effects of normal physiological TD. However, motor NCS in the upper extremity are infrequently performed proximal to the arm and the lower extremity is infrequently conducted proximal to the knee. Depending on an individual’s limb length, these techniques may exclude between 20 and 50% of the motor nerve from analysis. Motor NCS in these segments are more technically difficult to perform because of the distance of the excitable nerve fibers from the skin surface where conventional surface electrical stimulation is usually applied. Since the electric field is generated between the cathode and anode, physical separation of the anode and cathode will enlarge the electrical field and depolarize all excitable tissue within that region if * Correspondence to: Daniel L. Menkes, MD, Department of Neurology, University of Tennessee Health Sciences Center at Memphis, 855 Monroe Avenue, Link Building, Room 415, Memphis, TN 38163, USA. E-mail address: [email protected] Tel: +1-901-448-6199; fax: +1-901-448-7440.

the stimulus is of sufficient intensity. RS methodologies enlarge the size and current density of the electrical field so that the proximal portions of the motor nerve roots are stimulated. Therefore, RS methods provide a means by which motor NCS may be performed from the motor nerve roots to the target muscle. Similar to conventional motor NCS, pathological conditions may manifest as either latency prolongation or changes in waveform morphology such as non-physiological TD or proximal conduction block (PCB). Nerve conduction velocity calculations are less useful as the exact distance traversed is difficult to measure accurately. While F-waves and H-reflexes provide some assessment of these proximal segments, they only evaluate a small proportion of the largest and fastest conducting fibers evaluated by NCS. If only one large motor fiber capable of conducting an F-wave is unaffected by the underlying pathology, then a normal F-wave minimum latency (FWML) may be recorded. Conversely, significant degrees of either myelin dysfunction or axon loss may result in absent F-waves. Thus, the observation of absent F-waves associated with a normal distal CMAP is not pathognomonic for acquired demyelination. The same analogy may be applied to H-reflexes wherein a normal H-reflex latency may be noted when one large sensory fiber, the interneuron and one large motor fiber are unaffected by the proximal pathology. Similarly, an absent H-reflex may be noted in a variety of conditions other than proximal demyelination. For example, one could consider a patient with multifocal motor neuropathy with conduction block in the tibial nerve 5 cm distal to the popliteal fossa. The tibial late responses, F-waves or H-reflexes, may or may not be prolonged to the degree necessary to diagnose acquired demyelination with confidence. However, stimulation of the tibial nerve at the popliteal fossa that demonstrated non-physiological TD or PCB would confirm myelin dysfunction as the underlying pathological mechanism. Had the pathology been located in the thigh instead of distal to the knee, then motor NCS would not demonstrate either of these findings as the stimulation

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site is distal to the affected site. In this situation, stimulation of the tibial nerve proximal to the pathologic site could demonstrate these findings. RS is the method by which proximal stimulation is accomplished. However, RS techniques should be viewed as complementary tests to late response testing. The correlation between F-wave minimum latencies prolonged into the demyelinating range and RS abnormalities was reported in one case series of 31 patients where demyelinating abnormalities were noted in a total of 77 motor nerves (Menkes et al., 1998). F-wave minimum latency prolongation associated with a PCB was noted in 11 nerves, prolonged F-waves alone in 24 nerves and PCB alone in 42 nerves. Therefore, failure to have performed RS studies would have missed PCBs in over 50% of the affected nerves. Therefore, RS should be performed in cases of suspected proximal demyelination even when the FWML is not prolonged into the “demyelinating range.” However, RS need not be performed when sufficient F-wave and other neurodiagnostic abnormalities that meet established criteria for demyelination are observed. In summary, RS techniques extend the range of traditional motor NCS. This chapter will review RS methods, discuss the evidence for site of stimulation, discuss the merits and drawbacks of these techniques and discuss the clinical utility of this methodology. 11.2. Overview of root stimulation methods Although RS techniques activate the motor nerve roots, they have many similarities to conventional motor NCS. Both techniques rely upon supramaximal stimulation of the largest, most heavily myelinated subpopulations of the motor nerves. Strict adherence to proper technique with regard to stimulation and recording parameters to insure reproducibility is essential. As with any normative data, each laboratory ought to obtain its own normal controls from a representative sample of the general population. Alternatively, the technique must be performed in exactly the same manner as published in the literature. In each case, stimulation must be performed at least twice in order to demonstrate waveform presence and reproducibility. With the exception of thoracic and upper lumbar magnetic nerve root stimulation (MNRS) techniques, all RS methodologies are highly reproducible. Various techniques and their normal values have been summarized in another text along with illustrations of these techniques (Oh, 1993). Normal values used in the author’s laboratory are summarized in the appended tables.

DANIEL L. MENKES

It cannot be overemphasized that root stimulation techniques activate these structures distal to the axon hillock. Therefore, RS techniques do not evaluate the entire length of the motor axons in the peripheral nervous system. This may be one explanation as to why there are cases of acquired demyelinating neuropathies that do not manifest PCB. There are two main methods in root stimulation studies: electrical and magnetic. Electrical nerve root stimulation (ENRS) utilizes an invasive method with monopolar needles wherein the electrical field distribution is widened by physically separating the cathode and the anode. In contrast, magnetic nerve root stimulation (MNRS) is a painless, noninvasive method wherein a magnetic field is used to induce an electrical field within the proximal portions of the motor nerves. Since MNRS is noninvasive, it can be undertaken on people with a bleeding diathesis or on those who require anticoagulation. Its main limitation is that most magnetic stimulators produce a lower electrical field density than is generated by ENRS techniques. A complete discussion of magnetic stimulation physiology is beyond the scope of this chapter but is discussed elsewhere (Chokroverty, 1990). The main reason is that invasive ENRS insures that the electrical field is generated in close proximity to the motor nerve roots whereas the magnetic stimulator induces an electrical field at a greater distance. Since the electrical field strength decreases as an inverse square of the distance, MNRS techniques are more susceptible to small changes in distance. One main criticism of RS techniques is that it is highly technique-dependent, requiring an additional degree of expertise; analogous to the differences between routine needle EMG and single fiber EMG, (SFEMG). While it is true that SFEMG and RS require an advanced skill set, the information that may be obtained from RS increases the sensitivity of diagnosing pathology in the proximal portions of the motor nerves. Since small changes in distance will result in a significant change in electrical field density, strict adherence to technique and avoidance of common pitfalls is essential. MNRS techniques are even more prone to small changes in magnetic stimulator positioning because of the greater distance from the targeted excitable nerve tissue. Unlike electrical fields, the energy in a magnetic field is not conserved so all of the energy is not transferred into the induced electrical field. Moreover, the greatest magnetic field intensity is generated at the skin surface, resulting in a smaller electrical field density at the motor nerve root. This effect is greatly pronounced in persons of large girth that results in an even greater distance between the skin and the target site.

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RS techniques also impose strict interpretation guidelines to insure specificity at the expense of sensitivity. Since suboptimal techniques will have a greater impact on proximal conduction studies, profound changes in CMAP morphology must be observed in order to diagnose proximal motor nerve pathology with confidence. As in more distal segments of the peripheral nervous system, significant CMAP morphology changes are identified based on significant changes in amplitude, duration or both. A simplified mathematical model of normal physiological TD demonstrated that phase cancellation could not account for greater than a 50% reduction in amplitude and area, (Rhee et al., 1990). Physiological conduction block is thus defined electrophysiologically as a 50% reduction in amplitude and area, whereas TD is defined as a 50% reduction in amplitude but not area that is associated with a significant increase in CMAP duration, (Ad Hoc Subcommittee on AIDS, 1991). Since etiologies other than acquired demyelinating polyneuropathies can result in such findings, it is important to exclude confounding possibilities such as electrode shift, compression neuropathies, pseudo-conduction block, and changes due to physiological TD in CMAPs with greatly reduced amplitude. Latency abnormalities may be used but they tend to be less sensitive and specific. 11.2.1. Overview of common root stimulation pitfalls Electrode shift may occur when the examiner asks the patient to shift position during the procedure or if the electrode shifts position because it is not tightly applied to the skin surface. Just as the ulnar nerve needs to be stimulated with the elbow bent in a certain position, nerve root stimulation requires that the patient maintain the same, specific posture throughout the procedure. In the author’s laboratory, the patient is seated in a backless chair with the elbows and knees bent at a 90–110° angle with the feet resting on a stool. The recording electrodes are placed snugly on the skin surface so that electrode shift is minimized during the procedure. The patient is also instructed to minimize any movement during the procedure. Any motor nerve conduction studies previously performed will have to be repeated so that the root stimulation data are compared to the distal CMAP data with the limbs and the recording electrodes in the same relative position. The median, ulnar and peroneal nerves are liable to compression even in those persons who do not have an underlying neuropathy. For example, a patient with an

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ulnar neuropathy across the elbow may demonstrate a 60% reduction in CMAP amplitude and area when comparing the below elbow stimulation CMAP waveform to the one obtained by stimulating above the elbow. For this reason, it would be incorrect to conduct the ulnar nerve at the wrist and proceed directly to nerve root stimulation and attribute the 60% reduction in amplitude and area to a PCB at the level of the motor nerve root. For this reason, the median nerve must be conducted up to the antecubital fossa, the ulnar nerve above the elbow and the peroneal nerve above the fibular head. Eliminating the confounding variable of a compression neuropathy is an important preliminary step in all RS techniques. Pseudo-conduction block is another common source of error for these studies. Consider a patient with a C8–T1 radiculopathy who presents one week after the onset of neck pain and weakness in the hand intrinsic muscles. Since Wallerian degeneration may not be completed until 3 weeks after the precipitating event, the distal portions of the motor axons may still be excitable. The ulnar nerve in this hypothetical patient might demonstrate a normal CMAP amplitude of 7 mV at the wrist, at the below elbow site, and at the above elbow site in addition to a value of 3 mV obtained from a cervical root stimulation. It would be incorrect to attribute this finding to PCB as the finding is due to incomplete Wallerian degeneration and the appearance of a pseudo-conduction block. Electromyographers who perform RS techniques within 3 weeks of symptom onset need to be aware of this potential pitfall. Another common source of a systematic error is that due to normal physiological temporal dispersion, as all CMAPs manifest less synchrony with more proximal stimulation given that the subcomponent motor fibers have different maximum conduction velocities. The amount of separation depends on the total distance traversed. Therefore, a distal CMAP is likely to have a greater amplitude and a shorter duration than a CMAP obtained by proximal stimulation. This becomes more problematic with lower CMAP amplitudes as fewer subcomponent fibers contribute to the CMAP. Thus, the CMAP duration increases in inverse proportion to the CMAP amplitude. This will lead to accentuated degrees of TD with increasing distance in CMAPs with a reduced amplitude. Root stimulation techniques are especially prone to this potential systematic error as the stimulus site to target muscle distance is significant greater than is generally noted with more distal stimulation. In order to insure

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specificity, the target muscle to be examined must demonstrate a minimum amplitude of 1mV at its distal stimulation site. Moreover, the median, ulnar and peroneal nerves must be conducted above common sites of compression such that the CMAP obtained from root stimulation may be properly analyzed. One final source of systematic error is the failure to appreciate that absence of evidence is not evidence of absence. Just as normal late response studies do not exclude the diagnosis of proximal demyelination, RS studies that demonstrate neither PCB nor significant TD do not exclude PCB either. This has led to significant confusion in the literature wherein new syndromes such as MAMA and DADS are invoked in order to explain cases in which there is no conduction block noted and yet the patient responds to immunomodulating therapy (Saperstein et al., 2001). Most of these reports were done in the absence of RS techniques so that proximal demyelination with secondary axon loss cannot be excluded as the underlying pathophysiology. RS techniques minimize the probability of arriving at an erroneous conclusion. In those cases where RS failed to demonstrate evidence of proximal demyelination, distal nodal conduction failure or distal demyelination would result in relative inexcitability of numerous motor axons. This would result in low amplitude CMAPs, relatively normal conduction velocities and the absence of PCB; features traditionally associated with axonal neuropathies. While the appearance may be “axonal,” distal demyelination cannot be excluded (Reid et al., 2001). Thus, a RS study that fails to document PCB does not exclude the presence of distal myelin dysfunction, mild degrees of proximal myelin dysfunction or both (Menkes, 2002b). While findings of PCB or TD strongly support the diagnosis of myelin dysfunction, the absence of these findings does not exclude the diagnosis. In summary, motor NCS, late responses and RS studies are far more specific then they are sensitive.

