Spontaneous Miniature Endplate Potentials Research Articles

Acetylcholine release at identified nerve terminals in the organ‐cultured frog neuromuscular preparation.

1. The frog cutaneous pectoris neuromuscular preparation was maintained in organ culture for a few days, at either 24 or 14 degrees C. The synaptic activity of identified nerve terminals was repeatedly examined in order to describe the sequence and time course of changes from normal synaptic activity to a complete synaptic silence. 2. We found that the 'transient stage' (stage II, according to Ko, 1981) consists of at least three distinct periods each characterized by a unique trend of change in synaptic activity. The initial change involved a simultaneous decay of both spontaneous and nerve stimulation-evoked release of acetylcholine (sub-stage II1). Subsequently, the frequency of spontaneous miniature endplate potentials (MEPPs) increased gradually, while the evoked release continued its monotonous decay (sub-stage II2). During the third sub-stage spontaneous MEPPs, but no evoked endplate potentials (EPPs), were observed (sub-stage II3). 3. Statistical properties of acetylcholine release in still-transmitting junctions at sub-stage II2 and in non-transmitting junctions at sub-stage II3 were investigated. MEPPs with skewed amplitude histograms and bursting behaviour were evident at both sub-stages. However, the incidence and the extent of these distortions were higher in the non-transmitting junctions. 4. An inverse relation between the quantal content of evoked release and the rate of spontaneous secretion was found in the transmitting junctions. 5. These results suggest that some of the deterioration of synaptic activity in organ culture is caused by an impairment of the release process itself. Possible cellular mechanisms are discussed.

Organic calcium channel antagonists provoke acetylcholine receptor autodesensitization on train stimulation of motor nerve

The effects of nicardipine and other organic Ca 2+-channel antagonists on the responses induced by indirect train stimulation (3 s, 50–100 Hz) were studied in mouse phrenic nerve diaphragm preparations. Nicardipine at 1–10μM, which alone did not affect single or tetanic contractions or the amplitude of evoked endplate potentials and spontaneous miniature endplate potentials, caused tetanic contraction to fade completely in the presence of 0.3 μM neostigmine or 50 μM diisopropylfluorophosphate. In combination with these anticholinesterases, nicardipine caused a severe run-down and shortening of endplate potentials in 1–2 s. This effect on endplate potentials was dependent on stimulus frequency and on extracellular Ca 2+. The effect was accelerated by intracellular injection of Ca 2+, but retarded by injection of EGTA. The amplitudes of miniature endplate potentials and the evoked endplate depolarization were also depressed during repetitive stimulation. On termination of repetitive stimulation, all postsynaptic responses, including evoked endplate potentials, miniature endplate potentials and single twitches, recovered to pre-train level in 3–10s. These results suggest that the postsynaptic nicotinic receptors had lost the functional activity during repetitive stimulation. The time-courses of the aforementioned changes initiated by repetitive stimulation were similar to the fast phase of desensitization induced by acetylcholine. The irreversible action of α-bungarotoxin on acetylcholine receptor was attenuated in the presence of nicardipine and neostigmine if repetitive stimulation was applied. The same effects were observed with other organic Ca 2+-channel antagonists (diltiazem, verapamil and nifedipine) as well as agonist (methyl-1,4-dihydro-2,6-dimethyl-3-nitro-4-(2-trifluoromethylphenyl)-pyridine-5-carboxylate, BAY K8644), but not with Mn 2+, theophylline or caffeine. It is inferred that organic Ca 2+-channel antagonists interact directly with acetylcholine receptor ion channel, enhance its autodesensitization liability and thus cause extinction of endplate potentials on repetitive stimulation.

Effects of organic Ca antagonist, diltiazem, on neuromuscular transmission.

