Chapter 45. Electrophysiologic and Laboratory Aids in the Diagnosis of Neuromuscular Disease

The clinical suspicion of neuromuscular disease, disclosed by any of the symptoms or syndromes in the succeeding chapters, finds ready confirmation in the laboratory. The intelligent selection of ancillary examinations requires knowledge of the biochemistry and physiology of nerve action potentials, neuromuscular transmission, and muscle fiber contraction. These basic subjects, with the relevant anatomy, serve as an introduction to the descriptions of the laboratory methods and the subject matter of the chapters on the diseases of muscle and nerve that follow.

Since the early studies of Hodgkin (1951) and of Hodgkin and Huxley (1952), tomes have been written on the subject of the conduction of electrical impulses in neural tissues. Such conduction in nerve and muscle depends first on the maintenance of a fluid internal environment that is distinctly different from the external or interstitial medium. The main intracellular constituents are potassium (K), magnesium (Mg), and phosphorus (P), whereas those outside the cell are sodium (Na), calcium (Ca), and chloride (Cl). In both nerve and muscle the intracellular concentrations of these ions are held within a narrow range by both passive electrical and active chemical forces, which maintain the membranes in an electrochemical equilibrium termed the resting membrane potential. These forces are the result of selective permeability of the membranes to various ions and to the continuous expulsion of intracellular Na through specific channels by a pump mechanism (the sodium pump). The function of the sodium pump is dependent on the enzyme Na-K-ATPase (adenosine triphosphatase), which is localized in the membranes.

This resting membrane potential is the result of the differential concentrations of K and Na. The interior of the cell is some 30 times richer in K than the extracellular fluid, and the concentration of Na is 10 to 12 times greater in the extracellular fluid. In the resting state, the chemical forces that promote diffusion of K ions out of the cell (down their concentration gradient) are counterbalanced by electrical forces, the internal negativity, that opposes further diffusion of K to the exterior of the cell. The situation of Na ions in the equilibrium state is the opposite; they tend to diffuse into the cell, both because of their concentration gradient and because of the relative negativity inside the cell. Because the membrane at rest is less permeable to Na than to K, the amount of K leaving the cell exceeds the amount of Na entering the cell, thus creating the difference in electrical charge across the membrane. This discrepancy creates an electrical potential across the membrane such that, in the resting state, the intracellular compartment is 70 to 90 mV negative relative to the extracellular space (the resting membrane potential). The actions of the Na:K pump and the presence of impermeant, negatively charged intracellular proteins, contribute to maintaining this negative potential.

From this resting potential, any electrical discharge of neural and muscular tissue is predicated on the special property of excitable membranes; namely, the permeability of the cell to Na is linked to the electrical potential across the membrane by voltage-gated (i.e., voltage sensitive or voltage triggered) ion channels. Any depolarization of the membrane by a slight electrical or chemical change, creates an increased permeability to Na through the sodium channels and a consequent inward movement of sodium. Subsequently, K moves outward and repolarizes the membrane and thereby reduces its permeability to Na. If a greater degree of depolarization occurs, a situation arises in which the outward movement of K is unable to stabilize the membrane potential. The membrane then becomes even more depolarized and progressively more permeable to Na, and an "explosive," or regenerative Na current develops in which Na rushes down its chemical and electrical gradients into the cell. With the elevation in Na permeability, the Na ions continue to depolarize the intracellular compartment to about +40 mV (the Nernst equilibrium potential for Na). It is this depolarization of the intracellular compartment, lasting a few milliseconds, that constitutes the action potential.

However, an equilibrium at this level is not established because there is a rapid (within 5 ms) fall in membrane Na permeability back to baseline levels. This fall in Na permeability occurs because the depolarization inactivates Na conductance. The transient depolarization at the same time activates an increase in membrane permeability to K. The combined effects of the resulting efflux of K and a diminishing influx of Na repolarizes the membrane back to its resting level. During a brief period immediately after repolarization, the nerve and muscle fibers are refractory, at first absolutely then relatively, to another depolarizing stimulus. The duration of the refractory period is determined largely by the duration of the inactivation of the Na channel.

Conduction, or propagation, of the action potential along nerve or muscle, occurs as the current created by depolarization of a small segment of membrane flows into the contiguous membrane, which, in turn, becomes depolarized. When the depolarization reaches the threshold for development of an action potential, a new zone of increased Na permeability is created. In this manner, the action potential spreads in an "all-or-none" fashion, down the length of the nerve or muscle membrane. The action potentials in individual nerve fibers are too small and brief to be detected by conventional nerve conduction techniques, but a summated volley of all the fibers within a nerve is large enough to be recorded, and it is this electrical potential and the one it produces in the muscle that it innervates that is utilized to study nerve function.

As nerve impulses pass centrifugally from the axon into its terminal branches, transmission may "break down," especially if impulses arrive too frequently at branch points; they then fail to reach the neuromuscular junction of these fibers; or there may be failure of conduction at the neuromuscular junction, which is the characteristic feature of myasthenia gravis.

In large motor and sensory nerves, contiguous spread of action potentials along a fiber eventually decays over long distances. Conduction is aided by the structure and configuration of the myelin sheaths surrounding the axon. The sodium channels, which generate the action potential, are concentrated at short exposed segments of the axon, the nodes of Ranvier, lying between longer segments of myelinated axon, the internodes. In contrast, the internodes covered by myelin remain electrically insulated. This creates "flux lines" that converge on the nodes and allows current to be regenerated at each gap in the myelin. The speed of electrical conduction, which jumps in a "saltatory" fashion from node to node, is many times faster than conduction through an unmyelinated axon. The largest-diameter fibers have the thickest myelin sheaths and longest internodal distances, conferring on them the fastest conduction times.

Conventional laboratory studies of nerve conduction generally measure the speed of these fastest-conducting fibers. This dependence of nerve conduction on large, heavily myelinated fibers explains a number of electrophysiologic abnormalities that are a consequence of nerve disease. One common result, is a mild slowing of nerve conduction because of a dependence on the remaining smaller diameter, slower conducting axons. When myelin destruction is the prominent feature of a neuropathy (demyelinating neuropathy), conduction velocity is greatly slowed because of delays in regenerating the sodium current at nodes of Ranvier, or there may be total block of electrical conduction. An intermediate state of partial demyelination slows and desynchronizes the electrical volley, leading to temporal dispersion of the action potentials that reach the muscle. The cumulative effect of any of these changes is to reduce the number of nerve fibers that are capable of conducting an electrical volley, leading to a graduated reduction in the amplitude of the muscle action potential over longer segments of nerve. This reduction in amplitude of the compound motor action potential (CMAP) as the stimulating electrode is moved proximally is termed conduction block (discussed under "Studies of Nerve Conduction" further on). Blocked conduction of this restricted nature is a reliable marker of an acquired demyelinating neuropathy; of all the electrophysiologic changes, it also corresponds most closely to the degree of muscle weakness (see further on).

In contrast to these effects of demyelination of peripheral nerve, loss of axon fibers (axonal neuropathy) results in a reduction of the amplitude of summed electrical activity of the action potential in muscles uniformly all along the nerve, and to atrophic denervation of muscle as described further on.

This is the interface between the finely branched nerve fiber and the muscle fiber, where the electrical activity of the motor nerve is translated into muscle action (Fig. 45-1). The nerve fiber contacts the muscle membrane in a trough-like junctional space of 50 mm—the synaptic cleft—between the axolemma and sarcolemma (see Fig. 49-1). Within the nerve terminal, a relatively fixed number of packets, or quanta, of acetylcholine (ACh), each packet containing about 10,000 molecules, are liberated through an exocytotic process that is triggered by the arrival of axonal action potentials. Arrival of the electrical impulse opens calcium channels in the presynaptic neural membrane, which serves to bind packets of ACh to the membrane and govern their release. Molecules of ACh diffuse into the synaptic cleft and attach to receptor sites on the postsynaptic membrane. Each impulse triggers the release of approximately 20 to 50 quanta of ACh and produces a depolarization of sufficient size to initiate an action potential in the postsynaptic muscle membrane through the same mechanism of a regenerative sodium current described earlier. Botulinum toxin interferes with exocytosis of ACh at the release site by cleaving "synaptic protein 25." There is also a nonquantal release of ACh through continuous leakage. This appears to play a role in the trophic influence of nerve on muscle.

The ACh molecule binds to the postsynaptic ACh receptor, a complex of 5 proteins that constitute the postsynaptic ion channel. Binding of ACh by this receptor causes a conformational change in the receptor that leads to a local increase in the conductance of Na and K and other small ions. This produces a depolarization known as the endplate potential. Small (miniature) endplate potentials (MEPPs) are continuously formed and regenerated as the membranes repolarize, much as in the process of passive decay previously described. These potentials are too small to be recorded by routine electromyography (EMG) testing or to trigger an action potential, although fortuitous needle placement adjacent to a synapse may detect them.

The bound ACh is hydrolyzed by cholinesterase, a glycoprotein enzyme that exists in free form in the neuromuscular junction clefts. Its main function is to terminate the action potential and permit the sequential activation of muscle. The postsynaptic membrane, once depolarized, is refractory to another action potential until it is repolarized.

The calcium that entered the presynaptic nerve terminal is sequestered and then extruded, and the choline from the hydrolyzed ACh enters the nerve terminal, where it is resynthesized to ACh near the release sites.

The analysis of a rapid series of electrically of voluntarily elicited muscle contractions is used to test the function of the neuromuscular junction by stressing conduction across the junction. In general, a decrement in the amplitude of serial muscle action potentials is typical of postsynaptic failure, and an increment in the amplitude from a train of stimuli is a reflection of presynaptic failure.

Myasthenia gravis is the principal disease affecting the neuromuscular junction and represents a failure of postsynaptic function (see Chap. 49). In this process, the fundamental defect is not a deficiency of ACh or its release, but rather a failure of ACh to attach to the postsynaptic receptor, as a result of blockade by an antibody at the receptor site or of destruction of the membrane by antibody attack. There are several quite different synaptic disorders caused by botulism, aminoglycoside antibiotics, and the antibodies of the Lambert-Eaton myasthenic syndrome, which impede presynaptic release of ACh.