DANIEL L. MENKES

Fig. 11.1 Electrical cervical root stimulation technique at C5–C6.

ipsilateral hand intrinsic muscles, this method uses a monopolar needle inserted over the transverse process of C7 with the anode placed at T1. Another methodology is to widen the electrical field distribution by stimulating all nerve root levels contributing to a limb’s innervation by placing a monopolar needle at the rostral edge of the nerve roots to be excited and placing the anode 1–2 at the caudal edge of the nerve roots to be excited, (Menkes et al., 1998). For RS studies to the same ipsilateral hand intrinsic muscles, the monopolar needle is placed at the C5–C6 interspace with the anode over T2. For lumbosacral nerve RS to peroneal and tibial nerve innervated muscles, the monopolar needle may be placed at L1 as depicted in Fig. 11.3 or at L5 as depicted in Fig. 11.4. Each of these techniques has its inherent advantages and disadvantages that will be discussed in subsequent sections.

11.3. Electrical nerve root stimulation (ENRS) Electrical nerve root stimulation techniques enlarge the size of the electrical field so that all electrically excitable tissues between the anode and the cathode are activated during the stimulus. Two main methods have been described as depicted in Figs. 11.1 and 11.2. One method involves placing a monopolar needle above the specific nerve root to be activated and the anode placed one level caudal to that site (Berger et al., 1987). When RS is used for conduction to the

Fig. 11.2 Electrical cervical root stimulation at C7.

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Fig. 11.3 Lumbosacral monopolar needle nerve root stimulation technique at L1.

11.3.1. Common electrical nerve root stimulation pitfalls Proper technique is absolutely essential to performing ENRS. All patients who are about to undergo this technique should give their verbal informed consent. The patient should be aware that this technique is invasive and should be queried as to whether or not they have a bleeding diathesis or are receiving anticoagulation. Patients who are at risk of prolonged or excessive bleeding should not undergo this procedure. The patient should also be informed that there may be discomfort on monopolar needle insertion and during the electrical stimulation. This is not only good medical practice but it increases the probability of patient cooperation. Once the informed consent has been obtained,

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the patient should be seated in a backless chair with their feet placed upon a stool in order to reduce the probability that the patient will move. The electrodes should be applied to the target muscles to be examined and the motor nerves should be conducted above the common sites of compression. Once accomplished, monopolar needle placement is in order. The monopolar needle electrode must be placed 1 cm lateral to the midline, ipsilateral to the side to be examined and inserted down to the periosteum of the transverse process of the vertebral level to be stimulated. It is essential that the person’s axial anatomy be taken into consideration. Longer monopolar needles will be required on individuals with greater mass. Irrespective of the needle length used, the monopolar needle must be placed such that the needle tip rests within the periosteum of the transverse process of the vertebral level needed for nerve root stimulation. Proper placement will usually result in a palpable resistance when the monopolar needle is advanced. The patient often winces slightly when the periosteum is contacted. If asked to describe the sensation, most patients will describe it as a dull “tweak” or “fullness” in that area. The needle should be inserted such that there is minimal lateral or rotational motion with manipulation. Lack of resistance or significant motion usually implies improper placement. It is also important to ensure that the anode is properly placed as well. The anode is usually applied on the skin surface in the midline of the spinal column at a caudal vertebral level. Failure to place these properly may result in submaximal stimulation of the underlying structures. It is also important to deliver two stimuli in order to insure reproducibility. Non-reproducibility is often due to one or more of the following technical factors; monopolar needle placement, monopolar needle movement, submaximal stimulation due to patient anatomical factors, electrode shift, and a distal CMAP of the target muscle demonstrating a CMAP amplitude less than 1 mV. There are additional pitfalls of ENRS that are unique to each region being examined that are discussed in subsequent sections. 11.4. Magnetic nerve root stimulation (MNRS)

Fig. 11.4 Lumbosacral monopolar needle nerve root stimulation technique at L5.

Irrespective of the vertebral level stimulated, most magnetic stimulation techniques apply a circular magnetic coil centered over the nerve root to be stimulated 1–2 cm lateral to the edge of the vertebral column, (see Figs. 11.5–11.9). Most commercially available magnetic stimulators generate a magnetic field strength of approximately 2 tesla. The full output of

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DANIEL L. MENKES

Fig. 11.5 Magnetic cervical root stimulation technique at C5–C6.

Fig. 11.7 Thoracic magnetic stimulation technique.

Fig. 11.6 Magnetic cervical root stimulation technique at C8–T1.

Fig. 11.8 Lumbosacral magnetic stimulation over vertebral L1.

the magnetic stimulator is used in order to induce an electrical field at a 90-degree angle to the magnetic field. This will result in painless activation of excitable electrical structures within the induced electrical field. As will be described in subsequent sections, the commercially available magnetic stimulators generate a less dense electrical field than is obtained with ENRS. This is due to a variety of factors including distance from the nerve root, characteristics of magnetic fields versus electrical fields and nerve root geometry.

nerve root stimulation techniques are applicable here as well. Since the magnetic stimulator is held against the skin surface of the spinal column, it is not necessary to query the patient regarding a bleeding diathesis or anticoagulation. However, contraindications to magnetic resonance imaging studies are also contraindications to MNRS. If no contraindications are present, the patient should be reassured that MNRS techniques are painless although there may some degree of associated axial muscle contraction. It is important that the distance between the nerve roots and the magnetic stimulator be minimized so that the probability of submaximal stimulation is reduced. It is also important to insure that maximum magnetic stimulator output is utilized in all MNRS techniques. Both stimulation methods and potential pitfalls are unique to each particular region of the vertebral column that is

11.4.1. Common magnetic nerve root stimulation pitfalls All of the pitfalls regarding patient positioning, proper performance of motor nerve conduction studies, and avoiding common diagnostic pitfalls common to all

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Fig. 11.9 Lumbosacral magnetic stimulation over vertebral L5.

assessed. For this reason, these issues are covered in greater detail in those particular subsections. 11.5. Stimulation sites of ENRS versus MNRS These comparisons are possible for motor nerve conduction studies (NCS) to target muscles with easily obtainable F-waves. These nerves include the median nerve to the abductor pollicis brevis (APB), the ulnar nerve to the abductor digiti minimi (ADM) and the tibial nerve to the flexor hallucis brevis (FHB). RS, motor NCS and F-wave minimum latency (FWML) techniques stimulate the largest and most heavily myelinated motor fibers. The FWML incorporates the time required to ascend the motor fibers in antidromic fashion, 1 ms of “turn around” time at the level of the axon hillock and the time required to descend in the normal orthodromic fashion down to the target muscle. If 1 ms is subtracted from the FWML, then the result is the amount of time required to travel antidromically from the site of stimulation to the axon hillock and down to the target muscle. If the distal motor latency (DML) is added to this value, this represents twice the time required for the signal to travel from the axon hillock to the target muscle. Thus, the calculated F-wave motor nerve root latency (FWRL) is defined as 1/2 [(FWML-1) + DML]. In this manner, motor nerve root latencies derived from ENRS and MNRS studies can be compared to those calculated from the FWML (Cros et al., 1990). Electrical root nerve stimulation studies performed by Mills and co-worker demonstrated that the cervical latency to the ADM was on average 1.6 ms less than the calculated FWRL consistent with a site of stimula-

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tion distal to the axon hillock and near the exit of the nerve roots through the intervertebral foramen, (Mills and Murray, 1986). Based on a proximal conduction velocity of 60 ms, this indicates that the site of stimulation is an average of 9.6 cm distal to the axon hillock (Kimura, 1974). However, a later study found no significant difference between these values and inferred that the site of excitation was within millimeters of the axon hillock (Cros et al., 1990). Cros and his coworkers also reported no statistically significant difference in the average cervical root latency between 12 pairs of ENRS and MNRS within the same subjects although there was a trend indicating a slightly longer latency from ENRS. These authors also noted a high degree of correlation, (r = 0.97), between the ulnar FWML and the latency derived directly from MNRS. However, the lumbosacral spine differs from the cervical spine in that there are two potential sites of stimulation: one near the T12 spinous process that activates the motor axons adjacent to the conus medullaris; the other, at the level where the motor nerve roots exited the spinal canal (Maertens de Noordhout et al., 1988). This was noted in a subsequent report wherein the latencies to the quadriceps and tibialis anterior were significantly shorter with MNRS when stimulated at the T12–L1 vertebral level, (Ugawa et al., 1990). A subsequent report with L5–S1 vertebral level stimulation did not observe any significant differences in latency between electrical and magnetic techniques to either the TA or the FHB (MacDonnell et al., 1992). FWML analysis to FHB by these authors indicated that the site of stimulation was approximately 13 cm distal to the anterior horn cell pool innervating this muscle. A subsequent study using L1 vertebral level stimulation demonstrated no significant differences between the FWRL and the ENRS latencies (Menkes et al., 1998). Analysis of these data also revealed a high degree of correlation (r = 0.91), between the calculated FWRL and the ENRS latency to FHB. This author previously reported that MNRS at L1 did not produce a reproducible CMAP in the FHB in 50% of the normal subjects (Menkes et al., 1994). Both MacDonnell et al. (1992) and Menkes et al., (1994) concurred that lumbosacral MNRS was very difficult to accomplish at levels rostral to L4 whereas Ugawa et al. did not report this difficulty (Ugawa et al., 1990). One possible explanation is that Ugawa and his colleagues used a more powerful magnetic stimulator that achieved a peak flux of 4.6 tesla as opposed to the magnetic stimulator used by the other groups that reached a peak flux of only 1.5 tesla. The average stature of Ugawa and his coworkers’ subjects