The effects of the organic "Ca antagonist" diltiazem on the frog neuromuscular junction was investigated. The addition of diltiazem to the bath solution increased the frequency of spontaneous miniature endplate potentials (MEPPs) and quantal content. The amplitude of the endplate current (EPC) recorded from a d-tubocurarine-blocked endplate was significantly decreased by diltiazem (20-200 microM). However, the amplitude of miniature endplate current (MEPC) was relatively unchanged by diltiazem. The same degree of decrease in the decay time constant was observed in both EPC and MEPC. ACh potentials induced by repetitive iontophoresis were depressed gradually by low concentrations of diltiazem (2, 5 microM), suggesting enhanced desensitization. Using a patch-clamp method, the effects of diltiazem on ACh-activated single current in chick cultured myotubes were examined. The addition of diltiazem to the bath solution shortened the mean open time in a dose-dependent manner between 10 and 50 microM. The voltage-dependence of mean open time disappeared in the presence of diltiazem, with little change in the channel conductance. The present experiments suggest that diltiazem acts on the ACh-activated channel as an open channel blocker.

Blocking of repetitive response from a neuromuscular preparation of Tenebrio Molitor by phosphotriesters

1. Diethylphosphate esters of m-cresol, o-, m- and p-nitrophenols, 2-nitro- p-, 4-nitro- m- and 5-nitro- o-cresols, thymol, 3,5-xylenol, α-naphthol, p-hydroxybiphenol, p-cyanophenol, p-chlorophenol and 3,4,5-trichlorophenol were synthesized and tested for their inhibitory activity toward the neuromuscular preparation of meal worm. T. molitor. 2. Remarkable depression of excitatory postsynaptic potentials (EPSPs), possibly due to acetylcholinesterase inhibition, was observed to occur by p-nitrophenol, 2-nitro- p-cresol, p-chlorophenol and 3,4,5-trichlorophenol derivatives of diethylphosphate ester, each at a concentration from 10 −5−10 −6M. 3. The spontaneous repetitive miniature end-plate potentials (MEPPs) on a post synaptic membrane was predominantly induced by p-cyanophenol, m-cresol, 4-nitro- m- and 5-nitro- o-cresols esters of the diethylphosphate. Following inducement of the spontaneous excitation, neural transmission was blocked completely.

Lambert-Eaton myasthenic syndrome: The lack of short-term in vitro effects of serum factors on neuromuscular transmission

Serum was obtained from 7 patients with the Lambert-Eaton myasthenic syndrome (LES), 3 patients with small-cell carcinoma of the lung (SCCL), and 9 healthy control subjects. Serum samples were applied in vitro to the rat neuromuscular junction (for 1–3 h for control LES sera; 4 h for SCCL sera), following which the pre- and postjunctional physiological effects of serum factors were studied in the presence of 10 mM [Mg 2+] o. All sera produced a marked reduction in the frequency of spontaneous miniature end-plate potentials (MEPPs), while causing slight to moderate changes in MEPP amplitude. There were no consistent changes in the quantum content of the impulse-evoked end-plate potentials, though the serum from one LES patient significantly and reversibly inhibited the evoked quantal release. No significant effect was found when a human intercostal muscle was exposed to serum from another LES patient for 2 h. Therefore, when applied in vitro on a short-term basis, the putative LES autoantibodies do not consistently react with voltage-dependent calcium channels in the motor nerve terminal and thus fail to reproduce the physiologic abnormality of the syndrome. We suggest that the pathogenic IgG molecules may require more than 3 h of incubation in order to gain access to, and inhibit the function of, the prejunctional Ca 2+ channels.

Stationary and non-stationary occurrences of miniature end plate potentials are well described as stationary and non-stationary Poisson processes in the mollusc Navanax inermis

Protractor muscles in the gastropod mollusc Navanax inermis exhibit typical spontaneous miniature end plate potentials with mean amplitude1.71 ± 1.19 (standard deviation) mV. The evoked end plate potential is quantized, with a quantum equal to the miniature end plate potential amplitude. When their rate is stationary, occurrence of miniature end plate potentials is a random, Poisson process. When non-stationary, spontaneous miniature end plate potential occurrence is a non-stationary Poisson process, a Poisson process with the mean frequency changing with time. This extends the random Poisson model for miniature end plate potentials to the frequently observed non-stationary occurrence. Reported deviations from a Poisson process can sometimes be accounted for by the non-stationary Poisson process and more complex models, such as clustered release, are not always needed.