A number of pharmacologic agents also interfere with neuromuscular transmission by combining with the cholinergic (nicotinic) receptor on the postsynaptic membrane, thereby competitively blocking the transmitter action of ACh. The curariform drugs, derived from curare and termed nondepolarizing neuromuscular blockers because they do not alter the postsynaptic membrane potential are the main examples. Other drugs, notably succinylcholine and decamethonium, cause neuromuscular blockage by producing direct depolarization of the endplate and adjacent sarcoplasmic membrane (depolarizing neuromuscular blockers). Agents that inactivate cholinesterase have the opposite effect, i.e., they enhance the action of ACh. The ones in clinical use for the treatment of myasthenia gravis are the carbamates neostigmine, physostigmine, and pyridostigmine, the effects of which are reversible. The organophosphates are irreversible blockers of cholinesterase function, for which reason they are feared weapons of chemical warfare. Because the potent cholinergic antagonist atropine is active only at muscarinic sites, it has no effect at the neuromuscular junction.

The sarcolemma, the transverse tubules, and the sarcoplasmic reticulum each play a role in the control of the activity of muscle fibers. Figure 45-2 illustrates the structural components involved in excitation, contraction, and relaxation of muscle. Following nerve stimulation, an action potential is transmitted by the sarcolemma from the motor endplate region to both ends of the muscle fiber. Depolarization spreads quickly to the interior of the fiber along the walls of the transverse tubules, probably by a conducted action potential. The transverse tubules and the terminal cisternae of the sarcoplasmic reticulum come into close proximity at points referred to as triads. Here, depolarization of the transverse tubules alters the conformation of a voltage-sensitive calcium channel in the transverse tubule membrane. This causes a large protein (the ryanodine receptor) in the membrane of the adjacent sarcoplasmic reticulum (SR) to open an internal calcium pore, allowing stored calcium to exit the SR and flood into the cytoplasm. The released calcium binds to the regulatory protein troponin, thereby removing the inhibition exerted by the troponin–tropomyosin system upon the contractile protein actin. This allows an interaction to occur between the actin molecules of the thin filaments and the cross-bridges of the myosin molecules in the thick filaments and enables myosin ATPase to split adenosine triphosphate (ATP) at a rapid rate, thereby providing the energy for contraction. This chemical change allows the filaments to slide past each other, thereby shortening the muscle fiber. Relaxation occurs as a result of active (energy-dependent) Ca reuptake by the sarcoplasmic reticulum.

The pyrophosphate bonds of ATP, which supply the energy for muscle contraction, must be replenished constantly by a reaction that involves interchanges with the muscle phosphagen creatine diphosphate, where high-energy phosphate bonds are stored. These interactions, in both contraction and relaxation, require the action of creatine kinase (CK).

Myoglobin, another important muscle protein, functions in the transfer of oxygen, and a series of oxidative enzymes are involved in this exchange. The intracellular Ca, as noted earlier, is released by the muscle action potential and must be reaccumulated within the cisternae of the sarcoplasmic reticulum before actin and myosin filaments can slide back past one another in relaxation. This reuptake of Ca requires the expenditure of considerable energy. When there is defective generation of ATP, for example, from an enzyme deficiency, the muscle remains shortened, as in the contracture of phosphorylase deficiency (McArdle disease) or of phosphofructose kinase deficiency. The same sort of shortening contracture occurs under normal conditions in some of the "catch muscles" of certain mollusks and is the basis of rigor mortis in mammals.

Many glycolytic and other enzymes (transaminases, aldolase, CK) are utilized in the metabolic activity of muscle, particularly under relatively anaerobic conditions. Muscle fibers differ from one another in their relative content of oxidative and glycolytic enzymes; the latter determine the capacity of the muscle fiber to sustain anaerobic metabolism during periods of contraction with inadequate blood flow. Muscle cells rich in oxidative enzymes (type 1 fibers) contain more mitochondria and larger amounts of myoglobin (therefore appearing red), have slower rates of contraction and relaxation, fire more tonically, and are less fatigable than muscle fibers poor in oxidative enzymes. The latter (type 2 fibers) fire in bursts and are used in quick phasic, rather than sustained, reactions. The amount of myosin ATPase activity, which governs the speed of contraction, is low in oxidative-rich fibers and high in glycolytic-rich fibers. The myosin ATPase stain at pH 9.4 has been used to differentiate these two types of fibers in microscopic sections. Type 1 fibers have a low content of myosin ATPase, and type 2 (phosphorylative-rich) fibers have a high content of this enzyme; hence type 1 fibers stain lightly and type 2 darkly (the reverse reaction occurs at pH 4.6). All the fibers within one motor unit are of the same type, a feature that is used to advantage to identify the reinnervation of muscle fibers by a single motor neuron after adjacent neurons have died and denervated their constituent muscle fibers (fiber-type grouping).

The chemical energy required to maintain the various activities of the muscle cell is derived mainly from the metabolism of carbohydrate (blood glucose, muscle glycogen) and from fatty acids (plasma-free fatty acids, esterified fatty acids, and ketone bodies). There is a lesser contribution to energy from branched-chain and other amino acids, but their activity increases during prolonged exercise.

The most readily available source of muscular energy is glycogen, which is synthesized and stored in muscle cells. It provides more than 90 percent of the energy needs of muscle under conditions of high work intensity and during the early stages of submaximal exercise. Blood glucose and free fatty acids supplement intracellular glycogen as exercise proceeds. The free fatty acids are obtained from endogenous triglycerides (found mostly in type 1 fibers), from the triglycerides released by circulating lipoproteins, and from the lipolysis of adipose tissue. Most of the energy needs of resting muscle are provided by fatty acids.

The enzymatic reactions involved in the transport of these substrates into muscle cells and their intracellular synthesis and degradation during anaerobic and aerobic cell conditions have been thoroughly investigated and most of the participating enzymes have been identified. This subject is too extensive to be presented in a textbook of neurology, but enough is known about these matters to state that there are diseases that impair the contractile functions of muscle without destroying the fiber. In particular, specific enzymatic deficiencies alter carbohydrate utilization (myophosphorylase, debrancher enzyme, phosphofructokinase, and phosphoglyceromutase deficiencies), fatty acid utilization (carnitine and carnitine palmitoyl transferase deficiencies), pyruvate metabolism, and cytochrome oxidase activity (in the mitochondrial muscle diseases). These diseases are discussed in later chapters.

The contraction of a muscle fiber may be viewed as a series of electrochemical events that culminate in a mechanical effect. The mechanical change far outlasts the electrical one and extends through the period when the muscle fiber is refractory to another action potential. When a second muscle action potential arrives after the refractory phase of the previous action potential, but before the muscle has relaxed, the contraction will be prolonged. Thus, at frequencies of anterior horn cell firing of more than 100 per second, twitches fuse into a sustained voluntary contraction or fused tetanus. In most sustained contractions, there is incomplete tetanus, attained by firing rates of 40 to 50 per second. In this fashion, the mechanical contractions of individual muscle fibers are smoothed into a continuous process, even though the electrical potentials present as a series of depolarizations, separated by intervals during which the muscle membrane resumes its resting polarized state.

As pointed out in Chap. 3, the physiology of muscle activity is best considered in terms of motor units, i.e., the group of muscle fibers innervated by a single anterior horn cell. The strength of muscle contraction is in turn a function of the number and rates of firing of many adjacent motor units. The smoothness of contraction depends on the integrated and sequential enlistment of motor units of increasing size. The electrical signal of this summated contraction, the compound motor action potential, or CMAP, as recorded at the skin surface overlying a muscle is the main feature of the surface EMG and serves as the basis for quantifying the size of the motor nerve potential in the nerve conduction examinations.

The CMAP can be visualized on the screen of an oscilloscope or a computer and its onset used to measure the speed of motor nerve conduction (conduction velocity) as well as the summated amplitudes produced by all the innervating nerve fibers. It can also be converted into an audible noise. Further study involves the insertion into the muscle of a coaxial needle, which samples several motor units in the vicinity of the electrode. When elicited by a sustained voluntary contraction, the flurry of electrical activity from many muscle fibers at different distances from the electrodes is referred to as an interference pattern (see further on, under "Studies of Nerve Conduction" and "Needle Examination of Muscle [Electromyography]").

Various biochemical changes may cause not only an impairment of muscular activity (paresis, paralysis) but also excessive irritability, tetany, spasm, and cramp. In these instances, spontaneous discharges may occur from instability of axon polarization; hence a single nerve impulse may initiate a train of action potentials in nerve and muscle, as in the tetany of hypocalcemia. The common cramps of calf and foot muscles (painful, sustained contractions with motor unit discharges at frequencies up to 200 per second) may be a result of increased excitability (or unstable polarization) of the motor axons. Quinine, procainamide, diphenhydramine, and warmth reduce the irritability of nerve and muscle fiber membranes, as do a number of antiepileptic drugs that act by blocking sodium channels, thereby limiting spontaneous membrane discharges.

The muscle fiber, which is wholly dependent on the nerve for its stimulus to contract, may be physiologically activated or paralyzed in a number of ways that are explained by the principles described above. The main disturbances of nerve, muscle, and neuromuscular function occur when the motor nerve cell or its axon is injured or inexcitable, in which case the muscle cannot be stimulated; or, the nerve cell in the anterior horn of the spinal cord may be disinhibited, permitting the discharge of continuous action potentials, as in tetanus and the "stiff man" syndrome (see Chap. 48); the nerve fiber may fail to conduct impulses (demyelinating neuropathies) or the number of fibers may be inadequate to produce a fused and sustained contraction (axonal neuropathies); the distal axon may not distribute the nerve impulse simultaneously to all parts of the motor unit; ACh may not be released at the presynaptic region of the neuromuscular junction (as occurs in botulism and in the Eaton-Lambert syndrome) or, once released, ACh may not be inactivated by cholinesterase (physostigmine, organophosphates); the receptor zone on the postsynaptic membrane may be destroyed or blocked by antibodies or pharmacologic agents (myasthenia gravis or curariform drugs); and, finally, the metabolic or contractile elements of the muscle may not react or, once contracted, may not relax. One type of this last category is due to genetically determined abnormalities of voltage-gated ion channels, the "channelopathies," that are the subject of Chap. 50.

Similarly, there may be an unstable polarization of the nerve fibers, as in tetany and in dehydration with salt depletion, or hyperirritability of the motor unit, as in amyotrophic lateral sclerosis. The threshold of mechanical activation or electrical reactivation of the sarcolemmal membrane may be reduced, as occurs in myotonia, or impairment of an energy mechanism within the fiber may slow the contractile process, as in hypothyroidism; or a deficiency of phosphorylase, which deprives muscle of its carbohydrate energy source, may prevent relaxation, as in the contracture of McArdle disease. Lesions of the most peripheral branches of nerves, which allow nerve regeneration, may give rise to continuous activity of motor units. This is expressed clinically as a rippling of muscle, or myokymia.

