Evoked responses in normal and diseased muscle with particular reference to twitch potentiation

The compound muscle action potential and isometric twitch tension evoked by single and repetitive electrical stimuli are indicators of the number of motor units activated and of the contractile properties of the muscle. The action potentials and mechanical responses were recorded in proximal and distal muscles in patients with myasthenia gravis and myopathy and compared with findings in normal subjects. In normal muscle, at low rates of stimulation (2-3 s-1) the decrement was at most 5% in the action potential and 15-24% in the twitch tension. Tetanic stimuli (50 s-1) were unsuitable for diagnostic purposes because of movement artefact. In patients with myasthenia gravis, the incidence and size of the decrement of evoked responses were greater in the platysma than in the elbow flexors and the adductor pollicis (ADP) muscles. The 2-3 times greater post-tetanic facilitation (PTF) of the action potential in the platysma than in extremity muscles also indicates a more severe functional block in facial muscle. The PTF is an indicator of recruitment of blocked fibres. The maximal decrement was grossly related to the titre of antibodies against the acetylcholine receptor. To reveal failure of neuromuscular transmission in patients with myasthenia gravis without a decrement, a small dose of d-tubocurarine (0.2 mg in 30 ml of saline) was injected i.v. in the upper arm in a regional curare test. The sensitivity was greater in patients with myasthenia gravis than in controls and in patients with myopathy. Potentiation of twitch tension reflects contractile properties. In normal muscle twitch potentiation in the staircase (1-3 s-1, 100 s in duration) and after tetanus (50 s-1, 1.5 s in duration) was 2-3 times greater in the platysma than in the elbow flexors and ADP, presumably related to the greater proportion of fast-twitch fibers in facial muscle. The amplitude of the action potential and the twitch tension varied proportionally with the number of fibers activated and the difference in the decrements of the action potential and the twitch during the staircase in some patients with myasthenia gravis showed that the staircase phenomenon was diminished suggesting abnormalities in the excitation-contraction coupling. The diminution of the staircase and post-tetanic potentiation (PTP) in myopathy also indicates impairment of the excitation-contraction coupling. In rats with severe chronic myasthenia gravis, the staircase and PTP were decreased even when the failing neuromuscular transmission was circumvented by applying direct stimuli to the extensor digitorum longus muscle (EDL).     

Further electrophysiological studies of neuromuscular transmission in an animal model of myasthenia gravis

The development of experimental autoimmune myasthenia gravis (EAMG) in rats produces a significant reduction in the amplitude of spontaneously occurring miniature end-plate potentials (MEPPs) and impulse-evoked end-plate potentials (EPPs). This junctional abnormality, however, does not impair the ability of the affected fibers to produce propagated muscle action potentials of normal amplitude. Quantal content analysis indicates that in Mg²⁺-blocked EAMG muscle the mean number of acetylcholine (ACh) quanta released per nerve impulse closely approximates the corresponding normal value. At EAMG end-plates, the depolarization produced by either saturating or nonsaturating doses of carbamylcholine was significantly less than that seen at normal end-plates, suggesting a reduced acetylcholine receptor (AChR) content. When the depolarizing response to 250 μm carbamylcholine was examined at various EAMG end-plates with different sizes of MEPPs, there was a direct correlation between the carbamylcholine-induced depolarization and the MEPP amplitude; this correlation, however, was much less pronouced in normal end-plates, which supports the concept that the MEPP size in the receptor-immunized muscle reflects the content of the functional postsynaptic AChR. In control solution, the indirect twitch tension produced in EAMG muscle was normal. The twitch tension in EAMG muscle however, was almost completely abolished by a dose of d-tubocurarine that reduced the tension in normal muscle by only 50%. When 4-aminopyridine, a drug known to increase the quantum content of EPPs, was applied to curarized normal or EAMG muscles, normal muscle contraction was restored and the decremental response to repetitive nerve stimulation was abolished. We conclude that the major defect of neuromuscular transmission in EAMG results from postsynaptic abnormalities at the end-plates, presumably secondary to reduction of the number of functional AChRs. Chronic EAMG in rats is a reasonable model of human myasthenia gravis in which a similar defect of neuromuscular transmission is present.     