In vitro formation of neuromuscular junctions between adult Rana muscle fibres and embryonic Xenopus neurons.

Adult muscle fibres of the frog Rana temporaria were cultured with neurons from embryos of the frog Xenopus laevis. Electron microscopical and electro-physiological examination of the cultures showed that hetero-specific (Xenopus-Rana) neuromuscular junctions were formed in vitro. Nerve processes, without any Schwann cell covering, made contacts anywhere along a muscle fibre, and the junctions resembled those seen during early regeneration of neuromuscular synapses in situ. Functional contacts, as inferred by the presence of spontaneous miniature endplate potentials, or currents, were more common if the muscle fibres were denervated prior to culturing with neurons. Miniature endplate currents (m.e.p.cs) had a skewed amplitude distribution, with many small events lost in the recording noise, and their mean amplitude was much smaller than that of m.e.p.cs in the original lumbricalis muscle. The time constant of decay of m.e.p.cs in the hetero-specific junctions formed in vitro was several times longer than the decay of m.e.p.cs in the original muscle. Analysis of membrane current noise elicited by ionophoretically applied acetylcholine (ACh) suggests that the slower decay of m.e.p.cs in the junctions formed in vitro is due to a prolonged lifetime of the channels opened by ACh and to repetitive activation of ACh-receptors, which becomes possible because of a comparative lack of cholinesterase in the junctions.

Statistics of neuromuscular transmitter release in young and old mouse muscle.

1. It was reported previously that in limb muscles of old (27-30 months) CBF-1 mice, quantal content (m) of evoked transmitter release was increased compared to that in young (9-12 months) mice. In diaphragm muscles there was no change with age. The object of the present study was to determine whether the age-related increase in transmitter release was due to increase in the binomial parameter n or the parameter p. The analysis also involved consideration of goodness-of-fit between observed and expected binomial distribution of the data. 2. Spontaneous miniature end-plate potentials (m.e.p.p.s) and evoked end-plate potentials (e.p.p.s) were recorded with intracellular techniques from soleus and diaphragm muscles bathed in low-Ca high-Mg medium. The goodness-of-fit between the observed e.p.p amplitude distribution and that expected from a binomial distribution was evaluated by chi 2 test. 3. In different muscles and at different ages, the percentage of fibres with binomial e.p.p. distributions varied from 17 to 44%, even though in all fibres there was a similar proportionality between direct quantal content and the reciprocal of the square of the coefficient of variation of e.p.p. amplitudes. In addition, apparent graphical agreement between observed and theoretical binomial e.p.p. distributions was often not substantiated by the chi 2 criterion. 4. In soleus muscles from young mice, lowering the stimulus frequency from 10 to 0.5 Hz and shortening the train length from 250 to 100 pulses increased the prevalence of binomial e.p.p. distributions, but the same result was not obtained in diaphragm or soleus muscles from old mice. If the mean amplitude of groups of 10 e.p.p.s in any train showed any drift (greater than 10%) then that train was excluded from the results. Thus, in order to make valid age comparisons, only fibres with binomial e.p.p. distributions were analysed further. 5. There was no change with age in m, n or p in diaphragm muscles, but in soleus muscles from old animals a nearly 2-fold increase in n entirely accounted for the increase in m. 6. If, as proposed by others, n represents the number of release sites, then the ageing soleus neuromuscular junction may have increased numbers or length of active zones or associated membrane components.

Reduction of quantal size by vesamicol (AH5183), an inhibitor of vesicular acetylcholine storage

Isolated unparalysed mouse phrenic nerve-hemidiaphragm preparations were indirectly stimulated in the presence of 2-(4-phenylpiperidino)cyclohexanol (AH5183, vesamicol). Spontaneous miniature endplate potentials were subsequently studied. They exhibited a large depression of amplitude which was more profound at higher stimulation frequencies and drug concentration. No postjunctional effects of the drug were observed. Since the drug blocks storage of acetylcholine by isolated synaptic vesicles, it is argued that the results support the theory of vesicular release of acetylcholine.