In recent years, special techniques have made it possible to study each of the proteins and channels involved in neuromuscular transmission and the excitation-contraction-relaxation of muscle fibers. The amino acid composition of these proteins has also been determined. This information is being increasingly applied to the analysis of genetic diseases of muscle in the normal state and under conditions of disease. Pertinent comments and references to this subject are found in the chapters that follow.

Diffuse muscle weakness or muscle twitching, spasms, and cramps should always raise the question of uremia or an abnormality in serum electrolytes. These disorders reflect the concentrations of electrolytes in the intra- and extracellular fluids. If the plasma concentration of K falls below 2.5 mEq/L or rises above 7 mEq/L, weakness of extremity and trunk muscles results; below 2 mEq/L or above 9 mEq/L, there is almost always flaccid paralysis of these muscles and later, of the respiratory ones as well; only the extraocular and other cranial muscles are spared. In addition, the tendon reflexes are diminished or absent. The normal reaction of muscle to direct percussion is also reduced or abolished, suggesting impairment of transmission along the sarcolemmal membranes themselves. Hypocalcemia of 7 mg/dL or less (as in rickets or hypoparathyroidism) or relative reduction in the proportion of ionized calcium (as in hyperventilation) causes increased muscle irritability and spontaneous discharge of sensory and motor nerve fibers (i.e., tetany) and sometimes convulsions from similar effects upon cerebral neurons; frequent repetitive and prolonged spontaneous discharges grouped in couplets or triplets appear in the EMG. Hypercalcemia with Ca levels above 12 mg/dL (as occurs in vitamin D intoxication, hyperparathyroidism, carcinomatosis, sarcoidosis, and multiple myeloma) causes weakness perhaps on a central basis, and lethargy. Extreme hypophosphatemia, observed most often with intravenous hyperalimentation or bone tumor, can cause acute areflexic paralysis with nerve conduction abnormalities. Reduction in the plasma concentration of magnesium also results in tremor, muscle weakness, tetanic muscle spasms, and convulsions; a considerable increase in magnesium levels leads to muscle weakness and depression of central nervous function. The weakness of muscle in hypermagnesemia may also be partly a result of reduced release of ACh at the motor endplate.

The electrocardiogram (ECG) also becomes abnormal with many of these systemic electrolyte derangements as a result of alterations of the intracellular levels of electrolytes in the myocardium; ECG patterns are most sensitive to extreme changes in serum K levels.

In all diseases that cause extensive damage to striated muscle fibers, intracellular enzymes leak out of the fibers and enter the blood. Those measured routinely are the transaminases, lactic acid dehydrogenase, aldolase, and, especially, CK. The concentration of CK in serum has proved to be the most consistent and most sensitive measure of skeletal muscle damage. Because high concentrations of this enzyme are found in heart muscle and brain, raised serum values may be a result of myocardial or cerebral infarction, as well as of the necrotizing diseases of striated muscle (polymyositis, muscle trauma, muscle infarction, and the more rapidly advancing muscular dystrophies). For serum CK levels to be interpretable, one has to be certain that the enzyme released into the serum is not derived from heart or brain. This can be determined by the quantitation of serum isoenzymes of CK, referred to as MB, MM, and BB (M, muscle; B, brain); their measurement provides a means for the detection of damage to myocardium (MB), skeletal muscle (MM), and nervous tissue (BB), respectively.

The MM form of CK is found in highest concentration in striated muscle, but these muscles also contain 5 to 6 percent MB isoform. Myocardial tissue contains 17 to 59 percent MB; hence, the diagnosis of myocardial infarction requires that the CK-MB fraction be greater than 6 percent (or that troponin, which is overrepresented in heart muscle, be elevated in the serum). Embryonic and regenerating muscle contains more CK-MB than mature normal muscle. In patients with destructive lesions of striated muscle, serum values of CK often exceed 1,000 U/L and may reach 50,000 U or more (the upper limit of normal varies from 65 to 300 U/L, depending on the method of measurement). It is notable that in some black men the level may normally be in the range of 500 U/L in the absence of muscle or nerve disease (see below). The serum of the healthy adult contains only the MM isoenzyme, but in healthy children, as much as 25 percent of serum CK may be derived from the MB fraction.

As interesting is the rise in serum enzyme concentration in some children with progressive muscular dystrophy before there is enough destruction of fibers for the disease to be clinically manifest, at least as judged by the relatively crude test of muscle strength. Moreover, the unaffected female carrier of Duchenne dystrophy may be identified by an elevated serum level of CK; these changes reflect a "leaky" membrane that allows the transgression of the large enzyme molecules. Alterations of serum enzyme levels are, however, nonspecific as they occur in all types of disease that damage the muscle fiber. Moreover, in the more slowly evolving types of dystrophy, the serum levels of CK may be normal.

It would be expected that enzyme values would be normal in paralysis due to denervation with secondary muscular atrophy, but elevations are sometimes observed in patients with progressive spinal muscular atrophy and amyotrophic lateral sclerosis, particularly if there is relatively rapid progression of the disease. Even vigorous exercise or surgical operations involving muscle elevate CK, sometimes with the MB fraction exceeding 6 percent.

Idiopathic "HyperCKemia"

In some normal individuals, CK may be persistently elevated without evidence of muscle or other disease. There is a particularly high incidence of this finding, albeit of mild degree, in African American men. An unexplained alteration of the sarcolemma with elevated serum CK occurs in hypothyroidism and in alcoholism. Toxic myopathies, for example, the type caused by the cholesterol-lowering drugs (statins), are another common cause of elevation in CK in current practice. Among the causes of persistently elevated CK are numerous underlying genetically determined muscle defects, usually one of the varieties of abnormal dystrophin or another membrane protein. Detailed examination may reveal slight proximal weakness and perhaps minimal calf enlargement characteristic of the mildest forms of Becker dystrophy. Closely related is the finding of CK elevations that occur in females who are asymptomatic carriers of Duchenne or Becker dystrophies. These diseases are described in Chap. 48.

Several nondystrophic muscle diseases also present as elevated CK levels in the blood, including polymyositis and dermatomyositis, carnitine palmityl transferase deficiency, some of the mitochondrial myopathies, McArdle disease, central core disease, multicore disease, and some forms of inclusion body myopathy. Finally, the rapid denervation of muscle that accompanies some cases of ALS may cause CK elevation.

If other sources of persistently elevated CK levels have been excluded, particularly those caused by exercise, trauma, and by drug-induced muscle damage, it is appropriate to follow the patient over time so that mild weakness may be detected, or to perform a biopsy to resolve the issue of an inflammatory myopathy or a mild muscular dystrophy. Because treatment would likely be withheld until weakness or pain arises, in idiopathic cases of raised enzyme levels, it seems prudent to delay biopsy unless the enzyme levels are massively elevated.

An isolated elevation of aldolase, the serum enzyme other than CK that is derived predominantly from skeletal muscle, generally has less clinical significance. Measurement in the serum of various transaminases or lactate dehydrogenase is not particularly useful for the diagnosis of muscle disease because of the ubiquitous distribution of these enzymes in many mammalian tissues. Nevertheless, the neurologist should be aware that unexplained elevations in all of the muscle-derived enzymes (CK, lactate dehydrogenase [LDH], serum glutamic-oxaloacetic transaminase [SGOT], aldolase) can be caused by inevident muscle trauma and by many other processes that damage the muscle membrane. As mentioned, cardiac muscle-specific enzyme, troponin, is not expressed in skeletal muscle and therefore is not found in the serum in cases of muscle disease, except in unusual circumstances in which regenerating muscle fibers in muscular dystrophies may transiently express the enzyme (the T, but the I, isotype) as noted in the series reported by Jaffe and colleagues. More often, elevations or troponin in these muscular diseases reflects a parallel myocardial disorder that accompanies several varieties of dystrophy.

The red pigment myoglobin, responsible for much of the color of muscle, is an iron-protein compound present in the sarcoplasm of striated skeletal and cardiac fibers. Of the hematin compounds, approximately 25 percent are in muscle and the remainder are in red blood corpuscles and other cells. Destruction of striated muscle, regardless of the cause liberates myoglobin, and because of its relatively small size, the molecule filters through the glomeruli and appears in the urine, imparting to it a burgundy-red color. Because of the low renal threshold for myoglobin, excretion of the pigment is so rapid that the serum remains uncolored. In contrast, the high renal threshold for hemoglobin colors both the serum and the urine if there is destruction of red blood corpuscles. Myoglobinuria should thus be suspected when the urine is deep red and the serum is normal in color. It is estimated that 200 g of muscle must be destroyed to color the urine visibly (Rowland).

As with hemoglobinuria, the guaiac and benzidine tests performed on urine are positive if myoglobin is present. However, the colored urine does not fluoresce, as it does in porphyria. On spectroscopic analysis, myoglobin shows an absorption band at 581 nm but the most sensitive method for measuring myoglobin in the urine and serum is by radioimmunoassay. Some years ago, the measurement of creatine and creatinine in blood and urine was a standard method of estimating damage to striated muscle. This technique is now seldom used, having been replaced by the measurement of CK and its isoenzymes. However, measurement of urine creatinine in large-volume samples of urine that have been collected for other testing assures that an adequate amount of renal excretion is being captured in the sample.

In a number of disorders of the endocrine glands, muscle weakness may be a prominent feature, and occasionally may be the chief complaint. These diseases are discussed in detail in Chap. 48 but it should be noted here that such metabolic causes of muscle weakness, acute or chronic, may occur in the absence of changes in serum electrolytes or enzymes. Specific hormone assays are then necessary for diagnosis. This is particularly true of patients with thyrotoxicosis or Cushing disease, and of those receiving prolonged corticosteroid therapy. In thyrotoxicosis, muscle weakness may appear without the signs of Graves disease.

It was long ago discovered that muscle would contract when a pulse of electric current was applied to the skin, near the point of entrance of the muscular nerve (the motor point). The electrical pulse required is brief, less than a millisecond, and is most effectively induced by rapidly alternating (faradic) current. If there has been muscle denervation, an electrical pulse of several milliseconds induced by a constant electrical (galvanic) stimulus is required to produce the same response. This change, in which the galvanic stimulus remains effective after the faradic one has failed, was the basis of the "Erb reaction" of degeneration, and varying degrees of this change were formerly plotted in the form of strength-duration curves. For decades, this was the standard electrical method for evaluating denervation of muscle. Although still valid, it was replaced by nerve conduction studies and by the needle electrode examination. The latter test, based on the sherringtonian concept of the "motor unit" described in Chap. 3, is accomplished by the insertion into muscle of needle electrodes to measure spontaneous and voluntarily evoked muscle fiber activity. The terms electromyography and electromyogram were used originally to describe the needle electrode examination but are now a common shorthand designation for the entire electrodiagnostic evaluation, including the nerve conduction studies.