Evoked responses of the elbow flexors in control subjects and in myopathy patients

Electrical and mechanical responses were evoked in the elbow flexors (EFs) of normal subjects and myopathy patients by maximal stimulation of the musculocutaneous nerve by a wire electrode in the axilla. In normals, the decrement in the mechanical responses during short trains of twitches was small (10%). The potentiation during a staircase decreased with increasing frequency of stimuli, but the potentiation 30 sec after the 100-sec staircase was the same at all frequencies. Posttetanic potentiation was twice as large as the maximal potentiation attained during the staircase. The staircase phenomenon probably results from two opposing events, diminution and potentiation. Findings in 11 patients with myopathy varied according to the severity of involvement. Slight weakness of the EFs was associated with an increased twitch:tetanus ratio, decreased tetanic tension, and decrease in the staircase and posttetanic potentiation. Additionally, severe weakness of the EFs was associated with a diminution in twitch tension.     

Long-term effects of estrogen on rat skeletal muscle

The long-term effects of estrogen on the development of rat extensor digitorum longus (EDL) and soleus (SOL) muscles were examined using physiological and histochemical methods. The rats were in three groups: group 1, ovariectomized; group 2, sham-operated; and group 3, ovariectomized followed by estradiol administration. Isometric twitch and tetanic tensions of both the EDL and SOL obtained from 10-week-old rats were evoked by electrical stimulation. The isometric twitch tensions of the EDL in groups 2 and 3 were significantly lower than in group 1, however, there was no significant differences in isometric twitch tensions of the SOL among the three groups. The isometric tetanic tensions of the EDL in groups 2 and 3 were also significantly lower than in group 1, and the isometric tetanic tension of the SOL in group 3 was significantly lower than in group 1. There were no changes in the total number of muscle fibers or in the ratios of fiber types. But the mean fiber diameter of all fiber types (particularly in types IIA and IIB) was significantly lower in group 3 than in groups 1 and 2. Therefore, the possibility that estrogen may play an inhibitory role in the development of skeletal muscle fibers has to be considered.     

Facilitatory effects of 4-aminopyridine on neuromuscular transmission in disease states

The in vitro effects of 4-aminopyridine (4-AP) on neuromuscular transmission were determined by microelectrode techniques in intercostal muscles from patients with myasthenia gravis (MG) and the Eaton-Lambert syndrome (ELS), and in forelimb muscles from rats with experimental autoimmune myasthenia gravis (EAMG). In MG and EAMG, the amplitudes of miniature endplate potentials (MEPPs) and endplate potentials (EPPs) were reduced, and there was increased sensitivity to the blocking action of d-tubocurarine (dTc). In ELS, MEPP amplitude was normal but the average number of acetylcholine quanta released by nerve impulses was reduced, causing subthreshold EPPs. In EAMG muscle, 4-AP produced dose-dependent increases in EPP amplitude and in the duration of indirectly elicited muscle action potentials but no changes in MEPP amplitude and resting membrane potential. 4-AP completely reversed the postsynaptic blockade produced by dTc and EAMG. 4-AP appears to facilitate neuromuscular transmission in EAMG, MG, and ELS by increasing the neurally evoked transmitter release, thus overcoming either the pre- or the postsynaptic neuromuscular blockade.     

Changes in isometric contractile properties of fast-twitch and slow-twitch skeletal muscle of C57BL/6J dy2J/dy2J dystrophic mice during postnatal development

Our primary aim was to determine if there exists a preferential involvement of the fast-twitch or slow-twitch skeletal muscle fibers in the dy2J/dy2J strain of murine dystrophy. The changes in the contractile properties of the slow-twitch soleus (SOL) and the fast-twitch extensor digitorum longus (EDL) muscles of normal and dystrophic mice were studied at 4, 8, 12, and 32 weeks of age. Isometric twitch and tetanus tension were decreased in the 4- and 8-week-old dystrophic EDL compared with controls, this situation being reversed in the older animals. At 12 weeks, the dystrophic EDL generated 15% more tetanic tension than normal EDL and by 32 weeks no significant difference was seen between normal and dystrophic EDL twitch or tetanus tension. By 8 weeks, dystrophic EDL exhibited a prolonged time-to-peak twitch tension (TTP) and half-relaxation time (1/2RT) of the isometric twitch which continued to 32 weeks. For the dystrophic SOL, decreased twitch and tetanus tension was observed from 4 to 32 weeks. At 8 and 12 weeks, TTP and 1/2RT of dystrophic SOL were prolonged. However, by 32 weeks there was no longer a significant difference seen in TTP or 1/2RT between normal and dystrophic SOL. Our results appear to indicate that a loss of the primary control which is determining the fiber composition of the individual muscles is occurring as the dystrophic process advances.     