Sustained transmitter output by increased transmitter turnover in limb muscles of old mice

The ability of neuromuscular junctions in old animals to maintain tetanic output was tested in phasic and tonic limb muscles and the physiologic mechanism of maintenance was elucidated by analysis of the turnover of a false transmitter during prolonged tetani. Transmitter release during and after tetani was compared in limb muscles of young (8-9 month) and old (28-30 month) male CBF-1 mice. Amplitudes of end-plate potentials (epp's) in curarized preparations and of spontaneous miniature end-plate potentials (mepp's) were measured in vitro at 30 degrees C in soleus and extensor digitorum longus (edl) muscles. In both young and old soleus muscles, epp amplitude was maintained at about 45% of resting level during the latter part of trains of 1,200 stimuli at 10 Hz but recovered to about 90% control within a few seconds after stimulation ceased. In edl muscles of young mice, epp amplitudes during a 20 Hz train of 1,200 impulses steadily declined to about 20% of control and gradually recovered over 2 min after the tetanus. In old edl muscles, tetanic decay of the epp's was greater and recovery slower than in young muscles, but absolute epp amplitudes were invariably greater. During trains of 6,000 impulses at 10 Hz, plateau epp amplitude decayed to 40-50% in young soleus muscle and 30-40% control in old muscle, but recovery was similar and absolute epp amplitudes were greater in old soleus muscle. A false transmitter precursor, homocholine (HoCh), was used to investigate the mechanism of this prolonged output, and, therefore, the use of HoCh in this system was first validated.(ABSTRACT TRUNCATED AT 250 WORDS)

The origin of (+)-tubocurarine resistance in dystrophic mice.

Intracellular recording, twitch responses and radio-ligand binding techniques were used to study the causes of resistance to (+)-tubocurarine (curare) of extensor digitorum longus (EDL) muscles from dystrophic mice (129 ReJ/strain). The indirectly evoked twitch response of muscles from dystrophic mice was more resistant to block by curare than the twitch response of muscles from normal littermates. The IC50 (concentration producing 50% inhibition of stimulus-evoked contractions) values for the curare block of muscle twitch were 0.78 +/- 0.03 microM and 1.32 +/- 0.05 microM (mean +/- 95% confidence limits) for muscles from normal and dystrophic mice, respectively. There was no difference between muscles from normal and dystrophic mice in the number of alpha-bungarotoxin binding sites per endplate. The amplitudes of both spontaneous miniature endplate potentials (m.e.p.ps) in unblocked preparations and of evoked endplate potentials (e.p.ps) in 1.91 microM curare were greater in muscles from dystrophic mice than in muscles from normal mice. The ratio dystrophic/normal was greater for the e.p.p. amplitudes than for the m.e.p.p. amplitudes. The quantum content of e.p.ps in magnesium-blocked and in cut-fibre preparations was greater in muscles from dystrophic mice than in muscles from normal littermates. Calculation of the binomial parameters n and p in the cut-fibre preparations indicated that this increased quantum content was caused by an increase in the value of p. It is concluded that at least part of the increased resistance to curare of the indirectly evoked twitch response of muscles from dystrophic mice is due to an increase in the quantum content of e.p.ps in these muscles.

Passively transferred Lambert-Eaton syndrome in mice receiving purified IgG.

Purified IgG antibodies were prepared by ion-exchange chromatography from the plasma of a patient with nonneoplastic form of the Lambert-Eaton myasthenic syndrome (LES). The antibodies were injected into mice with daily doses of 0.15-10 mg for 20-22 days, following which the integrity of neuromuscular transmission was assessed in vitro in phrenic nerve-diaphragm muscle preparations. The injected animals manifested electrophysiologic features of human LES, which were characterized by: dose-dependent reduction in the quantal content of nerve-evoked endplate potentials, an abnormally small increase in the frequency of spontaneous miniature endplate potentials (MEPPs) with elevated [K+]o, and normal MEPP amplitude with no evidence of postjunctional deficiency. Crude immunoglobulins (Igs) from the same patient and two LES patients with associated malignancy similarly transferred the defects in quantal transmitter release. In contrast, animals receiving Igs from control subjects or from a patient with small-cell carcinoma of the lung manifested no functional impairment of neuromuscular transmission. Instead, the evoked release in these animals was significantly enhanced relative to that found in normal untreated mice. These results suggest that an IgG antibody produces the presynaptic impairment that is characteristic of LES and support the concept that LES with and without cancer has an autoimmune pathogenesis.