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was significantly less (range 153–178 cm) than the studies by MacDonnell and his co-workers (range 157–193 cm) and Menkes and his colleagues (range 155–196 cm). It is also likely that Ugawa’s controls had significantly smaller body mass indices than those encountered by the other authors. The combination of a shorter, leaner subject and a more powerful magnet would greatly lessen the probability of nerve root inexcitation at the T12–L1 vertebral level. All of these studies suggest that there are two potential sites of motor nerve root stimulation, one near the axon hillock and the other at the exit of the nerve root through the intervertebral foramen. Thus, it is important to insure that normal values used in assessing a patient are done in exactly the same manner as the published technique. It should also be emphasized that RS techniques that activate the more distal site will not detect pathology affecting the motor nerve roots between the site of proximal stimulation, adjacent to the axon hillock, and the intervertebral foramen. 11.6. Cervical root stimulation techniques Cervical MNRS and ENRS techniques are performed in a similar manner although published ENRS techniques vary the site of monopolar needle placement. Both techniques utilize maximum stimulation using the same band pass filter settings as those used for routine motor NCS. ENRS utilizes a maximum current of 100 mA or a voltage of 400 V at a duration of 1 ms. MNRS utilizes 100% output of the magnetic

stimulator. Therefore, it is imperative that the electromyographer use normal values that were obtained in an identical manner. For MNRS, the identical magnetic stimulator should be used. Tables 11.1 and 11.2 summarize some normal values for both cervical MNRS and ENRS methodologies used in the authors’ laboratory. Latencies are provided for the biceps, triceps, ADM and APB. Regardless of whether MNRS or ENRS is used for stimulation, recordings over the median nerve innervated APB merit a separate discussion because these studies require a collision technique. 11.7. Abductor pollicis brevis collision technique The C8 and T1 motor nerve roots innervate both the thenar and hypothenar muscles. Therefore, RS at these levels will activate all motor fibers in both the median and ulnar nerves. The CMAP recorded from the APB in this situation will be the algebraic sum of the subcomponent sine and cosine waves from all nearby muscles including those with ulnar nerve innervation. Depending on the waveforms of these adjacent ulnar innervated muscles, there may be an additive effect on the APB CMAP that might obscure a conduction block. Conversely, there may be a phase cancellation effect resulting in a false manifestation of abnormal temporal dispersion or a conduction block. In order to overcome this confounding variable, a collision technique must be employed in order to eliminate the contribution of the ulnar nerve innervated muscles to the APB CMAP. This technique is similar to the one used

Table 11.1 Summary of magnetic cervical root stimulation data Latency data

Amplitude data

Target muscle

Reference

Mean ± SD (ms)

Upper limit (mean ± 3 SD)

Biceps Biceps Biceps Deltoid Triceps APB– no collision APB– no collision APB– no collision ADM ADM

Ingram et al, 1988 Cros et al., 1990 Ugawa et al., 1990 Ugawa et al., 1990 Cros et al., 1990 Ingram et al, 1988 Oh et al., 1993 Ugawa et al., 1990 Cros et al., 1990 Oh et al., 1993

5.3 ± 0.42 6.3 ± 0.8 5.3 ± 0.8 5.0 ± 1.0 6.0 ± 0.85 13.0 ± 0.96 14.0 ± 1.3 11.8 ± 1.0 13.8 ± 1.5 13.84 ± 1.51

6.6 8.7 7.7 8.0 8.55 15.9 17.9 14.8 18.3 18.37

Amplitude (mean ± SD)

6.6 ± 5.5

5.7 ± 5.4

5.3 ± 2.5

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Table 11.2 Summary of electrical cervical root stimulation data Latency data

Amplitude data

Target muscle

Reference

Mean ± SD (ms)

Upper limit (mean ± 3 SD)

Deltoid Biceps Biceps Biceps Triceps Triceps APB–no collision APB– w/collision ADM ADM

Ugawa et al., 1990 Menkes et al., 1998 Ugawa et al., 1990 Cros et al., 1990 Cros et al., 1990 Menkes et al., 1998 Ugawa et al., 1990 Menkes et al., 1998 Cros et al., 1990 Menkes et al., 1998

4.7 ± 0.6 5.7 ± 0.45 5.0 ± 0.8 5.8 ± 0.8 5.4 ± 0.85 5.2 ±1.0 12.0 ±1.2 14.3 ± 1.1 14.1 ± 1.5 14.4 ± 1.6

6.5 7.05 7.4 8.2 7.95 8.2 15.6 17.6 18.6 19.2

to evaluate a potential Martin-Gruber anastomosis, (see Figs. 11.10 and 11.11). The ulnar nerve is stimulated with simultaneous recordings over the APB and ADM. This technique requires an electromyograph capable of delivering two separate stimuli at a fixed interstimulus interval. The stimulus intensity is slowly increased until the ulnar nerve is being supramaximally stimulated with no negativity noted over the APB CMAP that is being recorded simultaneously. This APB CMAP should

Amplitude (mean ± SD)

16.3 ± 4.5 9.9 ± 5.5 10.0 ± 5.4 16.3 ± 5.3 8.4 ± 2.8 6.6 ± 2.5 8.0 ± 2.6

only demonstrate an initial positivity, as depicted in Fig. 11.11. This value is the amount of stimulation that will be given over the ulnar nerve prior to the RS that will occur 6 ms later. This will result in an ulnar nerve antidromic impulse that will “collide” with the descending RS volley that will eliminate the contribution of the C8–T1 nerve roots to the ulnar nerve that would normally activate muscles adjacent to the APB. Figure 11.12 demonstrate the APB and ADM CMAP waveforms without the collision technique. The APB Fig. 11.10 Ulnar nerve collision technique.

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demonstrates different morphology from that usual observed on conventional nerve conduction studies because of the contribution to the APB CMAP from adjacent ulnar nerve innervated muscles. By contrast, Fig. 11.13 demonstrates normal APB waveform morphology and the absence of an ADM CMAP due to an effective collision technique. 11.8. Electrical nerve root stimulation techniques

Fig. 11.11 Waveforms showing supramaximal ulnar stimulation and no median stimulation. [Note the initial positivity of the APB CMAP.]

Fig. 11.12 Waveforms in a normal subject without the collision technique. [Note the initial positivity of the APB CMAP.]

Many different techniques have been advocated but all of them attempt to increase the probability of nerve root stimulation by widening the distribution of the electric field by separating the anode and the cathode. The cathode is a monopolar needle placed at the appropriate vertebral level down to the periosteum 1 cm lateral to the midline on the side to be stimulated and in contact with the transverse spinous process. The anode, usually a circular ground electrode, is placed caudal to the cathode so that the electrical field can activate all excitable tissue between these sites. The maximum output of the electromyograph is used; 100 mA or 400 V at a duration of 1 ms with the filters set at a band pass

Fig. 11.13 Waveforms in a normal subject with the collision technique. [Note that the normal morphology of the APB CMAP has been restored and that no ADM CMAP is observed]

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of 10 Hz to 10 kHz. The gain is set at 2–5 mV per division at a sweep of 5 ms per division. Stimulation may be performed at C5–C6 for the biceps, triceps, APB and ADM as depicted in Fig. 11.1. If the ADM and APB are the only target muscles of interest, then stimulation at the C7 transverse process may be performed as depicted in Fig. 11.2 (Berger et al., 1987). 11.8.1. Specific cervical electrical nerve root stimulation pitfalls Although cervical ENRS with the cathode at C7 and the anode at T2 produces a higher electrical field density at the C8–T1 nerve roots, pneumothorax is a known risk of this procedure (Sander et al., 1997). Stimulation at C5–C6 produces similar waveforms and has no reported incidence of pneumothorax, (Sander et al., 1999). Therefore, cervical nerve root stimulation should be performed first at the C5–C6 level. Stimulation at the C7 level may be performed at the electromyographer’s discretion in order to confirm an abnormality noted at C5–C6. Prior to performing this technique, the patient should be informed of the potential for pneumothorax and instructed to report any symptoms of dyspnea. Any such symptoms should be evaluated emergently. Great care must be taken in order to insure that the needle is inserted 1 cm lateral to midline so that the thecal sac is not pierced with a resulting traumatic myelopathy. For similar reasons, the needle must be advanced slowly in order to avoid trauma to surrounding tissue. 11.9. Magnetic nerve root stimulation techniques Unlike ENRS, MNRS techniques are noninvasive and painless. Most studies have reported data using magnetic field strengths of less than 2.2T. It must be emphasized that the spinal cord is not directly stimulated. One of the first publications by Barker and his co-workers noted that optimal cervical MNRS stimulation site was just lateral to the spinal column as close to the exit foramina as possible as illustrated in Fig. 11.3 (Barker et al., 1987). Other authors confirmed that the maximum amplitude from cervical MNRS is obtained by centering the coil over the spinous processes of C3, C4, C5, or C6 (Chokroverty et al., 1991). Another study reported that MNRS to the biceps, triceps and ADM were highly reproducible (Cros et al., 1990). These authors noted that the latencies from MNRS were statistically similar to those obtained from ENRS but that the amplitudes observed

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from MNRS were significantly less than those derived from ENRS. 11.9.1. Specific cervical magnetic nerve root stimulation pitfalls While there is no risk of traumatic neural structural injury, it should be recalled that metallic objects in or around the stimulated region may react to the sudden change in magnetic field density. The safety of this technique in patients with vagal nerve stimulators, cardiac pacemakers and previous penetrating trauma from metallic objects is unknown. Until such data are available, patients should be questioned as to whether or not they have any contraindication to magnetic or electrical stimulation in this region. If there is any question, then neither ENRS nor MNRS should be performed. Another potential pitfall is failure to maintain the proper positioning of the magnetic stimulator throughout the MNRS study. Small changes in magnetic stimulator orientation can have profound effects on the electrical field density and geometric distribution. It is essential that both the patient and the examiner maintain the same relative position throughout the study. 11.10. Summary/Comparison Magnetic nerve root stimulation and ENRS both produce statistically similar latencies with cervical RS. A statistically prolonged latency with either technique may be found with motor nerve root pathology. However, the amplitudes derived from MNRS are significantly lower than those obtained from ENRS. The report by Cros and his co-workers demonstrated that the ratio of the CMAP from MNRS to ENRS was 0.66 for the biceps, 0.57 for the triceps and 0.8 for the ADM. However, the range of these ratios to the ADM was 0.9 to 1. In another publication that examined 30 normal subjects, MNRS did not elicit CMAPs from the ADM in 4 subjects and from the APB in three subjects, (Oh, 1993). However, no collision technique was used in order to eliminate the contribution of ulnar-innervated muscles from the APB CMAP. While it is possible that phase cancellation may have contributed to the APB findings, the ADM abnormalities cannot be explained on this basis. Another group of authors reported similar findings as well as the observation that ENRS was more painful than MNRS especially at the C5–C6 site, (Evans et al., 1990). Thus, MNRS may be used as a screening test for proximal pathology. If no significant latency prolongation or