Functional evaluation of acute vincristine toxicity in rat skeletal muscle

Vincristine sulfate (VCR) was administered intravenously to rats at doses of 0.25, 0.5, and 0.75 mg/kg. During the first week following VCR treatment, extensor digitorum longus (EDL) muscle contraction strength and fiber electrophysiologic parameters were measured. At all doses tested, VCR strongly reduced twitch and tetanic tension. EDL fiber resting membrane potential was affected in a dose-dependent manner, and membrane depolarization was associated with the loss of excitability. Local membrane hyperpolarization by intracellular current application restored the capacity to produce action potential (AP). However, to elicit APs with a normal rate of rise, polarizing current had to be maintained for 3-5 minutes, indicating that the removal of Na+ channel inactivation followed a slow kinetics. Minor alterations in spontaneous synaptic transmission and in evoked transmission during high-frequency repetitive stimulation were seen only at the highest dose. It is suggested that VCR impairs skeletal muscle function by affecting primarily the contractile apparatus, whereas sarcolemmal alterations are evident at increased doses of the drug.     

Neural regulation of glucose 6-phosphate dehydrogenase in rat muscle: effect of denervation and reinnervation

We tested the hypothesis that glucose 6-phosphate dehydrogenase (G6PD) in rat extensor digitorum longus (EDL) muscle is under neural control by studying changes in G6PD activity in EDL muscles following nerve crush-induced denervation and reinnervation. Changes in G6PD were correlated with choline acetyltransferase activity, as well as with neurological function, muscle weights, and muscle isometric twitch tension. The data show a dramatic increase in G6PD following denervation. The gradual recovery of enzyme activity toward normal levels correlates with the return of functional synaptogenesis manifested by the return of neurological function, choline acetyltransferase, and muscle twitch tension. We conclude, therefore, that muscle G6PD is under neural control. G6PD activity provides a facile biochemical indicator of muscle reinnervation.     

Isometric contractile properties of fast- and slow-twitch muscles in normal and spastic mice

Isometric contractile properties of the fast-twitch extensor digitorum longus (EDL) and slow-twitch soleus (SOL) muscles of 24 male C57 mice were studied in vitro at 35°C. Ten animals exhibited the hereditary movement disorder known as spasticity. EDL muscles of spastic and normal mice developed similar specific tensions and they had similar twitch tension:tetanic tension ratios. The time-to-peak tension and the time to half relaxation in the twitch of spastic EDL muscles were, on average, slightly longer. SOL muscles of spastic and normal mice also developed similar specific tensions and their twitch contractions had similar times to half-relaxation. The time-to-peak tension in the twitch and the twitch tension: tetanic tension ratio of spastic SOL muscles were, on average, higher than in normal SOL. The significance of our findings remains somewhat uncertain.     

Contractile properties of rat fast-twitch skeletal muscle during reinnervation: effects of testosterone and castration

The isometric contractile properties of skeletal muscle were examined after nerve crush to establish the temporal sequence of recovery during reinnervation of normal, castrated, and testosterone-treated rats. Extensor digitorum longus muscles of male rats were studied in vivo 8 to 21 days after crushing the peroneal nerve 1 cm from the muscle. The earliest signs of functional reinnervation in normal animals were observed 8 to 9 days after nerve crush when faint muscle twitches with markedly prolonged twitch contraction times were recorded. By days 10 and 11, twitch tension was 9 to 20% of control, twitch contraction time was 149 to 183% of control, and tetanic tension was 4 to 9% of control values. The optimal frequency of stimulation was 58 to 64 Hz, the twitch:tetanus ratio was three times control values, and little or no posttetanic potentiation of twitch tension was observed. During the next 9 days there was a gradual return of all experimentally measured contractile properties toward control values; the relative rate of return was twitch tension greater than twitch contraction time greater than twitch:tetanus ratio greater than tetanic tension greater than optimal frequency of stimulation greater than posttetanic potentiation. Neither testosterone nor castration significantly altered either the rate or extent of functional reinnervation 8 to 21 days after nerve crush (P greater than 0.05). During this period the twitch:tetanus ratio for any given animal was highly correlated (r = 0.83, P less than 0.001) with the extent of functional recovery of neurally evoked muscle tension and was determined to be the most reliable index of the degree of muscle reinnervation. These data provide valuable baseline information for future studies of reinnervation of skeletal muscle.     