Studies of Nerve Conduction

The main laboratory technique for the study of peripheral nerve function involves the transcutaneous stimulation of motor or sensory nerves and recording of the elicited action potentials in the muscle (CMAP) and the sensory nerve action potential (SNAP). The results of these motor and sensory nerve conduction studies, expressed as amplitudes, conduction velocities, and distal latencies, yield certain quantitative information and additional qualitative observations regarding the waveform and dispersion of electrical neural and muscular impulses.

Hodes and coworkers, in 1948, were the first to describe nerve conduction studies in patients, and the techniques used currently are not much changed. An accessible nerve is stimulated through the skin by surface electrodes, using a stimulus that is large enough to recruit (cause a discharge in) all the available nerve fibers. The resulting action potential is recorded by electrodes on the skin (1) over the muscle distally in the case of motor fibers stimulated in a mixed or motor nerve (CMAP), (2) over the nerve more distally, using antidromic techniques for sensory nerve conduction studies (this has technical advantages over orthodromic techniques), and (3) over the nerve more proximally for mixed (sensory and motor) nerve conduction studies (Fig. 45-3). These techniques are the ones used most often in clinical work. An alternative but much more demanding technique uses "near-nerve" needle electrodes to record action potentials as they course through the nerve. The main characteristics of the conventional nerve conduction studies are described below.

Distal (Terminal) Latencies, Conduction Times, and Conduction Velocities

The conduction times from the most distal stimulating electrode to the recording site over a muscle, in milliseconds, as determined by the latency from the stimulus artifact to the onset and to the peak of the CMAP, are termed the distal (or terminal) and peak motor latencies, respectively (see Fig. 45-3). The former is the one used more often as a reflection of conduction time in routine work. A stimulus may then be applied to the nerve at a second site more proximally (or if recording electrodes can be placed more proximally in the case of sensory fibers), and a conduction time can be measured over a longer segment of nerve. When the distance (in millimeters) between the two sites of stimulation is divided by the difference in conduction times (in milliseconds), one obtains a conduction velocity (in meters per second), which describes the maximal velocity of propagation of the action potentials in the largest-diameter and fastest-conducting nerve fibers. These velocities in normal subjects vary from a minimum of 40 or 45 m/s to a maximum of 65 to 75 m/s, depending upon which nerve is studied (e.g., slower in the legs than in the arms; Table 45-1). Values are lower in infants, reaching the adult range by the age of 2 to 4 years, and declining again slightly with advancing age. Conduction velocity also is diminished with exposure to cold, a potentially important factor if these recordings are taken when the patient's skin is cool; consequently, measurement of skin temperature is routinely done prior to performing the nerve conduction tests.

Normal values have been established for distal latencies from the usual sites of stimulation on various mixed nerves to the appropriate muscles. Stimulating the median nerve at the wrist, for example (see electrode 1 and segment A in Fig. 45-3), has a latency for motor conduction through the carpal tunnel to the median-innervated thenar muscles of less than approximately 4.5 ms in healthy adults. Similar normal values have been compiled for orthodromic and antidromic sensory conduction velocities and for distal latencies in all the main peripheral nerves (see Table 45-1).

Disease processes that preferentially injure the fastest-conducting, large-diameter fibers in peripheral nerves reduce the maximal conduction velocity because the remaining thinner fibers conduct more slowly. In most neuropathies, all of the axons are affected either by a fairly uniform "dying-back" phenomenon or by wallerian degeneration as described in Chap. 46, and nerve conduction velocities are then less informative. This is true, for example, in typical alcoholic-nutritional, carcinomatous, uremic, diabetic, and other metabolic neuropathies, in which conduction velocities range from the low-normal range to mildly slowed. In these "axonal neuropathies," the motor and sensory nerve amplitudes are diminished.

By contrast, demyelinating neuropathies of the acute (Guillain-Barré) and chronic types, such as chronic inflammatory, metachromatic leukodystrophy, and the common type of Charcot-Marie-Tooth disease, show marked slowing of conduction and, in the case of the acquired demyelinating diseases, there is also dispersion of the motor action potential and a highly characteristic conduction block (see later).

Amplitude of the Compound Muscle Action Potential

In addition to the study of distal latency and conduction velocity, the amplitude of the evoked muscle action potential (CMAP) yields valuable information about peripheral nerve function. These amplitudes are a semiquantitative measure of the number of nerve fibers that respond to a maximal stimulus (and of the innervated volume of muscle). Demyelinative lesions affecting the large, fast-conducting fibers are detected by the finding of differential slowing among various caliber fibers that causes a dispersal of the CMAP response. Reduction in motor and sensory amplitudes is a more specific and sensitive indicator of axonal loss than is slowing of conduction velocity or prolongation of distal latencies. Conversely, prolonged distal latencies and slowed motor conduction velocities, as well as conduction blocks and dispersed responses (described below), are the hallmarks of demyelinative lesions. Table 45-1 shows the range of normal amplitudes for the CMAPs that are elicited by stimulation of the main motor nerves.

It is usually possible to obtain a reliable motor conduction study as long as some functioning nerve fibers remain intact. The conduction velocities then reflect the status of the surviving axons, and the velocity may be normal or nearly so despite widespread axonal degeneration. This is most apparent following incomplete transection of a nerve; the maximal motor conduction velocity may be normal in the few remaining fibers, although the muscle involved is almost paralyzed and the compound muscle potential recorded from it is very low.

Sensory Nerve Action Potentials

When motor fibers in a mixed nerve are stimulated, an amplified CMAP of many hundreds of microvolts can easily be recorded from electrodes on the skin over the muscle. However, when one attempts to measure sensory potentials, where activity must be recorded from nerve fibers themselves, the "amplification" provided by many motor units is not available and electronic amplification is required. Sensory potentials are sometimes very small or absent even when powerful computer-averaging techniques are used, and sensory conduction measurements may then be difficult to determine. Table 45-1 gives the range of normal values for sensory nerve action potential amplitudes and velocities.

Conduction Block

By stimulating a motor nerve at multiple sites along its course, it is possible to demonstrate segments in which conduction is partially "blocked" or is differentially slowed. From such data one infers the presence of a multifocal demyelinative process in motor nerves. This contrasts with the findings in certain of the inherited and metabolic demyelinating neuropathies, in which all parts of the nerve fiber are altered to more or less the same degree, i.e., there is uniform slowing and reduction in amplitude and no conduction block.

As a technical matter, conduction block is demonstrated by a reduction in the amplitude of the CMAP elicited from the proximal site along the motor nerve, compared to stimulation at a distal site. Generally, a 40 percent reduction in amplitude over a short distance of nerve, or 50 percent over a longer distance, qualifies as a block, one possible exception being along the tibial nerve, in which there is some degree of physiologic dispersion (it is also difficult to stimulate all the motor nerve fibers of this nerve in obese patients); therefore a slight drop in amplitude over the length of the nerve is normally expected. It is important to be sure that any reduction in amplitude along the course of the nerve is not solely a result of dispersion of the waveform. The presence of a conduction block can also be inferred from the finding of poor recruitment of muscle action potentials and the concurrent absence of active denervation (see further on). The finding of conduction block is a main feature of a number of acquired immune demyelinating neuropathies including Guillain-Barré syndrome, chronic inflammatory demyelinating neuropathy, and multifocal conduction block associated with the GM1 antibody, which are discussed in Chap. 46.

Focal conduction block may be caused simply by nerve compression at certain common sites (fibular head, across the elbow, flexor retinaculum at the wrist, etc.) rather than to an intrinsic disease of the peripheral nerves. Focal compression of nerve, as occurs in these entrapment syndromes, produces localized slowing or blocks in conduction, perhaps because of segmental demyelination, only at the site of compression. The demonstration of such localized changes of conduction affords ready confirmation of nerve entrapment; for example, if the distal latency of the median nerve (see A, Fig. 45-3) exceeds 4.5 ms while that of the ulnar nerve remains normal, compression of the median nerve in the carpal tunnel is likely. Similar focal slowing or partial block of conduction may be recorded from the ulnar nerve at the elbow and from the peroneal nerve at the fibular head.

Electrodiagnostic Studies of Nerve Roots and Spinal Segments (Late Responses, Blink Responses, Evoked Responses)

H Reflex

Information about the conduction of impulses through the proximal segments of a nerve is provided by the study of the H reflex and the F wave. In 1918, Hoffmann, after whom the H reflex was named, showed that submaximal stimulation of mixed motor–sensory nerves, insufficient to produce a direct motor response, nonetheless induces a muscle contraction (H wave) after a latency that is far longer than that of the direct motor response. This reflex is based on the activation of afferent fibers from muscle spindles (the same axons that conduct the afferent volley of the tendon reflex), and the long delay reflects the cumulative time required for the impulses to reach the spinal cord via the sensory fibers, synapse with anterior horn cells, and to be transmitted along motor fibers to the muscle (see Fig. 3-1). Thus the H reflex is the electrical representation of the tendon reflex circuit and is a useful measure because the impulse traverses both the posterior and anterior spinal roots. The H reflex is particularly helpful in the diagnosis of S1 radiculopathy and of polyradiculopathies. Its status generally parallels that of the clinically elicited Achilles reflex. However, it is difficult to obtain an H reflex from nerves other than the tibial. Stimuli of increasing frequency but low intensity cause a progressive depression and finally obliteration of H waves. The latter phenomenon has been used to study spasticity, rigidity, and cerebellar ataxia, in which there are differences in the frequency-depression curves of H waves. In parallel with the Achilles tendon reflex, the H-reflex is transiently obliterated in spinal shock (see Chap. 44).

F Response (Wave)

The F response, so named because it was initially elicited in the feet, was first described by Magladery and McDougal in 1950. It is evoked by a supramaximal stimulus of a mixed motor–sensory nerve. After a latency that is longer than for the direct motor response (latencies of 28 to 32 ms in arms, 40 to 59 ms in legs), a second small muscle action potential is recorded. This F wave is the result of the impulses that travel antidromically in motor fibers to the anterior horn cells, a small number of which are activated and produce an orthodromic response that is recorded in a distal muscle. The F response is a more reliable test than the H wave of proximal motor nerve and root conduction in that the F wave traverses only the ventral root and can be elicited from a number of muscles. The combination of a normal F response and an absent H reflex is found in diseases of sensory nerves and roots. Both of these "late responses" find their main use as corroborative tests that are interpreted in the context of the entire nerve conduction examination (see Wilbourn). As with the H reflex, the F wave may be absent in the state of spinal shock (see Chap. 44). Table 45-1 gives the normal F wave response latencies.