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amplitude reduction is noted, then ENRS need not be performed. However, any abnormalities noted on MNRS must be confirmed with ENRS prior to diagnosing abnormal temporal dispersion or a PCB. 11.11. Thoracic root stimulation techniques 11.11.1. Electrical nerve root stimulation The techniques for the thoracic region follow the same basic principles as those described in the previous section. However, there is a paucity of human subject data on ENRS in this region as there have been concerns regarding inducing cardiac arrhythmias. However, a canine model demonstrated no adverse effect on cardiac function with direct spinal cord electrical stimulation at T9–T10 at 50 mA or greater at 50 Hz for a duration of 1–2 seconds, (DiMarco et al., 1999). Another study of lumbosacral root stimulation with the dorsal insertion of a needle cathode at S1–S2 and the anode placed ventrally over the midpoint between the umbilicus and the tip of the xiphoid process did not report any adverse cardiac events (Troni et al., 1996). Despite these studies, it is likely that additional animal model data will be required before local IRBs will approve thoracic ENRS studies in human volunteers so that normal data may be obtained. 11.11.2. Magnetic nerve root stimulation This technique has been used to evaluate motor nerve root latencies from the thoracic region. One publication described thoracic region magnetic stimulation with a circular coil placed over the posterior axillary line and recording CMAPs from the upper rectus abdominus, external oblique and intercostals (Chokroverty et al., 1995). These authors reported that one patient with diabetic radiculoneuropathy demonstrated prolonged latencies to these muscles. The utility of this technique remains to be confirmed by other authors. Neither these authors nor another group that stimulated the thoracic region between T2 and L3 with the coil centered over each interspace between these levels reported any adverse events with MNRS (Tsugi et al., 1993). Despite these studies, the same caveats regarding cervical MNRS apply to thoracic stimulation as well. 11.11.3. Discussion Thoracic MNRS has been conducted without any reports of adverse events. However, this technique has

DANIEL L. MENKES

limitations not encountered in other spinal regions. F-waves are difficult to obtain from thoracic musculature because of the relatively short distances involved between the axon hillock and the target muscle. In distal limb muscles, the distance along the nerve allows for the CMAP discharge to complete and the largest motor axons to return to their resting potentials prior to the arrival of the depolarizing F-wave volley. In shorter nerve segments, the largest motor axons have not returned to their resting potential by the time the F-wave volley arrives at the motor end plate. Thus, F-wave methods cannot be used to calculate the site of MNRS activation although it is likely that it does occur proximal to the motor nerve root exit from the intervertebral foramen (Chokroverty et al., 1995). These authors remark that the latency of the potentials remains relatively stable but the CMAPs have highly variable amplitudes and thus the waveforms do not always reproduce. Therefore, any electrophysiologic diagnosis made on the basis of thoracic MNRS should be based solely upon latency criteria. However, a prolonged latency is difficult to interpret without knowing the amplitude with certainty as motor latencies prolong in proportion to the degree of axon loss whereas pathology of the myelin sheath is more likely to produce a prolonged latency out of proportion to the degree of amplitude reduction. Therefore, thoracic MNRS techniques may not reliably distinguish a diabetic thoracic radiculopathy due to demyelination versus a variant of diabetic radiculoplexus neuropathy that is a vasculopathy (Dyck and Windebank, 2002). 11.12. Lumbosacral root stimulation techniques These techniques are relatively similar to those conducted in the cervical region with some minor differences. The first is that the usual target muscles examined are the femoral nerve innervated rectus femoris (RF), the peroneal nerve innervated tibialis anterior (TA) or extensor digitorum brevis (EDB) and the tibial nerve innervated flexor hallucis brevis (FHB). Of these target muscles, F-waves are most reliably obtained from the FHB, less reliably from the peroneal innervated muscles and are generally unobtainable from the RF for reasons described in the previous section. This region has two potential sites of stimulation, adjacent to the axon hillock and near the exit of the nerve roots from the intervertebral foramen (Maertens de Noordhout et al., 1988). More distal stimulation is likely whenever the caudal portions of the vertebral column are stimulated and when a more

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powerful magnetic stimulator is used for motor nerve root excitation. Therefore, all patient data must be compared against laboratory-specific normal values or in exactly the same manner as the technique published in the literature. This may be the explanation for the wide range of normal values published in the literature. Unlike the cervical spine, the distance between the anterior horn cell pool and the nerve root exit from the intervertebral foramen is much longer. If one assumes a proximal conduction velocity of 50 m/s in the lumbosacral spine, then RS techniques activate the S1 nerve root approximately 13 cm distal to the anterior horn cells (MacDonnell et al., 1992). Thus, this technique omits a significant portion of the proximal nerve root segments. By contrast, stimulation of the L1 region activates these nerve roots adjacent to the axon hillock (Ugawa et al., 1990). Another significant difference from cervical region RS is that there is relatively little contribution of muscles innervated by other nerves to the RF, TA and FHB CMAPs. Therefore, no collision technique is required. 11.12.1. Electrical nerve root stimulation Previous sections have discussed methodologies by which the nerve roots may be stimulated at or about the L1 vertebral level which should activate the motor roots near the axon hillock as depicted in Fig. 11.3. Alternatively, stimulation at the L5–S1 level is more likely to activate the motor nerve roots as they exit their intervertebral foramina as illustrated in Fig. 11.4. Tables 11.3 and 11.4 list some of the normative data for each of these techniques. Irrespective of the tech-

nique used, a monopolar needle is inserted down to the periosteum of the transverse process ipsilateral to the target muscles being evaluated. The target muscles commonly evaluated include the tibialis anterior (TA) and the flexor hallucis brevis (FHB). The stimulus is given as maximum output of the electromyograph, either 400 V or 100 mA at a stimulus duration of 1 ms. The filter settings are the same as for conventional NCS with a band pass of 10 Hz to 10kHz. The display gain is usually set at 2mV per division with a sweep of 10 ms per division. Figure 11.14 demonstrates normal TA and FHB CMAP waveforms acquired with ENRS at L1. Any abnormality noted at the L1 nerve root level should be confirmed with stimulation at the L5–S1 level. 11.12.1.1. Specific lumbosacral electrical nerve root stimulation pitfalls Unlike the cervical region, there is much less risk of inadvertently traumatizing the spinal cord when stimulating at L1 and virtually no risk when stimulating at the L5 spinal level. However, the distance to the transverse processes in this region are often significantly greater than in the cervical region. Therefore, longer monopolar needles must be used in order to insure contact with the periosteum. Performing all preparatory nerve conduction studies with the patient bent forward at the waist by 10–30° may minimize this pitfall. The tibial and peroneal nerves should be conducted in the same relative position. The tibial nerve should be conducted at the ankle and at the popliteal fossa. The peroneal nerve should be conducted at both the fibular head as well as in the popliteal fossa. The EDB CMAP is less reliable

Table 11.3 Summary of magnetic lumbar root stimulation data Latency data

Amplitude data

Target muscle

Reference

Mean ± SD (ms)

Upper limit (mean ± 3 SD)

Quadriceps Tib Ant Tib Ant Tib Ant FHB FHB FHB

Ugawa et al., 1990 MacDonell et al., 1992 Menkes et al, 1994 Ugawa et al., 1990 MacDonell et al., 1992 Menkes et al., 1994 Ugawa et al., 1990

6.1 ± 0.7 13.6 ± 1.2 14.3 ± 1.2 12.7 ± 1.6 24.7 ± 2.2 26.7 ± 3.2 22.9 ± 2.2

8.2 17.2 17.9 17.5 31.3 36.3 29.5

* Expressed as a percentage of the distal amplitude.

Amplitude (mV) (mean ± SD) 36% ± 25* 0.84 ± 0.65 (see text) 25% ± 17.4* 0.83 ± 0.99 (see text)

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Table 11.4 Summary of electrical lumbar root stimulation data Latency data

Amplitude data

Target muscle

Reference

Mean ± SD (ms)

Upper limit (mean ± 3 SD)

Quadriceps Rectus Femoris Tib Ant Tib Ant Tib Ant Tib Ant FHB FHB FHB Soleus

Ugawa et al., 1990 Zileli et al., 2002 MacDonell et al., 1992 Menkes et al., 1994 Ugawa et al., 1990 Zileli et al., 2002 MacDonell et al., 1992 Ugawa et al., 1990 Menkes et al., 1994 Zileli et al., 2002

8.0 ± 1.4 9.8 ± 1.6 13.5 ± 1.2 15.6 ± 1.3 14.8 ± 1.5 14.6 ± 1.3 24.7 ± 2.2 22.9 ± 2.2 27.0 ± 1.5 15.2 ± 1.4

12.2 14.6 17.1 19.5 19.3 18.5 31.3 29.5 31.5 19.4

Amplitude (mV) (mean ± SD)

66% ± 31* 3.0 ± 1.2

64% ± 19* 7.8 ± 2.0

* Expressed as a percentage of the distal amplitude.

because the motor fibers to this muscle are liable to compression at the fibular head as well as at the extensor retinaculum. For this reason, the TA should be used instead of the EDB for assessing proximal conduction block affecting the motor fibers of the peroneal nerve.

Another potential pitfall is the failure to stimulate at both the L1 and L5 nerve root levels. Clinical weakness in a target muscle with unremarkable RS studies at L5 should suggest an abnormality proximal to the motor nerve root exit from the its vertebral foramen. In such instances, stimulation at L1 should be performed in order to evaluate for evidence of proximal myelin dysfunction based upon abnormal degrees of temporal dispersion or PCB. If L1 is stimulated first and such abnormalities are noted, then stimulation at L5 may help localize the pathological site as being either proximal or distal to the motor nerve root exit from the intervertebral foramen. 11.12.2. Magnetic nerve root stimulation

Fig. 11.14 Waveforms from electrical nerve root stimulation technique performed at L1.