Characteristics of tension recruitment and mechanical activation in mammalian skeletal muscle

Characteristics of summation of isometric mechanical responses were studied in in situ flexor digitorum longus (FDL) (fast-twitch) and soleus (slow-twitch) muscles of the cat at 37°C. The pattern of tension recruitment in short tetanic contractions was examined at different stimulation frequencies. At frequencies leading to the development of maximum tetanic tension, the second stimulus activation made the largest individual contribution to tension development. The tension developed with two stimuli was dependent on the interval separating them. When the first two stimuli were fixed at a short interval such that the muscle developed the highest two-response tension, the tension developed following application of the third stimulus was independent over a wider range of two-to-three stimulus intervals. Application of a stimulus shortly after another, in general, lengthened the contraction. The results from the two muscles were basically similar when allowance was made for the difference in their speed of contraction. The marked tension potentiation seen in the twitch of FDL, with staircase effect and with cooling, was largely absent in the well-fused double response. We suggest that the mechanical activation occurring during the isometric twitch is incomplete in both muscle fiber types and that a second stimulus applied shortly after the first brings about nearly complete mechanical activation.     

Neurophysiology of fastest voluntary muscle contraction in hereditary neuropathy

In patients with hereditary motor and sensory neuropathy types I (demyelinative) and II (neuronal) and in normal subjects, isometric force and electromyographic activity of the first dorsal interosseous muscle were recorded during fastest voluntary contractions and during twitches evoked by nerve stimulation. The maximum voluntary force of the first dorsal interosseous muscle was also measured. In patients, fastest voluntary contraction time (i.e., time from onset of contraction to peak force) was prolonged and inversely proportional to maximum voluntary force. Maximum rate of rise of tension (i.e., slope of rise in force) was reduced and directly proportional to maximum voluntary force. In patients with hereditary motor and sensory neuropathy type I, contraction time was longer and the maximum rate of rise of tension was lower than in those with hereditary motor and sensory neuropathy type II. In patients and normal subjects, voluntary contraction time was closely correlated with the duration of electromyographic bursts. In patients, the twitch contraction time was prolonged and inversely proportional to maximum voluntary force. Twitch contraction amplitude was diminished and directly proportional to maximum voluntary force. Neither twitch contraction time nor amplitude were dependent on the type of hereditary motor and sensory neuropathy. Twitch contraction time evoked by proximal nerve stimulation was minimally longer than that evoked by distal stimulation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Twitch response in a myopathy with impaired relaxation but no myotonia

A patient with slow muscle relaxation but without accompanying motor unit or myotonic electrical activity had a unique staircase twitch response to repeated nerve stimulation. During 1-Hz stimulation, twitches recorded by measurement of the ankle dorsiflexor group displayed progressively increasing relaxation times with successive stimuli (37% increase) unlike the progressively decreasing relaxation time of the normal response (12-37% decrease). The response may be diagnostic of this unusual myopathy; the test methods are noninvasive and easily tolerated.     

Glycosaminoglycans treatment increases IGF-I muscle levels and counteracts motor neuron death: A novel nonanticoagulant action