Blink Responses

This special nerve conduction test is not in frequent clinical use but it serves a purpose in the diagnosis of certain demyelinating neuropathies and in any process that affects the trigeminal or facial nerve. The supraorbital (or infraorbital) nerve is stimulated transcutaneously and the reflex closure of both orbicularis oculi muscles is recorded with surface electrodes. Two CMAP bursts are observed: the first (R1) appears ipsilaterally 10 ms after the stimulus and the second (R2), ipsilaterally at 30 ms and contralaterally up to 5 ms later. The amplitudes of the responses vary considerably and are not in themselves clinically important. The first response is not visible as a muscular contraction but may serve some preparatory function by shortening the blink reflex delay. R1 is mediated by an oligosynaptic pontine circuit consisting of one to three neurons located in the vicinity of the main sensory nucleus; R2 uses a broader and less-well-defined reflex pathway in the pons. It has been established that R1 and R2 are generated by the same facial motor neurons.

The elicitation of blink reflexes establishes the integrity of the afferent trigeminal nerve, the efferent facial nerve, and the interneurons in the pons (R1) and caudal medulla (related to the bilateral R2 response). The test may also be helpful in identifying a demyelinating neuropathy when the facial and oropharyngeal muscles are affected and those of the limbs are relatively spared, thereby leaving conventional nerve studies normal. In such cases, the blink responses are delayed ipsilaterally and contralaterally as a result of conduction block in the proximal facial nerve. Direct facial nerve stimulation often fails to demonstrate this block because only the distal segment of the nerve is amenable to study. Although the blink responses are rarely necessary for diagnosis, most patients with hereditary neuropathy have blink response abnormalities. In Bell's palsy there is a delay or absence of R1 and R2 responses only on the affected side. Large acoustic neuromas (vestibular schwannomas) may interfere with the afferent trigeminal portion of the pathway and give rise to abnormal responses on the affected side. Diseases of the brainstem have yielded inconsistent responses. It is noteworthy that the test is normal in patients with trigeminal neuralgia.

Segmental Motor, Cranial, and Somatosensory Evoked Potentials (See also Chap. 2)

These techniques find use in diseases that affect the spinal roots and in studying central pathways.

By applying a magnetic stimulus, which induces an electrical impulse, or by a directly delivered electrical stimulus over the lower cervical or lumbar spine, it is possible to activate the motor (anterior) roots and to measure the time required to elicit a muscle contraction (see review by Cros and Chiappa). These root stimulation tests can be quite uncomfortable for the patient as a result of the contraction of muscles surrounding the stimulation site.

Transcranial magnetic stimulation of the cerebral cortex permits measurement of the latency of muscle contraction after excitation of motor neurons in the cortex. Thus, the integrity of the entire corticospinal system, from the cortical motor neurons through spinal tracts, anterior horn cells, and the peripheral motor nerve can be determined. By combining this technique with the above-described root stimulation, it becomes possible to measure central and peripheral motor conduction times. These forms of motor testing have their main use in the study of multiple sclerosis, amyotrophic lateral sclerosis (ALS), and related disorders.

As described in Chap. 2, by applying repetitive electrical stimuli to a peripheral nerve, the sensory evoked responses can be recorded from sites along the nerve and plexus as well as in central pathways (the thalamus and somatosensory cortex). These somatosensory evoked potential tests find their main use in the diagnosis of multiple sclerosis and in disorders of the sensory nerve roots as discussed in Chaps. 2 and 36. Discussion of magnetic stimulation, collision techniques, and quantitative EMG, among other topics, can be found in several monographs, such as the ones by Kimura, by Aminoff, and by Brown and Bolton.

Repetitive Motor Nerve Stimulation (See also Chap. 49)

This test of the neuromuscular junction is based on Jolly's observation in 1895 that in myasthenia gravis the strength of muscular contractions progressively declines in response to a train of stimuli. By adjusting the amplitude of a stimulus over a nerve to the supramaximal range, a maximal CMAP may be obtained for each stimulus. With repeated stimuli, each response will have the same waveform and amplitude until normal fatigue supervenes. In a healthy individual, a muscular response follows each stimulus with rates of stimulation up to 25 per second for periods of 60 s or more before a decrement of the CMAP appears. In certain disorders, notably myasthenia gravis, a train of 4 to 10 stimuli at rates of 2 to 5 per second (optimally 2 to 3 per second), the amplitude of the motor potentials decreases and then, after four or five further stimuli, may increase slightly (Fig. 45-4A). A progressive reduction in amplitude is most likely to be found in proximal muscles, but these are not easily stimulated for which reason the locations most commonly used for clinical testing are the accessory nerve in the posterior triangle of the neck (trapezius), the ulnar nerve (hypothenar muscle), the median nerve at the wrist (thenar muscle), and the facial nerve (orbicularis oculi muscle). A decrement of 10 percent or more denotes a failure of a proportion of the neuromuscular junctions.

The sensitivity of the procedure is improved by first exercising the tested muscle for 30 to 60 s; a form of posttetanic potentiation. (The full procedure consists of testing the muscle with a train of stimuli before and immediately after exercise [or maximal voluntary contraction] and at 30-s intervals for several minutes. The posttetanic potentiation partially compensates for the depletion of ACh during slow rates of stimulation; this is followed by a decrease in the excitability of the neuromuscular junction during the approximately 2 to 4 min after exercise.) The induced failure of neuromuscular transmission in myasthenia is similar to the one produced by curare and other nondepolarizing neuromuscular blocking agents, and both cases can be partially corrected with anticholinesterase drugs such as neostigmine and edrophonium. Similar but lesser decremental responses may occur in poliomyelitis, ALS, and certain other diseases of the motor unit or motor nerve, particularly those resulting in the growth of reinnervating nerve twigs.

The Lambert-Eaton myasthenic syndrome, often associated with oat cell carcinoma of the lung, as discussed in Chap. 49, is characterized by a presynaptic blockage of acetylcholine release and produces the opposite defect of neuromuscular transmission to the one recorded in myasthenia gravis. During tetanic stimulation (20- to 50-per-second repetitive stimulation of nerve), the muscle action potentials, which are small or practically absent with the first stimulus, increase in voltage with each successive response until a more nearly normal amplitude is attained (see Fig. 45-4B). Exercising the muscle for 10 s before stimulation will cause a similar posttetanic facilitation in patients with the Lambert-Eaton syndrome (200-fold increases are not uncommon). A less important decremental response to slow stimulation may occur, but it is difficult to discern because of the greatly diminished amplitude of the initial responses. Neostigmine has little effect on this phenomenon, but it is reversed by guanidine and 3,4-diaminopyridine, which stimulate the presynaptic release of ACh. The effects of botulinum toxin and of aminoglycoside antibiotics are similar, i.e., being active at the presynaptic membrane, they produce an incremental response at high rates of stimulation.

The single-fiber EMG, discussed in a later section, is an even more sensitive method of detecting failure of the neuromuscular junction.

Needle Examination of Muscle (Electromyography)

This technique requires the use of monopolar or concentric bipolar needle electrodes, which are inserted into the body of the muscle to record the electrical activity generated by contraction. With concentric electrodes, the tip of the wire that runs in the hollow of the needle is in proximity to many muscle fibers belonging to several different overlapping motor units; this is the active recording electrode. The shaft of the needle, in contact over most of its length with intercellular fluid and many other muscle fibers, serves as the reference electrode. Monopolar electrodes use the uninsulated needle tip as the active electrode, while the reference electrode may be another monopolar needle electrode placed elsewhere in subcutaneous tissue or a surface electrode on the skin overlying the muscle. Patients almost invariably find this portion of the test uncomfortable and should be prepared by a description of the procedure. Rapid and brief needle insertion by the skilled examiner makes the test more tolerable.

As the electrical impulse travels along the surface of the muscle toward the recording electrode, a positive potential is recorded on the oscilloscope, i.e., the recorded signal is deflected downward by convention (at A in Fig. 45-5). When the depolarized zone moves under the recording electrode, it becomes relatively negative and the beam is deflected upward (at B). As the depolarized zone continues to move along the sarcolemma, away from the recording electrode, the current begins to flow outward through the membrane toward the distant depolarized region, and the recording electrode becomes relatively positive again (at C). There is then a return to the resting isopotential position. The net result is a triphasic action potential, as in Fig. 45-5. This configuration is typical of the firing of a single fiber.

The electrical activity of various muscles is recorded both at rest and during active contraction by the patient. As indicated earlier, muscle fibers do not normally discharge until activated together in motor unit activity. This involves the almost simultaneous contraction of all the muscle fibers innervated by a single anterior horn cell. Although the typical configuration of a motor unit potential (MUP) is triphasic, up to 10 percent of normal MUPs consist of four or more phases (polyphasic potentials); however, an excess of polyphasic potentials beyond this is pathologic.

Normal muscle in the resting state should be electrically silent; the small tension spoken of as muscle tone has no EMG equivalent. There are, however, two closely related types of normal spontaneous activities and another that is induced by the insertion of the needle itself. One is a low-amplitude, 10- to 20-μV monophasic (negative) potential of very brief (0.5 to 1 ms) duration. These represent single or synchronized miniature end plate potentials (MEPPs) because of the small number of ACh quanta that are being released all the time. They are normally sparse but are most evident when the recording needle electrode is placed near a motor endplate ("endplate noise"). Fortuitous placement of the needle electrode very close to or in contact with the endplate gives rise to a second type of normal spontaneous activity. That is characterized by irregularly discharging high-frequency (50- to 100-Hz) biphasic spike discharges, 100 to 300 μV in amplitude (i.e., large enough to cause an isolated muscle action potential). These potentials have been termed endplate spikes and represent discharges of single muscle fibers excited by spontaneous activity in nerve terminals. They must be distinguished from fibrillation potentials (see later). Finally, insertion of the needle electrode into the muscle injures and mechanically stimulates a number of fibers, causing a burst of potentials of short duration (300 ms). This is referred to as normal insertional activity, but the extent of this activity is greatly raised in certain pathologic states as noted below.

When a muscle is voluntarily contracted, the action potentials of motor units begin to appear. One can observe a pattern of force build up by watching the progressive recruitment of MUPs; the initial ones, representing smaller motor units, firing at rates of 5 to 10 per second. With increased force of contraction, there is recruitment of larger, previously inactive motor units as well as an increased rate of firing (40 to 50 per second; Fig. 45-6A). Because individual MUPs can no longer be distinguished during maximal voluntary contraction, this activity is referred to as a complete interference pattern (Fig. 45-6A, right). This is seen not only as a summated signal pattern but is also heard as a mixed high-frequency clicking when the electrical activity is made audible. As muscles relax, an increasing number of units drop out. If a muscle is weakened by denervation or if electrical conduction is blocked, there obviously will be fewer MUPs, but the firing rate is still rapid (reduced recruitment; see Fig. 45-6B). In contrast, with poor voluntary effort and with upper motor neuron lesions, the MUPs fire in decreased numbers, at slower rates, and often in an irregular pattern (termed poor activation).