Several studies have illustrated that MNRS produces a motor nerve root latency statistically equivalent to what is obtained from ENRS. However, the CMAPs are less reproducible and tend to be lower in amplitude than those obtained from ENRS. One report compared the ratio of L5–S1 ENRS and MNRS to the distally evoked CMAP. The ENRS ratio was 0.64 for the FHB and 0.66 for the TA whereas MNRS ratios were 0.25 for the FHB and 0.36 for the TA (MacDonnell et al., 1992). These authors hypothesized that MNRS did not generate enough magnetic flux density to insure supramaximal nerve root stimulation. This finding was more remarkable at the L1 nerve root level wherein 50% of normal subjects did not have an evocable CMAP over the TA or FHB and the others had amplitudes one order

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of magnitude less than those evoked by ENRS at the same level (Menkes et al., 1994). One solution advocated by Ugawa and his co-workers was to use a more powerful magnetic stimulator capable of generating 4.6 T (Ugawa et al., 1990). However, this magnetic stimulator is not commercially available. Another solution was to use a V-shaped coil analogous to the shape of a bird with a large wingspan (Maccabee et al., 1996). This large coil was then centered over the lumbosacral spine in order to generate the most reproducible CMAPs. While this modification allowed for stable CMAP latencies, there still remained a wide variation in amplitude. 11.12.2.1. Specific lumbosacral magnetic nerve root stimulation pitfalls. The same pitfalls noted in cervical MNRS apply to this region save for the fact that vagal nerve stimulators and cardiac pacemakers are unlikely to be affected by 2.2 tesla magnetic stimulators. In contrast, indwelling catheters containing metal or intrathecal pumps are much more likely to be affected by magnetic fields in this region. Until the safety of this technique is well established, such patients should not undergo either ENRS or MNRS procedures in this area. The commonly used magnetic stimulators do not generate an electrical field density similar to what is achieved with ENRS. This becomes even more problematic when the magnetic field is more distant from the electrically excitable tissue that is targeted. Therefore, patients with larger body frames or significant girth are less likely to have CMAPs similar in amplitude to those generated distally by conventional NCS. In such persons, ENRS is the preferred technique. 11.12.3. Summary/Comparison As with cervical RS techniques, MNRS in the lumbosacral spine can be used as a noninvasive means of determining the motor nerve root latency to a particular target muscle. It may also be used as a screening test for a PCB. If no block is found, then ENRS need not be performed. Conversely, all amplitude abnormalities must be confirmed with ENRS. 11.13. Indications for root stimulation studies Because RS studies are a means of extending the scope of motor NCS; they have essentially the same indications for evaluating motor nerve pathology in

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the proximal portions of the peripheral nerves. RS studies have been advocated for a variety of conditions that result in motor weakness from interference with normal motor nerve function including motor neuron diseases, radiculopathies, brachial plexopathies, and acquired demyelinating polyneuropathies. These disorders produce some combination of changes in motor nerve latency/conduction velocity or CMAP amplitude. Therefore, any disorder that can result in a change in either of these parameters may merit investigation with RS techniques. However, RS should only be performed when conventional studies are non-diagnostic. Each of these indications will be discussed briefly. Root stimulation was initially introduced as a technique used to assess patients for cervical radiculopathy. In one early study, RS detected all 18 patients who had clinical evidence of a radiculopathy whereas needle EMG detected only 11 (Berger et al., 1987). Radiography was abnormal in six of the seven patients examined but none of these patients were reported to have surgical confirmation of these lesion. Subsequent studies have not demonstrated the same degree of sensitivity and specificity reported by these authors. A review by Fisher stated that since there is no “gold standard” by which radiculopathies are diagnosed, evidence-based medicine cannot be applied to the various methodologies but needle EMG was the best established technique (Fisher, 2002). An independent review by Levin assessed much of the same literature and concluded that needle EMG was the most sensitive and specific superior to evoked potentials, late responses and RS studies (Levin, 2002). While RS studies may detect cases of nerve root pathology in the absence of needle EMG changes, it is unclear if these findings would result in a significant difference in the management of patients who present with a radiculopathy. A direct comparison of MNRS to motor NCS, late responses and needle EMG for lower extremity radiculopathies reported that MNRS studies had an overall sensitivity of 40% at L5 and S1 whereas needle EMG has a sensitivity of 90% for L5 and 80% for S1 (Weber and Albert, 2000).The consensus of opinion is that the clinical utility of RS studies for the diagnosis of radiculopathy is uncertain. By contrast, RS has been shown to be useful in assessing patients for the presence or absence of PCB; one of the hallmarks of saltatory conduction failure. It is important to differentiate patients with lower motor neuron variants of motor neuron disease from those with motor neuropathies that might respond to

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treatment. A study by Raynor and her co-authors, subsequently replicated in other laboratories, did not detect any electrophysiologic evidence of PCB in 59 nerves of 32 patients referred with motor neuron disease (Raynor et al., 1998). A subsequent study of RS studies performed with transcranial magnetic stimulation techniques did not demonstrate evidence of PCB in any of 121 patients with motor neuron disease including 22 patients with pure lower motor neuron presentations (Triggs et al., 1999). Restated, these studies suggest that PCB should not be observed in patients with motor neuron disease. Conversely, two independent studies reported that RS could identify PCB in acquired demyelinating polyneuropathies. One study that only assessed for PCB in the ulnar nerve, observed this finding in 8/31 patients suspected of having an acquired demyelinating polyneuropathy (Jaspert et al., 1995). These authors also reported that resolution of the PCB correlated with clinical recovery. A subsequent study reported that RS demonstrated PCB in 20/31 patients suspected of having an acquired myelinopathy whereas this finding was never observed in any of the normal or pathological controls (Menkes et al., 1998). Therefore, RS studies should be performed in any patient with a pure lower motor neuron-type of weakness. Those with PCB should be evaluated for the presence of an acquired myelin-dysfunction neuropathy. Conversely, patients suspected of having a lower motor neuron variant of motor neuron disease should undergo RS studies in order to document the absence of PCB that would suggest an alternative diagnosis. One caveat is that absence of evidence is not evidence of absence. A treatable neuropathy is not excluded if there is no evidence of PCB. Root stimulation has also been reported as being useful in detecting PCB across the brachial plexus. One case report discussed that C8 NERS demonstrated PCB across the lower trunk of the brachial plexus in a patient with thoracic outlet syndrome (Felice et al., 1999). Another publication reported that MNRS reliably differentiated brachial plexopathies with segmental demyelination from those with axon loss (Oge et al., 1997). These authors reported that acquired demyelinating polyneuropathies and “suspension” plexopathies demonstrated electrophysiologic features of PCB, whereas motor neuron disease, n-hexane neuropathy and “neurogenic thoracic outlet syndrome” did not. Evidence-based medicine analysis cannot be applied to these techniques given that there is no universal agreement on what constitutes the “gold standard” for the diagnosis of demyelinating brachial plexopathies, the

DANIEL L. MENKES

thoracic outlet syndrome and acquired demyelinating polyneuropathies. Therefore, any abnormalities noted on RS studies need to be placed in the clinical context of that patient’s presentation. 11.14. Conclusion Root stimulation techniques allow for an evaluation of the most proximal portions of the motor nerves. RS may be performed using magnetic or monopolar needle stimulation techniques. The latency and CMAP amplitude to the target muscle can be directly measured. Absolute latency prolongation or evidence of PCB or abnormal degrees of TD are the most clinically useful findings. For all routinely studies target muscles, the latency and amplitude may be assessed directly from RS techniques. However, a collision technique is required for motor NCS of the median nerve to the APB. MNRS techniques are less useful than ENRS techniques but may be used as a screening examination. If no abnormalities are noted on MNRS studies, then the more invasive ENRS studies may be omitted. Otherwise, ENRS studies should be performed. References Ad Hoc Subcommittee of the American Academy of Neurology AIDS Task Force (1991) Research criteria for the diagnosis of chronic inflammatory demyelinating polyneuropathy. Neurology, 41: 617–618. Barker, AT, Freeston, I, Jalinous, R, Eng, B and Jarratt, J (1987) Magnetic stimulation of the human brain and peripheral nervous system: an introduction and the results of an initial clinical evaluation. Neurosurgery, 20: 100–109. Berger, AR, Busis, NA, Logigian, EL, Wierzbicka, M and Shahani, BT (1987) Cervical root stimulation in the diagnosis of radiculopathy. Neurology, 37: 329–332. Chokroverty, S (1990) Magnetic stimulation in clinical neurophysiology. Boston, ButterworthHeinemann Medical, 308 pp. Chokroverty, S, Deutsch, A, Guha, C, Gonzales, A, Kwan, P, Burge, R and Goldberg, J (1995) Thoracic spinal and nerve root conduction: a magnetic stimulation study. Muscle Nerve, 18: 987–991. Chokroverty, S, Picone, MA and Chokroverty, M (1991) Percutaneous magnetic coil application of human cervical vertebral column: site of stimula-

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tion and clinical application. Electroencephalogr. Clin. Neurophysiol., 81: 359–365. Cros, D, Chiappa, KH, Gominak, S, Fang, J, Santamaria, J, King, PJ and Shahani, BT (1990) Cervical magnetic stimulation. Neurology, 40: 1751–1756. DiMarco, AF, Romaniuk, JR, Kowalski, KE and Supinski, G (1999) Pattern of expiratory muscle activation during lower thoracic spinal cord stimulation. J. Appl. Physiol., 86: 1881–1889. Dyck, PJ and Windebank, A (2002) Diabetic and nondiabetic lumbosacral radiculoplexus neuropathies: new insights into pathophysiology and treatment. Muscle Nerve, 25: 477–491. Evans, BA, Daube, JR and Litchy, WJ (1990) A comparison of magnetic and electrical stimulation of spinal nerves. Muscle Nerve, 13: 414–420. Felice, KJ, Butler, KB and Druckemiller, WH (1999) Cervical root stimulation in a case of classic neurogenic thoracic outlet syndrome. Muscle Nerve, 22: 1287–1292. Fisher, MA (2002) Electrophysiology of radiculopathies. Clin. Neurophysiol., 113: 317–335. Ingram, D, Thompson, A and Swash, M (1988) Central motor conduction in multiple sclerosis: evaluation of abnormalities revealed by transcranial magnetic stimulation of the brain. J. Neurol. Neurosurg. Psychiatry, 51: 487–494. Jaspert, A, Claus, D, Grehl, H, Kerling, F and Neurndorfer, B (1995) Value of PCB study in diagnosis of inflammatory neuropathies. Nervenartz, 66: 445–454. Kimura, J (1974) F-wave conduction velocities in the central segments of the median and ulnar nerves. Neurology, 24: 539–546. Levin, K (2002) Electrodiagnostic approach to the patient with suspected radiculopathy. Neurol. Clin., 20: 397–421. Maccabee, PJ, Lipitz, ME, Desudchit, T, Golub, RW, Nitti, VW, Bania, JP, Willer, JA, Cracco, RQ, Cadwell, J, Hotson, GC, Eberle, LP and Amassian, VE (1996) A new method using neuromagnetic stimulation to measure conduction time within the cauda equina. Electroencephalogr. Clin. Neurophysiol., 101:153–166. Macdonell, RA, Cros, D, Shahani, BT (1992) Lumbosacral nerve root stimulation comparing electrical with surface magnetic coil techniques. Muscle Nerve, 15:885–890. MacLean, IC and Taylor, RS (1975) Nerve Root Stimulation to Evaluate Brachial Plexus Conduc-