The present study shows that sciatic nerve crush in 2-day-old rats causes extensor digitorum longus (EDL) muscle atrophy and motor neuron loss and that treatment with glycosaminoglycans (GAGs) promotes muscle reinnervation, motor neuron survival, and markedly increases insulin-like growth factor-I (IGF-I) content in the denervated muscles. EDL muscle denervation-induced atrophy in saline-treated rats is progressive and reaches the greatest extent at 42 days after birth, which correlates with reduced EDL weight growth. There is also a partial reinnervation as shown by the number of reinnervated EDL muscle fibers (65.4% of control) and by the poor restoration of the indirect isometric twitch tension (62% of control) that is further reduced under tetanic stimulation (34% of control). The number of surviving motor neurons that innervate EDL muscle drops from 55 +/- 3 to 29 +/- 8. In GAGs-treated 42-day-old rats, the effects of neonatal nerve lesioning on EDL muscle atrophy and denervation are successfully reversed, and the isometric twitch tension and the capacity to hold tetanic stimulation are restored to almost control levels. The number of surviving EDL motor neurons is also increased to 43 +/- 4. Treatment with GAGs selectively affects IGF-I content in denervated hindlimb muscles, which is augmented from 7.02 +/- 0.71 ng/mg tissue to 25.72 +/- 0.7 in the EDL and from 3.2 +/- 0.18 to a robust 211 +/- 9.6 in the soleus.     

The regulation of synaptic strength within motor units of the frog cutaneous pectoris muscle

The physiological properties of frog neuromuscular junctions may vary widely in a single muscle. In order to understand the factors that contribute to this variation, we have studied populations of synapses belonging to individual motor units of the frog cutaneous pectoris muscle. Motor units in this muscle differ widely in twitch strength. A motor axon's synaptic contacts could be found throughout the muscle, at both singly and polyneuronally innervated endplates. Indeed, over 36% of the endplates contacted by each isolated motor axon were polyneuronally innervated. Comparisons of synapses on muscle fibers in large twitch motor units with those in small twitch motor units reveal that endplate potential amplitude, transmitter release, and muscle fiber diameter are positively correlated with the strength of the motor unit contraction. Large and small twitch motor units differ more in their transmitter release than in their nerve terminal length, indicating that larger twitch motor units have a higher release per unit length of terminal. Among motor units of roughly similar twitch tension, transmitter release at endplates receiving only one axonal input is remarkably constant, independent of postsynaptic muscle fiber input resistance, or, presumably, nerve terminal size. In cases where two different motor axons contribute to a single endplate, the synaptic strength of each input is again related to properties of the contributing motoneuron, although the individual synaptic inputs are markedly reduced in strength and size relative to singly innervated endplates. Additionally, the diameter of polyneuronally innervated muscle fibers appears related to properties of both innervating motoneurons. Thus, the pre- and postsynaptic characteristics of neuromuscular junctions may be determined both by the motoneuron and by peripheral interactions between motoneurons.     

Contractility of mdx skeletal muscle after denervation and devascularization.

To evaluate the regenerative capacity of mdx skeletal muscle, changes in contractile properties of the fast-twitch extensor digitorum longus (EDL) of normal and mdx mice were studied at 7 and 16 weeks of age, following denervation and devascularization (DD) at 4 weeks of age. At 7 weeks, DD EDL of both strains showed significantly decreased isometric twitch and tetanus tensions compared with their non-DD controls. By 16 weeks, normal operated muscle exhibited a recovery of 57% and 58% of absolute tetanus and twitch tensions while the mdx EDL recovered remarkably to 96% and 99% of 7-week values. At 7 weeks, the DD EDL of both strains exhibited significantly slower time-to-peak (TTP) and one-half relaxation time (1/2RT). By 16 weeks, TTP and 1/2RT of the mdx DD EDL no longer differed from non-DD controls, but the normal EDL showed slowed TTP. No differences were found in the maximum velocity of shortening (Vo) or in posttetanic potentiation (PTP). Following DD, there was an increase in resistance to fatigue in both strains at 7 weeks. This resistance persisted at 16 weeks in the normal mouse, but the operated mdx EDL returned to normal. It would appear that following a denervation/devascularization insult, the mdx EDL is able to recover contractile characteristics to a remarkably larger extent than normal EDL.     

A sensitive period in gestation for nicotine acceleration of neuromuscular maturation

Nicotine, administered to pregnant Sprague-Dawley rat dams during gestation, altered the maturation of the developing extensor digitorum longus (EDL) muscle-peroneal nerve complex of the 2-week-old offspring. Initial isometric twitch time to peak and rate of rise of tension and tetanus time to peak tension of the group treated with nicotine during gestational (G) days G3-G8 were faster than controls indicating accelerated maturation of EDL muscle. Contractile parameters of the group treated with nicotine during G9-G13, a period immediately prior to muscle innervation, differed little from control. Nicotine administered during G14-G21 significantly increased twitch and tetanus tension and twitch rate of rise. There was little to no effect on motor unit size or number of motor units with nicotine during any of the three prenatal periods. Thus, while late prenatal nicotine administration primarily altered the strength of muscle, early gestational administration of nicotine increased the rate of muscle development suggesting a possible sensitive period for the accelerative action of nicotine on muscle maturation at this time.     