Increase in polyneuronal innervation in frog muscle after muscle injury.

The proportion of polyneuronal innervation was evaluated electrophysiologically in curare-blocked frog cutaneous pectoris muscles after local injury to the muscle fibres on one side. Focal polyneuronal innervation was revealed by recording end-plate potentials evoked by a gradual increase in the stimulus intensity applied to the motor nerve. An increase in the proportion of focally polyneuronally innervated muscle fibres appeared in the injured muscle 3-5 days after injury. The difference between the values obtained 3-5 days and 7-9 days (31 and 38%, respectively) and the control value (18%) was highly significant. A similar increase in the proportion of pluri-innervated muscle fibres was observed in the contralateral muscle, but after a longer period. The different components of complex end-plate potentials (e.p.p.s) usually had similar latencies and rise times in control and experimental muscles. This may indicate that the axons had similar conduction velocities and that synapses were located close to each other. A repeated muscle fibre section 24 h after the initial injury resulted in an enhanced polyneuronal innervation (52%) 7-9 days after the first section. The experiments were repeated on partially blocked muscles in order to detect small e.p.p.s with an amplitude similar to that of spontaneous miniature end-plate potentials (m.e.p.p.s). The proportion of polyneuronally innervated fibres estimated by this technique in control muscles approximated 40%. Polyneuronal innervation was also found to be significantly increased in cut muscles 7-9 days after muscle injury and a week later in contralateral muscles. Combined silver and cholinesterase staining was used to detect morphologically polyneuronal innervation. The comparison of morphological and electrophysiological data indicated that the increase in polyneuronal innervation after muscle injury is likely due to nerve sprouting and formation of new synapses. The results suggest that the signal for nerve sprouting originates from the damaged muscle cell and that it is transferred transneuronally to the contralateral side.

Enhancement by Anemonia sulcata toxin II of spontaneous quantal transmitter release from mammalian motor nerve terminals

J. Molgo, M. Lemeignan and F. Tazieff-Depierre. Enhancement by Anemonia sulcata toxin II of spontaneous quantal transmitter release from mammalian motor nerve terminals. Toxicon 24, 441 – 450, 1986. — The action of Anemonia sulcata toxin II (ATX-II) on spontaneous quantal transmitter release from motor nerve terminals was investigated by recording miniature end-plate potentials (MEPPs) from isolated mouse phrenic nerve - hemidiaphragm nerve - muscle preparations. ATX-II (3.2 μM) when applied for 30 – 40 min to junctions bathed in a normal ionic medium enhanced about one hundred fold the rate of spontaneous MEPPs. Concomitantly, ATX-II depolarized the muscle fiber. The effect of the toxin on MEPP frequency was markedly reduced when junctions were exposed to Na-deficient solutions or pre-treated with dantrolene sodium (10 μM). ATX-II (0.24 – 3.2 μM) increased MEPP rate in junctions exposed to a Ca-free medium containing 2 mM EGTA and 2 mM Mg 2+ in a dose- and time-dependent manner. Tetrodotoxin (0.2 – 1 μM) prevented the effects of ATX-II on MEPP frequency and on the resting membrane potential of muscle fibers. Tetrodotoxin also antagonized the acceleration of MEPP induced by ATX-II. The experimental findings suggest that ATX-II acts to increase quantal transmitter output from motor nerve terminals by enhancing Na + influx through tetrodotoxin-sensitive presynaptic channels, since ATX-II action does not appear to depend upon entry of Ca 2+ from the extracellular medium. It is likely that ATX-II, by increasing intraterminal Na + concentration, may trigger calcium release from internal stores.