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tion. Abstracts of Communications of the Fifth International Congress of Electromyography. Rochester MN, 47 pp. Maertens de Noordhout, A, Rothwell, JC, Thompson, PD, Day, BL and Marsden, CD (1988) Percutaneous electrical stimulation of lumbosacral nerve roots in man. J. Neurol. Neurosurg. Psychiatry, 51: 174–181. Menkes, DL (2002a) Proximal conduction techniques: somatosensory evoked potentials, magnetic stimulation and root stimulation. In: T. Bertorini (Ed.) Clinical Evaluation and Diagnostic Tests for Neuromuscular Disorders, ButterworthHeinemann Inc., Woburn, pp. 209–238. Menkes, DL (2002b) Axonal multifocal motor neuropathy without conduction block or other features of demyelination. Neurology, 59: 1666. Menkes, DL, Hood, DC, Ballesteros, RA and Williams, DA (1998) “Root stimulation improves the detection of acquired demyelinating polyneuropathies.” Muscle Nerve, 21: 298–308. Menkes, DL, Ring, SR, Chiappa, KH and Cros, D (1994) Cortical Magnetic Stimulation to the Lower Extremities: Normal Controls and 14 Pathologic Cases. Abstracts of the American Academy of Neurology, Washington, DC. Mills, KR and Murray, NMF(1986). Electrical stimulation over the human vertebral column. Which neural elements are excited? Electroenceph. Clin. Neurophysiol., 63: 582–589. Oge, AE, Boyaciyan, A, Gurvit, H, Yazici, J, Degirmenci, M and Kantemir, E (1997) Magnetic nerve root stimulation in two types of brachial plexus injury: segmental demyelination and axonal degeneration. Muscle Nerve, 20: 823–832. Oh, SJ (1993) Clinical electromyography: nerve conduction studies. Magnetic and High-Voltage/LowImpedance Electrical Stimulation Tests. Williams and Wilkins Inc., Philadelphia, IInd ed., pp. 406–446. Raynor, EM, Shefner, JM, Ross, MH, Logigian, EL and Hinchey, JA (1998) Root stimulation studies in the evaluation of motor neuron disease. Neurology, 50: 1907–1909. Reid, V, Black, K, Menkes, DL, Siao, P and Cros, D (2001) Chronic inflammatory demyelinating polyneuropathy. In: D Cros (Ed.). A Practical Approach to Diagnosis and Management, Williams and Wilkins, Philadelphia, pp. 85–107. Rhee, EK, England, JD and Sumner, AJ (1990) A computer simulation of conduction block: effects

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produced by conduction block versus interphase cancellation. Ann. Neurol., 28: 146–156. Sander, HW, Menkes, DL, Triggs, WJ and Chokroverty, S (1999) Cervical root stimulation at C5–C6 excites C8–T1 nerve roots and minimizes pneumothorax risk. Muscle Nerve, 22: 766–768. Sander, HW, Quinto, CM, Murali, R and Chokroverty, S (1997) Needle cervical root stimulation may be complicated by pneumothorax. Neurology, 48: 288–289. Saperstein, DS, Katz, JS, Amato, AA and Barohn, RJ (2001) Clinical spectrum of chronic acquired demyelinating polyneuropathies. Muscle Nerve, 24: 311–324. Triggs, WJ, Menkes, D, Onorato, J, Yan, RS, Young, MS, Newell, K, Sander, HW, Soto, O, Chiappa, KH and Cros, D (1999) Transcranial magnetic stimulation

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identifies upper motor neuron involvement in motor neuron disease. Neurology, 53: 605–611. Troni, W, Bianco, C, Coletti Moja, M and Dotta, M (1996) Improved methodology for lumbosacral nerve root stimulation. Muscle Nerve, 19: 595–604. Tsugi, S, Murai, Y and Yarita, M (1993) Cortical somatosensory evoked potentials evoked by magnetic stimulation of thoracic and lumbar roots. Neurology, 43: 391–396. Ugawa, Y, Kohara, N, Shimpo, T and Mannen, T (1990) Magneto-electrical stimulation of central motor pathways compared with percutaneous electrical stimulation. Eur. Neurol., 30: 14–18. Zileli, B, Ertekin, C, Zileli, M and Yunten, N (2002) Diagnostic value of electrical stimulation of lumbosacral roots in spinal stenosis. Acta Neurol. Scand., 105: 221–227.

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CHAPTER 12

Motor unit number estimation in peripheral neuropathies Mark B Bromberg* Department of Neurology, University of Utah, UT, USA

12.1. Introduction Motor nerves are involved in the majority of peripheral neuropathies, and most neuropathies include axonal degeneration. Assessing the degree of axonal loss in peripheral neuropathies is challenging. Estimates of severity of sensorimotor neuropathies are primarily qualitative, based on clinical symptoms, clinical scales, or sensory nerve studies. Motor nerve denervation cannot be accurately measured from muscle strength or routine electrodiagnostic studies due to the compensatory effects of collateral reinnervation. However, the electrophysiologic technique of motor unit number estimation (MUNE) is not affected by collateral reinnervation, and remains the only method that can provide a quantitative estimate of the number of motor units innervating a muscle. This chapter reviews the pathophysiology of denervation and collateral reinnervation and its effect on routine electrodiagnostic testing. The underlying principles and methodology of MUNE are presented. Experience with MUNE assessing the degree of axonal loss in peripheral neuropathies is reviewed. The reader is directed to the proceedings of a recent symposium on MUNE for more details about the technique (Bromberg, 2003b). 12.2. Pathophysiology of denervation and reinnervation The concept of a motor unit is easily grasped, but it is experimentally difficult to visualize the muscle fibers that make up a single motor unit. Single motor units can be studied by the technique of glycogen depletion where single ventral root axons are isolated

*

Correspondence to: Dr. Mark B Bromberg, MD, PhD, Department of Neurology, University of Utah, 50 North Medical Drive, Salt Lake City, UT 84132, USA. E-mail address: [email protected] Tel.:+1-801-585-5884; fax: +1-801-585-2054.

and stimulated continuously to deplete glycogen stores in the muscle fibers of that motor unit (Edström and Kugelberg, 1968). From such studies in the rat, the following features of motor units have been elucidated. It is rare for muscle fibers from the same motor unit to lie adjacent to each other. The tesselation of muscle fibers observed on routine histologic sections of muscle represent fibers from many motor units. Muscle fibers of a single motor unit occupy restricted cross-sectional areas within the whole area of the muscle, and at any given site in a muscle, the areas of 20 or more motor units overlap. With partial axonal loss, surviving terminal motor axons sprout collateral branches to reinnervate the denervated muscle fibers. Glycogen depletion studies reveal the following motor unit changes with reinnervation (Kugelberg et al., 1970). Reinnervation leads to a higher density of muscle fibers belonging to the surviving motor units, and more muscle fibers of the same motor unit will be adjacent to each other. Reinnervation is shared by many motor units within the muscle, observed as fiber type grouping on histologic sections. With early axonal loss, collateral reinnervation successfully reinnervates most denervated muscle fibers. However, there are limitations to collateral reinnervation, and fascicular boundaries within muscle restrict the extent of sprouting. With extensive axonal loss reinnervation is inadequate and greater numbers of muscle fibers remain denervated (Cohen et al., 1987). In slowly progressive neuropathies, distal muscle weakness will not be apparent until 80% of motor neurons are lost (McComas et al., 1971b). 12.3. Limitations of routine electrodiagnostic testing Routine electrodiagnostic studies used to assess peripheral neuropathies measure axonal loss and conduction velocity. The sensory nerve action potential (SNAP) and compound muscle action potential (CMAP) are useful, particularly for determining

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conduction velocity. For measures of axonal loss, the SNAP may be unrecordable when 50% of fibers remain (Rosenfalck, 1978). CMAP amplitude reflects the number of innervated muscle fibers, and amplitude values fall only when the extent of denervation exceeds the capacity for reinnervation and a sufficient percentage of muscle fibers remain denervated. In slowly progressive denervating disorders, there will be sufficient time for maximal reinnervation, and CMAP amplitude may not fall below the lower limits of normal until 50–80% of motor axons have been lost (McComas et al., 1971b). Needle EMG is sensitive to the effects of denervation and reinnervation, signaled by abnormal spontaneous activity (positive waves and fibrillation potentials) and changes in motor unit action potential (MUAP) recruitment, amplitude and waveform configuration. However, changes are customarily expressed by subjective scales (1+ to 4+), which represent qualitative estimates of denervation (Kimura, 2001). Few studies compare subjective EMG scaling with objective motor unit estimates (Schulte-Mattler et al., 2000). Single fiber EMG (SFEMG) measurement of fiber density is the most sensitive electrodiagnostic test for denervation (Stålberg et al., 1975). Fiber density measurements are based on the recording radius of a special SFEMG electrode, and values represent the closeness of muscle fibers from the same motor unit.

Fiber density values rise with collateral reinnervation, and fall when decompensation occurs (Stålberg, 1982). Correlations, therefore, are poor between fiber density values and MUNE values, CMAP or MUAP measurements (Bromberg and Larson, 1996). 12.4. Principles of motor unit number estimation Motor unit number estimation is a physiologic estimate of the number of motor units innervating a muscle or group of muscles (McComas et al., 1971a). MUNE values are calculated from the ratio (Fig. 12.1): maximal compound muscle action potential (CMAP ) amplitude or area . average single motor unit potential (MUP ) amplitude or area The principle behind MUNE is simple in concept, but there are a number of issues that have been extensively reviewed (McComas et al., 1971a; Brown and Milner-Brown, 1976; Slawnych et al., 1990; McComas, 1991; Bromberg, 2003b). Several different MUNE techniques have been developed (Table 12.1) to deal with some of these issues. The various MUNE techniques differ in the manner in which S-MUPs are obtained. 12.4.1. Muscles studied

− 100 μV +

− 2 mV +

Fig. 12.1 Incremental MUNE technique. (Top trace): Incremental responses of extensor digitorum brevis following excitation of deep peroneal nerve with threshold and slightly suprathreshold stimuli. (Bottom trace): Maximal compound muscle action potential (Modified from McComas et al. (1971a) Functional compensation in partially denervated muscles. J. Neurol. Neurosurg. Psychiatry, 34: 453–460, with permission from the BMJ Publishing Group).

When MUNE is applied to distal lower extremity muscles involved in peripheral neuropathies, the extensor digitorum brevis (EDB) muscle may be studied in isolation, but most MUNE determinations include estimates from a group of muscles innervated by one nerve. Examples in the leg are tibial-innervated abductor hallucis and other intrinsic foot muscles, and examples in the arm are median-innervated thenar eminence and ulnar-innervated hypothenar eminence muscles. Proximal muscles, such as the biceps-brachialis muscle group, can also be studied with certain MUNE techniques (Table 12.1) (McComas, 1991). 12.4.2. Compound muscle action potential The maximal CMAP is determined by the same techniques used in routine motor nerve conduction studies. Recording electrodes are placed in a belly-tendon arrangement, and the nerve is electrically activated by percutaneous stimulation to achieve the maximal CMAP. The recording electrodes are left in place to

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283

Table 12.1 Different MUNE techniques used to assess axonal loss in peripheral neuropathiesa MUNE Technique

Advantages

Incremental stimulation

●

Multiple point stimulation

●

●

Applicable to any EMG machine Passive testing; patient cooperation not necessary Applicable to any EMG machine Avoids alternation Passive testing; patient cooperation not necessary Samples range of nerve fibers Passive testing; patient cooperation not necessary

●

Applicable to distal and proximal muscles

●

●

Can provide quantitative intramuscular motor unit action potential data

●

●

● ●

Statistical

Disadvantages

●

●

● ●

● ● ●

Spike triggered averaging

a

Alternation leading to an over estimate of the MUNE Applicable to distal muscles only Applicable to distal muscles only

Assumes Poisson statistics Requires proprietary software Applicable to distal muscles only Requires intramuscular needle EMG electrode Active testing; requires patient cooperation

Techniques differ in the manner in which surface motor unit potentials (S-MUPs) are obtained.