In the usual EMG examination, a plan for the study is made based on detailed knowledge of muscular innervation and focusing on the regions affected by weakness. In some patients, as in those with motor neuron diseases or polymyositis, a wider sampling of muscles is required to detect changes in asymptomatic regions.

The Abnormal Electromyogram

Clinically important deviations from the normal EMG include (1) increased or decreased activity upon insertion of the needle; (2) the occurrence of abnormal "spontaneous" activity during the relaxed state (fibrillation potentials, positive sharp waves, fasciculation potentials, cramp potentials, myotonic discharges, myokymic potentials); (3) abnormalities in the amplitude, duration, and shape of single MUPs; (4) a decrease in the number of MUPs and changes in their firing pattern; (5) variation in amplitude and number of phases of MUPs during voluntary contraction; and (6) the demonstration of special phenomena, such as electrical silence during shortening of the muscle (physiologic contracture and states of continuous muscle fiber activity).

Insertional Activity

At the moment the needle is inserted into muscle, there is a brief burst of action potentials that ceases once the needle is stable, provided that it is not in a position to irritate a nerve terminal. Increased insertional activity is seen in most instances of denervation as well as in many forms of primary muscle disease and in disorders that dispose to muscle cramps. In cases of advanced denervation or myopathy, in which muscle fibers have been largely replaced by connective tissue and fat, insertional activity may be decreased and there is a palpable increase in the mechanical resistance to the insertion of the needle.

Abnormal "Spontaneous" Activity

With the muscle at rest, spontaneous activity of single muscle fibers and of motor units, known respectively as fibrillation potentials and fasciculation potentials, is abnormal. The two phenomena are often confused. Fibrillation is the spontaneous contraction of a single muscle fiber. It occurs when the muscle fiber has lost its nerve supply and is ordinarily not visible through the skin (but may be visible in the tongue). Fasciculation represents the spontaneous firing of an entire motor unit, causing contraction of a group of muscle fibers, and may be visible through the skin. The irregular firing of a number of motor units, seen as a rippling of the skin, is called myokymia.

Fibrillation Potentials

Destruction of a motor neuron or interruption of its axon causes the distal part of the axon to degenerate, a process that takes several days or more. The muscle fibers formerly innervated by the branches of the dead axon—that is, the motor unit—are thereby disconnected from the nervous system. By mechanisms that are still obscure, the chemosensitive region of the sarcolemma at the motor endplate "spreads" after denervation to involve the entire surface of the muscle fiber. Then, 10 to 25 days after death of the axon, the denervated fibers develop spontaneous activity; each fiber contracts at its own rate and without relation to the activity of neighboring fibers. This spontaneous activity is associated with a random conglomeration of brief di- or triphasic fibrillation potentials (Fig. 45-7A) having a duration of 1 to 5 ms and rarely exceeding 300 μV in amplitude. When brief spontaneous fibrillation potentials of this sort are observed firing regularly at two or three different locations (outside the endplate zone) of a resting muscle, one may conclude that the fibers are denervated. Usually, fibrillation potentials discharge at an almost regular rate. In some early lesions (less than 6 to 8 weeks), irregularly firing fibrillation potentials may be observed.

Diseases such as poliomyelitis, which damage spinal motor neurons, or injuries of the peripheral nerves or anterior spinal roots, frequently produce only partial denervation of the involved muscles. In such muscles, one electrode placement may record fibrillation potentials at rest from denervated fibers and normal potentials during voluntary contraction from nearby healthy fibers. Fibrillation potentials continue until the muscle fiber is reinnervated by progressive proximal-distal regeneration of the interrupted nerve fiber or by the outgrowth of new axons from nearby healthy nerve fibers (collateral sprouting), or until the atrophied fibers degenerate and are replaced by connective tissue, a process that may take many years. In addition, fibrillation potentials may take the form of positive sharp waves, i.e., spontaneous, initially positive diphasic potentials of longer duration and slightly greater amplitude than the spikes of fibrillation potentials (see Fig. 45-7A).

Fibrillation potentials, while characteristic of neurogenic denervation, are not altogether specific; for example, they are seen in muscle diseases such as polymyositis and inclusion body myopathy which presumably damage the muscle fiber and make its membrane electrically unstable.

Fasciculation Potentials

As stated earlier, a fasciculation is the spontaneous or involuntary contraction of a motor unit or part of a motor unit. Such contractions may cause a visible dimpling or twitching under the skin, although ordinarily they are of insufficient force to move a joint. Large distal fasciculations can briefly displace a finger or toe. They occur irregularly and infrequently, and prolonged inspection of the skin overlying a muscle may be necessary to detect them. The accompanying electrical form of an individual fasciculation potential is relatively constant. Typically, a fasciculation potential will have 3 to 5 phases (i.e., they are "polyphasic" as described later, in contrast to normal biphasic muscle activity), a duration of 5 to 15 ms (longer than normal but somewhat less in the facial muscles), and an amplitude of several millivolts (see Fig. 45-7B). Fasciculation potentials are evidence of motor nerve fiber irritability. Thus, the combination of fibrillations and fasciculations indicates active denervation combined with more chronic reinnervation of muscle.

The precise source of fasciculation is still contested. Forster and colleagues challenged the original belief that the discharge originated in anterior horn cells by demonstrating that fasciculations persisted after nerve block in ALS and ceased only with the appearance of fibrillation potentials, signifying wallerian (axonal) degeneration. These observations favored a distal site of generation. Other physiologic and pharmacologic evidence pointed to the first segment of the motor axon, or to the distal axon, or even to the motor point (the site of insertion of the nerve into muscle), involving elements of the postsynaptic muscle membrane (particularly in the case of benign fasciculations) as the source of the spontaneous electrical activity. It seems that several regions of the axon are capable of spontaneous impulse generation, depending on the underlying disease. Most of the diseases that produce fasciculations involve the anterior horn cell or the motor root, but more distal sites in the motor axon are spontaneously active in cases of nerve compression and polyneuropathy.

Occasional fasciculation potentials, particularly in the calves, hands, and periocular or paranasal muscles, occur in many normal persons. They can be almost constant for days or weeks on end, or even for years in some individuals, without weakness or wasting; therefore they need not be taken as evidence of disease ("benign fasciculations"). Certain quantitative features of fasciculations, such as brief duration and a consistent pattern and location of firing, favor benign over pathologic discharges. Shivering induced by low temperature and twitchings associated with low serum calcium levels are other forms of fasciculatory activity.

Fasciculation potentials occur with great frequency in chronic, slowly advancing, destructive diseases of the anterior horn cells, such as ALS and progressive spinal muscular atrophy. In these diseases, both voluntary MUPs and fasciculation potentials may be of long duration (more than 15 ms) and of increased amplitude, indicating chronic denervation and reinnervation. They are seen often in the early stages of poliomyelitis but only occasionally in the chronic phase of the disease, perhaps because the affected cells die rapidly. When anterior horn cells degenerate once again in older individuals who had had poliomyelitis (postpolio syndrome), fasciculations may return. Occasionally, they are seen in one muscle as a result of a compressive anterior root lesion, such as those caused by a protruded intervertebral disc. Large numbers of axons may be affected in this case, with the result that the fasciculations (or even cramps) may be even more prominent than with disease of anterior horn cells but they are restricted to the territory of innervation of the root or nerve. Fasciculation potentials in lesser numbers are also observed with chronic nerve entrapments, e.g., ulnar neuropathy at the elbow and other peripheral nerve lesions and some polyneuropathies. In all these cases, the damaged neuron or its axon seems to leave intact axons in a state of hyperirritability. The blocking of axon conduction by local anesthesia does not abolish fasciculations, but curare-like drugs do so.

Other Types of Spontaneous and Elicited Electrical Activity

(See Chaps. 48 and 50) These various phenomena, including some discussed above, can be classified according to their generating source. The muscle fibers themselves are the source of fibrillations, positive sharp waves, and complex repetitive discharges (CRDs). The motor axons produce fasciculation potentials, myokymic discharges, neuromyotonia, and cramp syndromes; and the central nervous system is the source of complex ensembles of continuous motor activity such as occur in the stiff man syndrome.

The common phenomenon of complex repetitive discharges, referred to in the past as bizarre high-frequency discharges, consists of repetitive spontaneous potentials created by numerous single muscle fibers that fire in near synchrony; there is often an erratic configuration and abrupt starting and stopping of the discharges. They are seen in some myopathies, in hypothyroidism, and in certain denervating disorders, and are a mark of chronicity (lesions more than 6 months old). High-frequency coupling of action potentials into doublets, triplets, or higher multiples of single units, indicating instability in repolarization of the nerve fiber to a muscle, occurs in tetany and in the early stages of myokymia.

Myokymia is a persistent quivering and rippling of muscles at rest (colloquially called "live flesh"). The EMG picture is distinctive. The spontaneously firing MUPs are called myokymic potentials, or discharges and consist of groups of repetitive discharging units, each firing at its own rate, quasirhythmically, usually several times per second, followed by a briefer period of silence. The small motor unit discharges may occur singly or as doublets, triplets, or multiplets.

The site of generation of this activity has been contested, possibly because it may arise from several sites along the motor nerve. The discharge corresponds to an alteration in the calcium concentration in the microenvironment of the axon. Spontaneous discharges arising in large myelinated fibers have been implicated in the genesis of myokymia; indeed, demyelinating polyneuropathies are among the conditions that give rise to this phenomenon. Myokymia is also caused by peripheral nerve hyperexcitability because of both K channel mutations and antibodies against the channels. This activity may be blocked by lidocaine infusion around the peripheral nerve and may be diminished by carbamazepine or phenytoin. Central forms of myokymia also occur, as in multiple sclerosis; the mechanism is similar to the peripheral form, namely irritation or demyelination of the motor nerve, but in its fascicular (central) course.

Focal and segmental myokymias differ in small ways from the generalized form of myokymia with regard to the timing and duration of the discharges. The focal types refer mainly to facial myokymia, seen most often in multiple sclerosis, Guillain-Barré syndrome, large cerebellopontine angle tumors, or compression of the facial nerve by a small aberrant blood vessel, but it may follow any peripheral nerve injury and regeneration. The EMG patterns are complex, either high-frequency (30- to 100-Hz) recurrent bursts or brief lower-frequency bursts. Segmental myokymia is a common occurrence in demyelination and in radiation injuries of the brachial plexus. The EMG bursts tend to be longer and less frequent than in generalized myokymia, and the interburst frequency is highly variable. The origin of these discharges is probably in the distal peripheral nerve, where activity of afferent fibers, possibly via ephaptic transmission, irregularly excites distal motor terminals. Segmental myokymia refers to similar activity in the distribution of one or more adjacent motor roots again, usually related in some way to demyelination. This activity persists during sleep and general anesthesia.