Na+ currents near and away from endplates on human fast and slow twitch muscle fibers

Fast and slow twitch muscle fibers have distinct contractile properties. Here we determined that membrane excitability also varies with fiber type. Na⁺ currents (I_NA) were studied with the loose-patch voltage clamp technique on 29 histochemically classified human intercostal skeletal muscle fibers at the endplate border and <200 μm from the endplate (extrajunctional). Fast and slow twitch fibers showed slow inactivation of endplate border and extrajunctional I_NA and had increased I_NA at the endplate border compared to extrajunctional membrane. The voltage dependencies of I_NA were similar on the endplate border and extrajunctional membrane, which suggests thatboth regions have physiclogically similar channels. Fast twitch fibers had larger I_NA on the endplate border and extrajunctional membrane and manifest fast and slow inactivation of I_NA at more negative potentials than slow twitch fibers. For normal muscle, the differences between I_NA on fast and slow twitch fibers might: (1) enable fast twitch fibers to operate at high firing frequencies for brief periods; and (2) enable slow twitch fibers to operate at low firing frequencies for prolonged times. Disorders of skeletal membrane excitability, such as the periodic paralyses and myotonias, may impact fast and slow twitch fibers differently due to the distinctive Na⁺ channel properties of each fiber type. © 1993 John Wiley & Sons, Inc.     

Influence of temperature on the characteristics of summation of isometric mechanical responses of mammalian skeletal muscle

Characteristics of summation of isometric mechanical responses elicited by two supramaximal nerve stimuli were examined at different temperatures (18 to 38°C) in a fast twitch (flexor digitorum longus) and a slow twitch (soleus) muscle, in situ, in the cat hind leg. At 37 to 38°C, both muscles developed a peak tension that was more than twice the twitch tension when a second stimulus was applied shortly after the first. The rate of rise of tension in such a summated response was also about, or more than, twice that of the twitch tension, and the time to peak tension was about 1.5 times the time to peak of the twitch. The peak tension in summated responses decreased with increasing the stimulus interval so that, when the interval was equal to the time to peak twitch, the tension developed was significantly less. Cooling significantly alters these characteristics in the fast twitch muscle, so that, at 20 to 22°C, the tension, the rate of rise of tension, and the time to peak tension enhancement produced by a second stimulus are markedly reduced; little or no change in the characteristics is seen with cooling of the slow twitch muscle. These results are examined in the light of some observations made on the pattern of summation in frog sartorius muscle at different temperatures (2 to 28°C). Cooling significantly increases the degree of activation occurring during the twitch in the fast twitch muscle fibers, whereas it has little or no effect on the degree of mechanical activation in the slow twitch muscle fibers. Furthermore, the results suggest that the tension, the rate of rise of tension, and, perhaps the time to peak tension of the isometric twitch in both the fast twitch and the slow twitch muscle fibers at 37 to 38°C are limited by the degree of mechanical activation and that a second stimulus applied shortly after the first can greatly overcome the effect of that limitation.     

Excitation-contraction uncoupling. The effect of hyperosomolar glycerol solution and antrolene sodium on mammalian muscle in vitro.

In the rat hemidiaphragm, in vitro dantrolene sodium irreversibly reduced twitch tension up to 20% of the original value in 20 minutes of exposure. The phenomenon was dose related. Dantrolene sodium did not affect (1) resting membrane potential (IMP); (2) miniature endplate potential frequency of amplitude; or (3) endplate potential amplitude decrement of repetitively stimulated partially curarized muscle. A combination of hyperosmolar solutions of glycerol and dantrolene sodium abolished contraction of mammalian muscle with no demonstrable effect on neuromuscular transmission or muscle fiber RMP. With this technique, prolonged recording from repetitively activated muscle fibers is possible.