Synergistic interaction of 4-aminopyridine with neostigmine at the neuromuscular junction

The pre- and postsynaptic events at the neuromuscular junction were studied in vitro in rat skeletal muscle exposed to clinically significant concentrations of 4-aminopyridine (4-AP), neostigmine or combinations of the two drugs. Simultaneous application of 4-AP and neostigmine produced increases in the amplitudes of nerve-evoked end-plate potentials which were significantly greater than the summed effects of the drugs applied individually. Such synergism was present at the junctions where transmission was blocked either postsynaptically by d-tubocurarine or presynaptically by low [Ca 2+] 0 and high [Mg 2+ 0. Quantal content analysis in the latter preparation indicated that the evoked release of acetylcholine was potentiated significantly more than the amplitude of spontaneous miniature end-plate potentials, suggesting that the site of synergism is predominantly presynaptic. For symptomatic relief and long-term management of the neuromuscular junction disorders, we propose a combined medication of aminopyridine and anticholinesterase at reduced dosages. Such therapy would minimize adverse effects and be particularly effective in the treatment of such presynaptic disorders as the Lambert-Eaton myasthenic syndrome and botulism.

Is a nicotinic influence involved in denervation-induced depolarization of muscle?

The neurotrophic role of acetylcholine (ACh) in the denervation-dependent decline of muscle resting membrane potential (RMP) was evaluated. Freshly dissected rat hemidiaphragms with short or long (2 cm) nerve stumps attached ("-N" and "+N" preparations, respectively) were incubated in organ culture in the presence or absence of the nicotinic blockers, alpha-bungarotoxin (alpha-BTX) or d-tubocurarine (curare). Subsequently, RMPs and miniature endplate potentials (MEPPs) of the junctional region were measured. Spontaneous MEPPs disappeared with a half-life of 12 and 20 hr in -N and +N preparations, respectively. A 10- to 15-mV depolarization of RMP was observed between 15 and 20 hr in -N muscles and between 24 and 28 hr in +N muscles. This time course of disappearance of spontaneous potentials and of membrane depolarization agrees well with that observed in vivo. Although nicotinic transmission was blocked from the initiation of the incubation period in alpha-BTX- or curare-treated muscles, no acceleration of RMP decline in -N muscles in vitro was observed. Moreover, in +N preparations the effect of the nerve stump in delaying RMP depolarization persisted despite the continuous presence of alpha-BTX or curare. Since excess ACh triggers a lysosomal proteolytic response at the nerve-muscle junction and since this may occur early in denervation, the possible role of a nicotinic-induced proteolytic mechanism was tested in vitro with the potent protease inhibitor leupeptin. This inhibitor did not delay or prevent the denervation-dependent alterations.(ABSTRACT TRUNCATED AT 250 WORDS)

Influence of divalent cations on the phospholipase‐independent action of beta‐bungarotoxin at frog neuromuscular junctions.

The influence of different divalent cations on the phospholipase-independent inhibition of transmitter release caused by beta-bungarotoxin (beta-BuTx), has been investigated by measuring the frequency of spontaneous miniature end-plate potentials (m.e.p.p.s) at frog neuromuscular junctions. After adding the toxin to normal calcium Ringer solution the m.e.p.p. frequency fell quickly to very low values. This was followed by an increase in frequency characterized by bursts of m.e.p.p.s. The temperature had a negligible effect on the speed of the first inhibition. In Ringer solutions where calcium had been substituted by other divalent cations (5 mM) in the presence of ethyleneglycol-bis-(beta-aminoethylether)N, N'-tetraacetic acid (EGTA, 1 mM), this beta-BuTx-induced decrease in m.e.p.p. frequency was markedly slower. The potency of cations in promoting the initial phase of toxin action was in the sequence: calcium greater than magnesium greater than strontium congruent to cobalt greater than manganese. This phospholipase-independent inhibition of transmitter release followed approximately first-order kinetics, suggesting that it depends mainly on toxin concentration. In the absence of any divalent cations in the Ringer solution beta-BuTx had practically no effect on m.e.p.p. frequency. It appears that beta-BuTx requires divalent cations in order to bind to motor nerve terminals and exert its initial inhibitory action on spontaneous release of transmitter quanta.