The phenomenon of generalized myotonia, or neuromyotonia denotes a failure of voluntary relaxation of muscle because of sustained firing of the muscle membrane (see Chaps. 48 and 50), is characterized by high-frequency repetitive discharges generally having a positive sharp waveform. These myotonic discharges wax and wane in amplitude and frequency, producing a "dive-bomber" sound on the audio monitor. The discharges can be elicited mechanically by percussion of the muscle or movement of the needle electrode and are also seen following voluntary contraction or electrical stimulation of the muscle via its motor nerve. The MUPs may appear normal during voluntary contraction, but they are not followed by the silence that normally occurs on relaxation; instead, there is a "prolonged afterdischarge" consisting of long trains of fibrillation-like potentials that may take as long as several minutes to subside (Fig. 45-8A). These EMG findings can be seen with any myotonic disorder. If the muscle is activated repeatedly at short intervals, the late discharge becomes briefer and briefer and eventually disappears (see Fig. 45-8B), as the patient becomes able to relax the exercised muscle ("warmup" effect).

In paradoxical myotonia the myotonia worsens after each of a succession of voluntary contractions. This is the converse of what happens in myotonia congenita (Thomsen disease). As shown by single-fiber EMG studies, myotonia is generated by single muscle fibers and the mechanism of the membrane instability, at least in some forms, seems to involve changes in the chloride conductance. These disorders are discussed in subsequent chapters.

The cramp-like contracture of McArdle disease and phosphofructokinase deficiency is associated with electrical silence of contracting muscle. This feature is an important part of the definition of true physiologic muscle contracture (as distinguished from chronic shortening of a muscle and its tendon which, strictly speaking, is a pseudocontracture).

In the syndrome of continuous muscle fiber activity or Isaacs syndrome (see Chap. 50), which is a generalized form of myokymia, the EMG discloses high-frequency (up to 300-Hz) repetitive discharges of varying waveforms. In the stiff man syndrome, painful muscle spasms and stiffness are generated by a spinal mechanism; the EMG potentials resemble normal motor units but are abnormal by virtue of continuous firing at rest (see Chap. 50).

Abnormalities in Amplitude, Duration, and Shape of Motor Unit Potentials

Figures 45-9 (schematically) and 45-10 depict the ways in which disease processes affect the motor unit and the appearance of the MUP in the EMG.

Motor Unit Potentials in Denervation

Early in the course of denervation, many motor units with functional connections to the spinal cord are unaffected, and although the number of MUPs appearing during contraction is reduced, the configurations of the remaining ones are quite normal. In time, the remaining MUPs often increase in size and in electrical amplitude, perhaps two to three times normal, and become longer in duration and sometimes polyphasic (more than four phases).

Such large and sometimes giant polyphasic potentials (see Figs. 45-9C and 45-10B) arise from motor units containing more than the usual number of muscle fibers that are spread out over a greatly enlarged territory within the muscle. Presumably, new nerve twigs have sprouted from nodal points and terminals of undamaged axons and have reinnervated previously denervated muscle fibers, thus adding them to their own motor units. Soon after reinnervation, the MUPs generated will be low in amplitude, extremely prolonged, and polyphasic, findings that constitute a transitional configuration of early reinnervation. These amplitudes disappear as the motor unit is reestablished. Increased amplitude is usually associated with very chronic, proximal axon loss, for example, with remote poliomyelitis and chronic radiculopathy. These MUPs are to be differentiated from (1) polyphasic potentials of normal duration, which, as has been mentioned, make up as much as 10 percent of the total number of MUPs in normal muscle, and (2) polyphasic MUPs of short duration and low amplitude, which are characteristic of most myopathies and of myasthenia gravis and other disorders of neuromuscular transmission.

The Motor Unit Potential in Myopathy

As Fig. 45-9B shows, diseases such as polymyositis, the muscular dystrophies, and other myopathies that randomly destroy muscle fibers or render them nonfunctional, and obviously reduce the population of muscle fibers per motor unit. Therefore, when such a unit is activated, its potential is of lower voltage and shorter duration than normal, and it may also appear polyphasic as the compound MUP becomes fragmented into its constituent single-fiber potentials. Slowing of the propagated muscle fiber action potential in affected muscle fibers also contributes to the changes in the "myopathic" MUP. When most of the muscle fibers are affected, the MUPs are very small and of short duration and are recruited out of proportion to the tension generated, so-called early recruitment. Both types of alterations produce a characteristic high-pitched crackling sound from the audio monitor that has been likened to rain falling on a tin roof. They occur in all forms of chronic myopathies. Identical MUP changes are seen occasionally with other processes that cause disintegration of the motor unit, for example, early Guillain-Barré syndrome (because of conduction block along some of the terminal nerve fibers), and rarely with disorders of neuromuscular transmission (myasthenia gravis, other myasthenic syndromes), but they are most characteristic of primary muscle disease.

As mentioned earlier, fibrillation potentials, while typical of denervation, are sometimes seen in the myositides, and the rapidly progressive muscular dystrophies, perhaps because the muscle membrane has been made unstable; in the past the finding was attributed to isolation of a segment of the fiber from its nerve supply. In myasthenia gravis, where transmission of impulse fails at the neuromuscular junction, a single MUP may vary in amplitude during sustained weak contraction and some fibers cease to function. Electromyographic recordings of single muscle fibers belonging to the same motor unit disclose varying interpotential intervals on successive discharges; this phenomenon is called "jitter" and increases to the point of actual block, with deficits in neuromuscular transmission within a motor unit (see below and Chap. 50).

Abnormalities of the Interference Pattern

Diseases that reduce the population of functional motor neurons or axons within the peripheral nerve decrease the number of motor units that can be recruited in the affected muscles. The decreased number of motor units available for activation produces a low-amplitude interference pattern with only a few remaining units firing at a moderate to rapid rate. A severe reduction in the interference pattern may result in the recruitment of only a single unit (see Fig. 45-6B). Structural damage to nerve, as well as demyelinating block, can produce this pattern of reduced recruitment; indeed, a reduced recruitment pattern coupled with the absence of denervation indicates a conduction block.

If muscle power is reduced in diseases such as polymyositis or muscular dystrophy, in which individual muscle fibers are affected, there may be little or no reduction in the number of motor units available for recruitment until the process is far advanced and entire MUPs have been lost as a result of random loss of all their constituent muscle fibers. Nonetheless, each motor unit will consist of fewer muscle fibers than normal, so more motor units must be activated to reach a certain degree of force. A modest effort can thus produce a full interference pattern despite marked weakness (increased recruitment). Because fewer muscle fibers are firing, the amplitude of the pattern will be reduced from normal. This type of full, highly complex interference pattern of less-than-usual amplitude in the face of dramatic weakness is the hallmark of myopathy (see Fig. 45-6C).

Motor Unit Quantification

This experimental technique, developed by McComas and colleagues estimates the size of motor units and is thus exquisitely sensitive to changes of denervation and reinnervation. It is carried out by applying a weak stimulus to a motor nerve or motor point and increasing it gradually as the evoked muscle response is recorded. Each quantal increase in the compound-evoked response is presumed to be caused by the addition of a single motor unit. In reinnervated muscles, the additional units are reduced in number and are abnormally large. The technique is used mainly for the investigation of motor neuron disorders. When a normal number and configuration of motor units is found, it has been helpful in distinguishing benign fasciculations from those of serious diseases.

Single-Fiber Electromyography

This is a special technique for the recording of single-muscle-fiber action potentials that has found utility in measuring muscle fiber density and in detecting so-called jitter in disease of the neuromuscular function, particularly myasthenia gravis. Fiber density is an index of the number and distribution of muscle fibers within a motor unit. Jitter is the variability of the interpotential interval of successive discharges of two single muscle fibers belonging to the same motor unit. This phenomenon is largely a result of the very slight variability of delay at the branch points in the distal axon and by synaptic delay at the neuromuscular junction, especially in myasthenia gravis where it has found its main clinical use. Fiber density and jitter may, however, also be increased in neuropathic disorders that cause denervation with reinnervation. Both are usually normal or only slightly increased in myopathic disorders.

Testing for jitter is carried out by having the patient voluntarily contract a muscle to the slightest degree possible so as to activate only one motor unit (requiring a great deal of cooperation by the patient) or by stimulating an intramuscular nerve twig (requiring great patience on the part of the examiner). The EMG needle is advanced until two muscle fibers from the same motor unit are recorded. If the oscilloscope sweep is triggered by the firing of the first fiber, a fluctuating latency of the second fiber potential can be seen on the screen as a movement (jitter) of the second peak. The degree of jitter can be quantitated by measuring the interval between the activation of the two muscle fibers (the result of slightly differing lengths of the terminal axons) from which a mean interpeak interval is determined. Approximately 20 fiber pairs are sampled, and an average of the mean consecutive intervals can be derived. In a muscle such as the extensor digitorum communis, the average variation should be no more than 34 ms. The acceptable average is lower for large proximal muscles. Also, in disease of the neuromuscular junction, one muscle fiber in a pair may fail to fire intermittently as a result of a blocking of conduction. Further details of this technique and its clinical applications are discussed by Stålberg and Trontelj.

Imaging of Muscle and Nerve

Imaging techniques—CT, MRI, and ultrasonography—enable one to measure muscle volume and to recognize qualitative changes in muscle structure (see review of DeVisser and Reimers). Such methods are finding some clinical and research use in the diagnosis of disorders of muscle and in gauging the effects of treatment. CT scans of dystrophic muscle show foci of decreased attenuation, representing masses of fat cells. The fatty masses spread gradually from multiple foci and eventually replace muscle fibers. The original shape of the muscle is retained; indeed, an enlarged weak muscle containing mostly fat confirms the clinical impression of pseudohypertrophy. In denervative atrophy, the muscles are obviously small and contain multiple punctate areas of decreased attenuation, which represent interstitial fat. Eventually, large portions of chronically denervated muscle may be replaced by fat. Blood, blood products, and calcium deposits are expressed by increased attenuation in CT. This may be helpful in the diagnosis of muscle trauma, myositis ossificans, and dermato- and polymyositis.