record the surface representation of single motor unit potentials (S-MUPs). It is preferable that the active recording electrode be accurately positioned over the motor point to achieving maximal CMAP amplitude, and this position is associated with a steep waveform rise time (Bromberg and Spiegelberg, 1997). Optimal positioning of the active electrode is important in serial MUNE studies to ensure reproducibility of the maximal CMAP, or to enhance the ability to detect changes over time. However, with respect to MUNE estimates, any variation from the optimal placement of the active recording electrode will be manifest equally in CMAP and S-MUP waveforms, and MUNE values will not be affected (Bromberg and Abrams, 1995). 12.4.3. Single motor unit potential waveforms Individual S-MUPs waveforms within a muscle or group of muscles vary in size and shape, (Fig. 12.2), but typically have an initial negative deflection followed by a terminal positive deflection (Fig. 12.2). Occasional S-MUPs are recorded with almost entirely positive waveforms. These are considered to most likely represent volume-conducted motor units from adjacent muscles. Accordingly, they are usually not included in calculating the average S-MUP value (Bromberg, 2003a). Total waveform duration can be challenging to assess due to difficulties in accurately determining the onset and termination points. Peak-topeak amplitude is readily measured, and negative peak

amplitude and area are measured by extending the prewaveform baseline across the negative portion of the waveform. The distribution of S-MUP amplitudes from normal muscle spans one order of magnitude, from approximately 20 to 200 μV peak-to-peak amplitude (Bromberg and Abrams, 1995). Occasionally very low amplitude S-MUPs are recorded, but they are considered to represent motor units from distant muscles. Accordingly, S-MUPs with negative peak amplitude <10 μV or negative peak area <25 μV*ms are not included in calculating the average S-MUP value (Bromberg, 2003a). S-MUP amplitudes increase in denervated muscle due to collateral reinnervation, but the degree of enlargement remains within the same order of magnitude as normal S-MUPs (measured in amyotrophic lateral sclerosis) (Bromberg and Abrams, 1995). 12.4.4. Phase cancellation Motor unit number estimation values can be calculated using a variety of CMAP and S-MUP metrics, including peak-to-peak amplitude, negative peak amplitude and negative peak area values. The issue of concern is the effect of phase cancellation. All routine metrics of the maximal CMAP incorporate the effects of phase cancellation between the constituent motor units. Variables that affect phase cancellation include differences in conduction velocities of motor units from the

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S-MUAPs

A 5 ms

− 20μV +

MU1

MU2

MU3 MU4 SUM AVG

B

p−p

-p

-area

248

170

432

159

77

342

72

44

100

40 519 130

17 308 77

33 907 227

111

67

218

(15%)

(13%)

(4%)

Data-point by data-point average wave-form

Fig. 12.2 Effects of phase cancellation on summation of S-MUP waveforms. Four S-MUP waveforms showing initial negative deflection and later positive deflection. (A) Waveforms are aligned by onset showing how phase cancellation could affect different methods used to calculate an average value. Table to the right indicates individual and averaged values as calculated from peak-to-peak amplitude (p–p), negative peak amplitude (−p), and negative peak area (−area) metrics. (B) Average S-MUP waveform calculated by datapoint-by-datapoint averaging. Table to the right indicates averaged S-MUP values by using different metrics. Percentages represent differences between simple arithmetic and datapoint-by-datapoint averaging. Note, use of the metric negative peak area is least affected by phase cancellation. (From Doherty, T and Brown, W (1993) The estimated numbers and relative sizes of thenar units as selected by multiple point stimulation in young and older adults. Muscle Nerve, 16: 355–366. Reprinted with permission of John Wiley and Sons, Inc.).

site of stimulation to the recording electrode, and differences in the shapes of S-MUPs (relative position of peak negativity and positivity within the waveform) (Fig. 12.2). The metric of peak-to-peak amplitude will be most affected by phase cancellation. Thus, when the average S-MUP is calculated by arithmetic addition of peak-to-peak amplitudes, phase cancellation will not be accounted for and the average S-MUP value will be artifactually high, leading to an artifactually low MUNE value. Metrics of negative peak amplitude or negative peak area are less affected by phase cancellation, but the position of peak negativity within individual S-MUP waveforms represents phase cancellation and can affect arithmetic addition to determine the average S-MUP (Doherty et al., 1993). The preferable method to determine the average S-MUP is point-by-point averaging of individual

S-MUP waveforms (Fig. 12.2). This technique will include the effects of phase cancellation, but requires special computer software (Doherty et al., 1993; Bromberg, 2003b). When different metrics (peak-topeak, negative peak amplitude or area) and different methods to determine the average S-MUP (arithmetic and point-by-point) are used, MUNE values can differ upto 35% (Doherty et al., 1993). The effect of differences in conduction velocity on phase cancellation will be least problematic if all waveforms, including the CMAP and all S-MUPs, are obtained from one nerve stimulation electrode site along the nerve, and most problematic if different stimulation sites are used for the CMAP and S-MUP. One method to neutralize the effect of different conduction velocities on phase cancellation is to obtain the maximal CMAP from a very distal stimulation site

MOTOR UNIT NUMBER ESTIMATION IN PERIPHERAL NEUROPATHIES

so as to minimize the effect of phase cancellation. Individual S-MUP waveforms elicited by stimulation at different sites along the nerve can be aligned by onset of the negative deflection before carrying out point-by-point averaging (Fig. 12.2) (Doherty et al., 1993). 12.4.5. Single motor unit potential sample size and sample bias The challenge in MUNE is to obtain a representative sample of S-MUPs to calculate an average value. There are two issues, sample size and sample bias. Normal muscles are innervated by 120 to 400 motor units (McComas, 1991). Although < 10% of the normal population (10 to 20 S-MUPs) is customarily sampled, there is little change in the calculated MUNE values when > 15 S-MUPs are used to determine the S-MUP value (Bromberg, 1993; Doherty and Brown, 1993). Sample bias is potentially more problematic. Most MUNE techniques rely on electrical activation of the nerve with graded currents, from low to high. It is known from in vitro single axon studies that large diameter axon have lower thresholds for electrical stimulation, and this was thought to impose a sampling bias in MUNE studies, particularly with techniques that use only low current strengths. However, subsequent investigations comparing S-MUP values obtained by a variety of MUNE techniques indicate little bias (Stein and Yang, 1990; Doherty and Brown, 1993). For MUNE techniques that rely on nerve stimulation, the overall geometry of the nerve, including orientation of fascicles and overlying tissue, neutralizes the excitability relationship between axonal size and current intensity. 12.4.6. Test–retest reliability Test–retest reliability has been assessed for most MUNE techniques in normal subjects and those with denervation. When expressed as a correlation coefficients, reliability is 15–20%. Of note, reliability is higher in denervating conditions when there are reduced numbers of motor units (Bromberg, 1993; Doherty and Brown, 1993). 12.4.7. Independent verification It is important to appreciate that there is no independent or anatomical method (gold standard) to count motor axons innervating a muscle. MUNE values

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remain estimates (McComas, 1991; Bromberg, 2003b). Animal studies allow some degree of comparisons between MUNE and anatomic verification and show good correlations (Arasaki et al., 1997). Human autopsy studies require assumptions about the percentages of motor efferent and muscle afferent fibers within a motor nerve, but also show good correlations. 12.5. Motor unit number estimation techniques A number of MUNE techniques have been developed that differ in how S-MUPs are obtained. Selection of a MUNE technique is guided by a number of factors, which include equipment availability, clinical experience, and study objectives. Direct comparisons show good concordance between techniques listed in Table 12.1, with no technique markedly better than another technique (Stein and Yang, 1990; Doherty and Brown, 1993; Lomen-Hoerth and Olney, 2000). Within a study, it is important to have internal validation of the chosen MUNE technique, including measures of test–retest reliability and comparisons with a control population. The most common MUNE techniques used to assess axonal loss in peripheral neuropathies (Table 12.1), and their modifications, are discussed, along with their advantages and pitfalls. 12.5.1. Incremental stimulation motor unit number estimation technique 12.5.1.1. Principles This is the original MUNE technique and is based on applying incremental increases in nerve stimulation intensity to generate an envelope of evoked responses (McComas et al., 1971a). Each step in the envelope is considered to represent the activation of single motor axons that are serially added to the growing evoked response. 12.5.1.2. Methods Surface electrodes are placed over the muscle or muscle group in a belly-tendon arrangement and a maximal CMAP is recorded. The stimulating electrode remains fixed in the same position on the nerve. The display sensitivity is raised to 100–200 μV/div to help visualize low amplitude steps in the response envelope. The stimulus intensity is lowered to 3–10 mA in order to activate the first axon, indicated by an all-or-none response. By small increases in stimulation

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intensity, an envelope of responses is obtained with 8–10 discrete steps before the increments in the envelope become indistinguishable (Fig. 12.1). The number of steps is divided into the peak-to-peak amplitude of the envelope to determine the average amplitude of each step. This average value represents the average S-MUP, and is used to calculate the MUNE value.

the serial addition of axons. This is termed “alternation” between different combinations of axons in the response. The result is uncertainty in the number of steps that are truly represented in the envelope; a larger number of perceived steps results in a smaller average S-MUP, and hence a larger MUNE value (McComas et al., 1971a; Brown and Milner-Brown, 1976).

12.5.1.3. Advantages (1) The incremental stimulation technique can be used on any EMG machine. (2) The stimulating electrode is fixed in the same position for the maximal CMAP and determination of the average S-MUP, and the effects of phase cancellation due to different axonal conduction velocities will be incorporated in the CMAP and S-MUP waveforms.

12.5.1.5. Modifications In the original technique, individual S-MUPs within the envelope are not isolated from the envelope. (1) An early modification used serial templates to isolate individual S-MUPs (Ballantyne and Hansen, 1974). This modification requires special computer software. The first potential is used as the first template and subtracted from the next larger potential to give the waveform of the second S-MUP; the first two potentials are then used as the second template and subtracted from the third larger potential in the envelope to identify the third S-MUP. This is repeated until 8–10 S-MUPs from the envelope are isolated. This modification also relies on measurement of S-MUP area rather than amplitude. (2) A later modification used an automated template subtraction technique where every subtracted waveform was compared to determine if any waveform appeared more than once. Those that did so were rejected from the average S-MUP calculation (Galea et al., 1991). This modification requires proprietary

12.5.1.4. Issues and pitfalls It was recognized that an axon does not have a discrete threshold current for activation, but activation follows an excitability curve from low probability of activation at lower currents to high probability at higher currents (Fig. 12.3). Thus, at a fixed current intensity within an excitability curve, an axon may or may not be excited with repeated stimuli. When a number of axons have similar and overlapping excitability curves, the steps in the envelope will reflect different combinations of axons being excited, and not simply

100 3 80 Firing index (%)

2 1

60 1 40

3

2

20 200 μV/div

2 ms/div 0

2

4

6

8 10 12 14 16

0 Stimulus current

Fig. 12.3 The effects of alternation on calculating an average S-MUP. (Left): Three waveforms generated by slowly increasing the stimulation current, as in the incremental stimulation MUNE technique. The assumption is that waveform number 3 represents the sum of the responses from axons 1 + 2 + 3. (Right): Excitability curves of axons 1, 2 and 3 show that at a constant current intensity (dashed line), the probability of firing for each axon is less than 100%. Therefore, waveform number 3 elicited at the current intensity indicated by the dashed line could represent the sum of the responses from axons 1 + 2 + 3, or axons 1 + 3, or axon 3 alone. Uncertainty in the number of axons contributing to the total response affects calculation of the average response.