Software that detects and analyzes bioelectrical events by assessing the fit of modeled subintervals

A software system is described which has been used to identify, average, and otherwise analyse different kinds of electrophysiological events. Spontaneous miniature end-plate potentials were identified by assessing the fit of subintervals within suspected event-containing intervals to flexibly defined model subevents. It was thus practical to apply complex, explicit, and stable criteria in the estimation of the frequency and mean time course of these spontaneous events. This software also mediated the objective rejection of noise-contaminated elements from samples of either evoked or spontaneous potentials. Average event time courses were comprised of a series of estimates of the standard deviations as well as the means. Each analysis generated thousands of individual event amplitudes which were used to improve estimates of quantal release and response parameters. This software allowed more complete analysis of tape-recorded experiments which conserved test compounds and animals, and enabled the observation of simultaneous and equal degrees of action upon spontaneous and evoked potentials, which suggested that a common mechanism was involved.

Passive transfer of the Lambert-Eaton myasthenic syndrome: neuromuscular transmission in mice injected with plasma.

Mice were given 1 to 67 daily injections of whole plasma from a control subject and a patient with the Lambert-Eaton myasthenic syndrome (LES), and the parameters of neuromuscular transmission in diaphragm muscles were studied in vitro. In animals that received 41 to 59 injections of LES plasma, mean quantum content of the endplate potentials, as measured under the conditions of low [Ca2+]o (1.2 mM) and high [Mg2+]o (10 mM), was reduced to 30% of the control, or 40% of the value found in normal untreated mice. When [K+]o was elevated from 5 mM to 17.5 mM, the frequency of spontaneous miniature endplate potentials increased by a factor of 98 and 105 respectively in the control and normal mice but only 39 times in the LES plasma recipients. Despite these presynaptic abnormalities, no evidence of postsynaptic deficit, nerve conduction failure, or inability of the muscle fibers to produce normal action potentials was found. These electrophysiologic features in mice thus represent a reproduction of the human condition, supporting the concept of humoral involvement in the pathophysiology of LES. Passively transferred LES in mice is a faithful animal model of the human disease which will be useful in exploring the molecular basis of the presynaptic impairment.

Reinnervation of normal and dystrophic skeletal muscle

A comparative study was conducted of resting membrane potential (RMP), extrajunctional acetylcholine (ACh) sensitivity, spontaneous and neurally evoked transmitter release, and directly and indirectly elicited action potentials in posterior latissimus dorsi (PLD) muscles of normal (line 412) and dystrophic (line 413) chickens during reinnervation after nerve crush. Control (nondenervated) dystrophic muscle fibers had a significantly greater RMP (−77.2 vs. −74.1 mV), extrajunctional ACh sensitivity (0.34 vs. 0.04 mV/nC), and incidence of repetitive firing of directly elicited action potentials than did normals. Miniature end-plate potential (MEPP) amplitude in dystrophic fibers was significantly reduced (0.26 vs. 0.38 mV). Six to 7 days after nerve crush, muscle fibers from both lines of chickens showed a significant reduction in RMP and an increase in extrajunctional ACh sensitivity. During this time spontaneous MEPPs were absent and the incidence of repetitive firing was decreased. No significant difference was noted between chicken lines in any of the properties studied. The return of normal properties associated with reinnervation occurred primarily between days 9 and 40. Repolarization of the RMP was clearly evident by day 9 in both lines, but dystrophic fibers showed a slightly earlier and greater degree of repolarization. Similarly, initial decreases in extrajunctional ACh sensitivity and the reappearance of MEPPs were observed on day 9 with dystrophic and day 12 with normal fibers. Neurally elicited action potentials were first recorded on day 11 for dystrophic and day 12 for normal fibers. Finally, multiple firing of directly elicited action potentials associated with reinnervation attained the same incidence (20 to 21% of fibers) on day 12 in dystrophic and day 14 in normal fibers. The results suggest that dystrophic chicken muscle has an enhanced capacity for reinnervation following nerve crush.