Fat and bone marrow have a high signal intensity in MR images, whereas fascia, ligaments, and cortical bone lack signal intensity. In T1-weighted images, normal muscle has a low signal and dystrophic muscle, a slightly increased signal; in T2-weighted images, dystrophic muscle has a slightly enhanced signal. Given its sensitivity to these dystrophic changes in muscle, MRI is particularly effective in determining the distribution of muscle involvement in a dystrophy. Transverse MR sections, for example, can help distinguish the topographic patterns of such disorders as the proximally predominant Becker dystrophy and the distally predominant Miyoshi myopathy, or subtle subtypes of Emery-Dreifuss dystrophy (Mercuri et al). Spectroscopic MRI in metabolically determined myopathies has the capacity to quantitate levels of selected biochemical constituents of muscle, including intracellular pH and levels of metabolic intermediates such as phosphocreatine. This technique is particularly effective in demonstrating subnormal generation of intracellular acidosis after a limb is exercised in disorders of glycogenolysis and of glycolysis. Some individuals with mitochondrial disease of muscle will demonstrate rapid depletion of energy supplies and profound delays in recovery that can be quantified and used as an end point for treatments.

New magnetic resonance techniques are also being developed that allow the imaging of nerves. This may be an aid in assessing traumatic nerve injury and in demonstrating neuromas, hypertrophy or atrophy of a nerve trunk, as well as neural tumors.

Muscle Biopsy

Muscle biopsy can be of great diagnostic value, but both surgical and microscopic techniques must be exacting. The muscle chosen for study should be accessible; there should be evidence that it has been affected but not totally destroyed by the disease in question; and it should not have been the site of a recent injection or EMG study, since the trauma of the needles produces focal necrotizing and inflammatory lesions. Muscle biopsy is helpful in distinguishing the following basic disorders in patients with neuromuscular disease.

1. Denervation atrophy. Reduction in the size of muscle fibers with an accompanying enlargement of intact motor units (because of collateral reinnervation) and degenerative changes in some fibers are the main changes of denervation atrophy. Group atrophy denotes enlarged motor units where all the fibers in the group are reduced to the same size; this is typical of progressive denervation. Normally, the fibers of each motor unit are not clustered, so that when grouping occurs it means that some fibers of a denervated unit have been adopted by an adjacent intact motor unit. This change typifies axonal neuropathies and many spinal cord diseases that affect the anterior horn cell. A related change is particularly well shown in histochemical stains for ATPase, phosphorylase, and oxidases, where the normal mosaic pattern of fiber types is altered. The use of these stains reveals fiber type grouping, the most specific histologic evidence of denervation and reinnervation. Here, muscle fibers of similar histochemical type form groups of 15 or more fibers as a result of reinnervation by a single motor neuron. The diagnosis of denervation atrophy can usually be made from the clinical and EMG examinations; seldom is biopsy necessary for this purpose, but it is still used in cases of possible ALS, for example, where the diagnosis remains uncertain after other testing.

2. Segmental necrosis of muscle fibers with myophagia and various manifestations of regeneration. These are the typical changes in idiopathic polymyositis (in combination with infiltrates of inflammatory cells), and infective polymyositis (e.g., in the presence of Trichinella or Toxoplasma). These changes may also be observed in more limited form in Duchenne and other rapidly progressive muscular dystrophies.

3. Inflammation and vasculitis. Lymphocytic infiltration of the endomysium is most characteristic of polymyositis and in dermatomyositis it is predominantly perivascular and perimysial. The lymphocytic infiltrate may be florid in these processes but, as often, it is less intense and scattered. Inclusion body myopathy is characterized by inflammation but has additional diagnostic features. Lesser degrees of inflammation are common in the myopathies associated with Sjögren syndrome, mixed connective tissue disease, and scleroderma. Numerous other processes, including the infections mentioned earlier and some dystrophies (especially the fascioscapulohumeral type)—may be associated with an inflammatory reaction. There is usually acute myofibrillar destruction in regions of maximal lymphocytic infiltration.

4. Vasculitis. The muscle is a site of inflammatory vascular destruction in systemic diseases such as polyarteritis nodosa, and for this reason it is often useful to obtain a small sample of muscle adjacent to a nerve biopsy. The finding of a granulomatous myopathy may indicate the presence of systemic sarcoidosis.

5. Alterations in the protein and histochemical composition of muscle fibers are shown by special stains for enzymes, glycogen, and structural proteins that are implicated in disease. It has become possible to detect the absence or deficiency of specific structural proteins of the muscle membrane that define each of the muscular dystrophies: dystrophin, sarcoglycan, laminin, dysferlin, and others as discussed in Chap. 48. These tests require rapid freezing (in a cryostat) rather than formalin fixation. Also, a number of enzymatic deficiencies and intrafiber glycogen storage that lead to weakness and muscle fatigue may be detected by appropriate histochemical staining (see Chap. 48).

6. Unusual changes of muscle fibers. Included here are sarcoplasmic masses and disorganized ring or serpentine collections of myofibrils and myofilaments (Ringbinden) in myotonic dystrophy; glycogen masses in glycogen storage diseases, rods (nemaline), central cores, aggregates of lipid bodies, and other cytoplasmic changes in certain congenital myopathies; and nuclear and cytoplasmic inclusions that characterize but are not isolated to inclusion-body myositis. Application of histochemical methods and electron microscopy are the important techniques in the diagnosis of these disorders. These are discussed in relation to each of these specified diseases in the following chapters.

7. Abnormalities of mitochondria. Several distinctive abnormalities are readily visualized in muscle biopsies and are virtually diagnostic of an entire class of mitochondrial diseases. Light microscopy of frozen sections of muscle stained with the Gomori trichrome stain show the main feature, so-called ragged red fibers, which are accumulations of subsarcolemmal mitochondria.

8. Disorders of the neuromuscular junctions. Here, an abnormality may be revealed by performing the demanding procedure of motor point biopsy (to include the motor endplate), and the use of electron microscopy and special staining techniques for nerve terminals, cholinesterase, and the outlining of acetylcholine receptors. Myasthenia gravis, botulism, Lambert-Eaton syndrome, and myasthenic syndrome with motor endplate cholinesterase deficiency fall into this category. Biopsy is rarely necessary for the diagnosis of these disorders, but it has added considerably to our understanding of them.

As a rule, the muscle biopsy procedure requires no more than a small cleanly excised block of muscle, 1.0 to 2.0 cm, which is prevented from contracting by a clamp or by tying at full length to a tongue depressor. As an alternative to the standard surgical technique, percutaneous needle biopsies are now being done routinely in neuromuscular clinics and may be adequate for the diagnosis of certain muscle dystrophies and for their study over time and for a number of other muscle diseases and denervation atrophy. Unfortunately, the erratic distribution of lesions in many of the common disorders of muscle lessens the diagnostic yield of all types of biopsies, but in general the needle biopsy is less satisfactory than an open biopsy. The study of the muscle biopsy requires special care in removal, transport, and fixation techniques. These are discussed in detail by Engel (1994b).

Nerve Biopsy

Nerve biopsies are processed for study by conventional histologic methods and by thin-section phase and electron microscopy. These methods, sometimes supplemented by study of teased fiber preparations and immunologic studies, can provide valuable histopathologic data. With ordinary microscopy and staining, one may find evidence of focal inflammation, demyelination, axon destruction, vasculitis, amyloidosis, leprosy, or sarcoidosis. As mentioned in reference to certain guidelines cited below, these procedures are most valuable in the diagnosis of inflammatory, vasculitic, and amyloid neuropathies. In children, the nerve biopsy may reveal the histologic features of metachromatic or globoid leukodystrophy, giant axonopathy, or neuroaxonal dystrophy. However, nerve biopsy is relatively unhelpful in most other polyneuropathies and should be used prudently because the procedure is not without complications, including the occasional occurrence of wound infections, painful stump neuromas, persistent dysesthesias of the lateral foot or heel, and thrombophlebitis. Guidelines given by England and associates from a review of the literature up to 2009, has suggested that nerve biopsy is useful in cases of mononeuropathy multiplex, suspected amyloidosis, and unusual forms of chronic inflammatory demyelinating neuropathy, all the subjects of discussion in Chap. 46, but not clearly useful in nondescript distal peripheral sensorimotor neuropathies.

Representative of the results of nerve biopsies in a neuromuscular clinic is a prospective study of 50 consecutive cases of sural nerve samples reported by Gabriel and colleagues in which nerve biopsy served only to confirm the clinical diagnosis in 70 percent of cases and altered the clinical diagnosis in only 14 percent. Nevertheless, in some instances of idiopathic polyneuropathy that are not clarified by clinical and electrophysiologic testing, and particularly if a treatable chronic inflammatory polyneuropathy is suspected, many neuromuscular experts resort to sural nerve biopsy as a final diagnostic step. Although the sural nerve is typically chosen for biopsy, the superficial peroneal nerve and the adjacent peroneus brevis muscle gives a higher yield in cases of vasculitis (see Collins et al).

In diseases that involve only the motor nerves, it is sometimes useful to sample a fascicle of the superficial radial nerve or the nerve to the extensor digitorum brevis, which may be taken with the muscle itself and leaves little or no clinical deficit. Chronic inflammatory neuropathy and vasculitic neuropathy may be disclosed by this selective biopsy technique when the sural nerve is unaffected. In selected circumstances we have undertaken biopsy of small radicles of upper lumbar roots (L1 or L2) to establish the diagnosis of an infiltrative lymphoma. Little deficit occurs from the removal of nerves from these sites if the procedure is done by an experienced surgeon.

Additional techniques that are used in the study of muscle biopsies and specific pathologic findings that characterize the many individual diseases, of both muscle and peripheral nerve, are discussed in the chapters that follow.

Other Laboratory Tests in the Study of Muscle and Nerve Disease

None of the diagnostic procedures previously described may be taken as an infallible diagnostic index of a specific disease of muscle or nerve. Each procedure is subject to technical error and the findings to misinterpretation. A biopsy specimen may be taken from an unaffected muscle or portion of an affected muscle, and such an error causes the sample to be normal in the face of obvious clinical evidence of disease. This is particularly true of the inflammatory myopathies that affect muscle in heterogeneous and spotty manner.

Rough excision and improper fixation and staining produce artifacts that may be misinterpreted as marks of disease when, in fact, the muscle (and nerve) is microscopically normal. Similarly, EMG study may fail to record fibrillations in obviously denervated muscle, particularly in slowly progressive disorders. Also, in some of the muscles of the feet, fibrillations and fasciculations may be found in normal asymptomatic older individuals (Falck and Alaranta). As in the study of all disease, laboratory data have significance only if evaluated in the light of the clinical findings.