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software on a single EMG machine that is no longer available.

200 μV/div

2 ms/div

12.5.2. Multiple point stimulation motor unit number estimation technique 12.5.2.1. Principles This technique was developed to avoid the problem of alternation by activating only single axons, one at a time (Kadrie et al., 1976). Different single S-MUPs are obtained by moving the stimulating electrodes to different sites along the nerve (Kadrie et al., 1976; Doherty and Brown, 1993). 12.5.2.2. Methods Surface electrodes are placed and a maximal CMAP is recorded. The display sensitivity is raised to 50–100 μV/div to record the low amplitude S-MUPs. The stimulus intensity is lowered to 3–10 mA to activate the first axon. This is verified by an all-or-none response to raising and lowering the stimulation current (Fig. 12.4). The stimulating electrode is moved to a different site along the nerve and another all-or-none response is obtained. Ten to fifteen responses are collected (Fig. 12.5) and averaged to obtain the average S-MUP used to calculate the MUNE value. 12.5.2.3. Advantages (1) The multiple point stimulation technique avoids the problem of alternation. (2) This technique can be used on any EMG machine.

200 μV/div

2 ms/div 0

2

4

6

8

10

12

14

16

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Fig. 12.4 (Top): All-or-none response. (Bottom): Apparent S-MUP that fractionated after many trials, indicating that the larger waveform was the sum of two or more waveforms.

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Fig. 12.5 S-MUPs waveforms obtained from multiple point stimulating MUNE technique.

12.5.2.4. Issues and Pitfalls (1) A suitable number of stimuli must be delivered to ensure that the response represents a single axon; two axons with similar excitability curves could be activated together as a single response most of the time (Fig. 12.4), and fractionation of the response into two S-MUPs might become apparent only if a suitable number of stimuli were given (>10). (2) The stimulating electrode is moved to different sites along the nerve to obtain single S-MUPs. If point-by-point averaging is used, the effects of phase cancellation from late arriving waveforms will be problematic, unless the waveforms are aligned by their onset. 12.5.2.5. Modifications Single motor unit potentials are obtained one at a time, which can be time consuming. A modification, called adapted multiple point stimulation technique (AMPS), has been developed that combines incremental and multiple point stimulation (Wang and Delwaide, 1995). At each site along the nerve, the stimulation intensity is raised to obtain a two or threestep envelope. Template subtraction is used to separate individual S-MUP waveforms. A suitable number of S-MUPs can be obtained from five to six sites along the nerve, reducing the time required for the study.

288

12.5.3. Statistical motor unit number estimation Technique 12.5.3.1. Principles The statistical technique is a novel approach using Poisson statistics to determine the average response based on the variability of the response (Daube, 1995). When multiple stimuli are delivered at a constant stimulation intensity, the evoked response will vary from trial-to-trial (envelope of responses). It can be assumed that this response variability reflects the addition and loss of axons to the response, and thus represents the phenomena of alternation. Poisson statistics assumes that the underlying changes represent discrete steps or axonal responses, and the statistical variance of the amplitude of the response over many trials is equal to the mean value of the change in the response, which is the average value of the S-MUP. 12.5.3.2. Methods Surface electrodes are placed and a maximal CMAP is recorded. The subject should remain at complete rest to prevent adventitious movements (Daube, 2003). The stimulating electrode is fixed and the nerve is “scanned” with a series of 30 stimuli delivered with increasing intensity from just subthreshold to just maximal to generate the total envelope of evoked responses (Fig. 12.6). This scan of the evoked response is used to identify portions of the response envelope that will be sampled. Usually, three to four regions are sampled to determine the variance and S-MUP amplitude value at each of these regions. The variance of the response in each region is determined by applying sets of 30 stimuli. For each set of 30 stimuli, the variance is calculated and an average S-MUP area determined. Repeated sets of 30 stimuli are performed until the standard error of the different determinations is <10%. The average S-MUP determined in a region is then used to calculate a MUNE value for that region. The other designated regions are similarly sampled. The regions selected for detailed study from the scan cure include regions with large steps reflecting large amplitude motor units, and one “average” region, considered to represent the unsampled region. A MUNE value is then calculated for the unsampled region using the smallest average S-MUP value. Finally, all regional and unsampled MUNE values are combined for a total MUNE value.

MARK B BROMBERG

12.5.3.3. Advantages (1) Alternation is the major issue in the incremental stimulation MUNE technique, but is the basis of the statistical technique. (2) The incremental and multiple point stimulation MUNE techniques rely on electrical activation of low threshold axons, and there has been concern for a sampling bias toward large diameter axons and hence higher amplitude S-MUPs. In the statistical technique, a wide range of axon thresholds and their S-MUPs are sampled. However, comparison studies between MUNE techniques do not support a significant bias due to axon diameter because the electrical threshold of an axon has greater dependence upon the overall tissue impedance to stimulus current flow around the nerve than the simple axon excitability threshold (McComas, 1991). (3) A degree of statistical robustness is included in the determination of the average S-MUP for each region by repeatedly applying sets of 30 stimuli until the standard error of the response is < 10% of the mean response. (4) The stimulating electrode is fixed for the maximal CMAP and all subsequent determinations, and the effects of phase cancellation due to different axonal conduction velocities over different conduction distances are incorporated in the waveforms. (5) Negative peak area is the metric used in all calculations. 12.5.3.4. Issues and Pitfalls (1) The statistical technique requires proprietary software available on a single brand of EMG machine. (2) An assumption of Poisson statistics is that the distribution of response amplitudes is skewed to the left, but that does not always occur during the 30 stimuli. Several methods have been used to adjust the stimulus intensity to force a skewed distribution (LomenHoerth and Olney, 2001; Daube, 2003). (3) With denervation there will be a reduced numbers of motor units and some will be enlarged from collateral reinnervation. This will appear in the scan curve as large jumps in the amplitude. There are several methods available to adjust and account for large and small motor units (Lomen-Hoerth and Olney, 2000; Daube, 2003). (4) The S-MUP values that are calculated from the variance of the response represent putative values and not physiologic single motor unit potentials. (5) There are a large number of variables in how data are obtained with the statistical technique. These include forcing the data into a Poisson distribution, managing large gaps, use of different window sizes and different methods to calculate the final MUNE value (see Modifications). Application of these variables in

MOTOR UNIT NUMBER ESTIMATION IN PERIPHERAL NEUROPATHIES

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Fig. 12.6 MUNE by statistical technique. (Upper left panel): CMAP scan envelope from threshold to supramaximal response evoked by 30 graded stimuli. (Middle panel): Same scan, but CMAP amplitudes (small horizontal marks) displayed as percent of maximal response, Note, 4 regions selected by the algorithm for determining response variances. (Right panel): Top portion shows maximal CMAP and envelope of responses evoked by sets of 30 stimuli at a constant intensity to determine variance in zone 4. Middle plot shows response in histogram format to determine whether distribution of responses is appropriately skewed to the left. Lower chart shows calculated variance of the response to sets of 30 stimuli. The variance is equal to the mean change in response amplitude, which is considered to reflect the average S-MUP amplitude for that region. A corresponding MUNE value is calculated. Three sets (groups) of 30 stimuli were required for the standard error to be < 10% of the mean value (96 ± 7). (Lower left panel): Results of testing all 4 regions (runs). An MUNE value is calculated for each region and summed to give an overall MUNE value for the total percentage of the CMAP tested (23%). The untested percentage (77%) is assumed to be innervated by the smallest S-MUPs, and an MUNE value is calculated accordingly. Tested and untested regions are summed for a final MUNE value of 131. An alternative method is to weigh the tested regions to determine a S-MUP value (waSMUP). This value is used to calculate and estimated MUNE value. See text for differences in final MUNE values by the different techniques.

different combinations influences on the final MUNE calculation, and MUNE values from the same normal subject that can vary by > 30% (Shefner et al., 1999; Lomen-Hoerth and Olney, 2001; Daube, 2003). 12.5.3.5. Modifications (1) A number of modifications have been proposed to determine the specific regions of the scan curve, which should be tested and the size of the tested regions (Olney et al., 2000; Lomen-Hoerth and Olney, 2001). The proprietary software automatically selects the regions to be tested, and each region includes about 6% of the total CMAP amplitude. The software also allows the operator to determine those regions. One paradigm uses four set regions of 15–25% of the

maximal response, 25–35%, 35–45%, and 45–55% (Bromberg, 2003a). (2) There are a number of different methods to calculate average S-MUP values that are used to determine the final MUNE. One source of variability in the final MUNE value is reliance on the smallest S-MUP value to calculate the MUNE for the untested regions of the CMAP. A modification weights the S-MUP values obtained from the tested regions to obtain a more representative average S-MUP value (Fig. 12.5) (Shefner et al., 1999). Note that the weighted average technique relies on the metric of negative peak amplitude while the conventional technique relies on negative peak area. A consensus for performing the statistical MUNE technique is available (Bromberg, 2003a).

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12.5.4. Spike triggered averaging motor unit estimation number technique 12.5.4.1. Principles The incremental, multiple point stimulation and statistical MUNE techniques rely on electrical stimulation of the nerve to activate S-MUPs. An alternative method to activate single motors is by voluntary contraction of the muscle (Brown et al., 1988). Individual motor units are identified from weak interference patterns and used to trigger a signal averager to extract the surface representation of the motor unit.

MARK B BROMBERG

T r c 1

500 μV

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12.5.4.3. Advantages (1) The intramuscular electrode can also be used to obtain quantitative motor unit action potential (MUAP) data, such as amplitude, duration, number of phases and turns. (2) This technique can be applied to proximal muscles because only limited access to the nerve is required to generate a maximal CMAP, whereas longer lengths of nerve are required for the controlled stimulation protocols with the other techniques. 12.5.4.4. Issues and Pitfalls (1) Care must be taken to avoid spurious trigger potentials (Bromberg and Abrams, 1995). (2) There is concern for a sampling bias to early recruited motor units. However, comparison studies between different MUNE techniques indicate no significant bias (Stein and Yang, 1990; Doherty and Brown, 1993). 12.5.4.5. Modifications In the conventional spike triggered averaging technique, single intramuscular potentials are isolated.

5 ms 500 μV

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12.5.4.2. Methods Two recording channels are used, one for surface recording and the other for intramuscular electrode recording. Surface electrodes are placed and a maximal CMAP is recorded. A weak interference pattern is generated and the intramuscular electrode is adjusted to isolate the discharge of one motor unit (Fig. 12.7). A voltage level trigger is set to detect the motor unit’s discharge pattern, and the trigger signals from that motor unit are used for spike triggered averaging of the surface response of the motor unit. The electrode is moved to another site in the muscle and another S-MUP is obtained. Ten to fifteen responses are collected and averaged to obtain the average S-MUP used to calculate the MUNE value.

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Fig. 12.7 Determ