Analysis of the organophosphate-induced electromyographic response to repetitive nerve stimulation: paradoxical response to edrophonium and D-tubocurarine.

We studied the mechanism underlying acute organophosphate intoxication (OPI) through in-vivo and in-vitro electrophysiologic studies in rats injected with diisopropylfluorophosphate. Intoxicated rats showed weakness, repetitive compound muscle action potentials (CMAPs) in response to a single stimulus, and decremental response to repetitive nerve stimulation that was most pronounced at the second CMAP. The decrement was worsened with the administration of edrophonium, and was completely reversed by D-tubocurarine. In-vitro microelectrode studies showed no reduction in the amplitude of miniature endplate potentials (MEPPs) or in the quantal content of end-plate potentials (EPPs). However, the half-decay times of MEPPs and EPPs were significantly prolonged. Trains of stimuli induced sustained end-plate depolarization via a "staircase phenomenon" of summation of prolonged EPPs, which was enhanced by edrophonium and abolished by D-tubocurarine. These results indicate that sustained end-plate depolarization can directly account for the decrement and weakness in acute OPI.     

Enhanced neuromuscular transmission efficacy in overloaded rat plantaris muscle

To better understand the effect of muscle hypertrophy on the physiological properties of transmitter release, we investigated neuromuscular transmission (NMT) efficacy in overloaded rat plantaris muscle in situ. In the overload group, following bilateral tenotomy of plantaris synergists, rats were confined to wheel-cages. Age-matched rats in the control group were confined to plastic cages. During the terminal experiment, muscle action potentials were blocked using micro-conotoxin, and full-sized endplate potentials (EPPs) were recorded at 25, 50, and 75 HZ to determine their amplitude rundown. Quantal contents for the control and overload groups were 37.0 and 74.3, respectively (P <0.01). There was a significant group difference in EPP amplitude rundown at all frequencies examined, with increased rundown occurring in the overload group (P < 0.01). Cumulative quantal release was 139% and 153% higher in the overload group at 25 and 50 HZ, respectively (P < 0.05). Together, these data suggest the safety factor for NMT is increased by neuromuscular overload. Furthermore, these findings support and supplement previously reported activity-dependent improvements in NMT efficacy that are probably mediated via presynaptic adaptations.     

Dynamical analysis of neuromuscular transmission jitter

Utilizing prolonged axonal stimulation single fiber EMG, neuromuscular transmission becomes a time-series of interpotential intervals (IPIs). In this form, the underlying processes of neuromuscular transmission can be studied using standard numerical techniques to determine whether these processes can be described by a simple mathematical model. In particular, neuromuscular transmission jitter can be examined in this way. In this article, we attempt to determine whether healthy jitter is noise or deterministic chaos. The presence of deterministic chaos was assessed by analysis of the IPI time-series using visual inspection of both phase-space plots and their principal component dimensions, and using the Grassberger–Procaccia algorithm to determine the correlation dimension of the time-series dynamics. These graphical and mathematical techniques provided little evidence for the existence of deterministic chaos. Linear autoregression time-series prediction also failed to account for the variability of the data and IPI histograms exhibited simple gaussian distributions. These results suggest normal neuromuscular transmission jitter is the result of intrinsic noise. © 1995 John Wiley & Sons, Inc.     

In vitro microelectrode study of neuromuscular transmission in a case of botulism.

We performed in vitro microelectrode studies in the anconeus muscle biopsy of a 6-week-old infant intoxicated with Clostridium botulinum toxin B. The most striking abnormalities were the severe reduction of the endplate potential (EPP) quantal content and the marked variability of EPP latencies. The increased variability was often limited to a "single quantum" component of the EPP. Neither the amplitudes nor the frequencies of spontaneous miniature endplate potentials (MEEPs) were decreased. However, there was a wide range of amplitudes and frequencies of MEPPs. This unique combination of electrophysiologic findings indicates a severe presynaptic failure of neuromuscular transmission, which appears to result from an impairment of the process of synaptic vesicle release taking place after the stimulus induced influx of calcium into the motor nerve terminals.     

Thiamine blockade of neuromuscular transmission

The effects of thiamine on neuromuscular transmission were studied. Thiamine (0.2-2 mM) decreased the quantal height (q) of the evoked end-plate current (EPC). In the presence of 2 mM thiamine, the shape of the decay phase of the EPC remained exponential but the decay time constant increased dramatically. Thiamine changed the EPC height/holding potential and log (EPC decay time constant)/holding potential relationships from linear to non-linear. The reversal potential and the rate of rise of the EPC were unaffected. Analysis of the ACh-induced noise revealed that thiamine increased the mean channel lifetime and decreased the single channel conductance. Under normal conditions, the ACh-induced single channel conductance had no membrane voltage dependency, while thiamine caused the single channel conductance to decrease with hyperpolarization. The effects of thiamine could be explained in computer simulation of the peak EPC/membrane potential relationship by the assumption that thiamine prolonged the channel lifetime and decreased the single channel conductance with hyperpolarization. It is concluded that thiamine modifies the kinetics of ACh-receptor ion channel complex and that this effect is dependent on the membrane voltage.     

Nitric oxide modulates neuromuscular transmission during hypoxia in rat diaphragm

Hypoxia impairs neuromuscular transmission in the rat diaphragm. In previous studies, we have shown that nitric oxide (NO) plays a role in force modulation of the diaphragm under hypoxic conditions. The role of NO, a neurotransmitter, on neurotransmission in skeletal muscle under hypoxic conditions is unknown. The effects of the NO synthase (NOS) inhibitor nomega-nitro-L-arginine (L-NNA, 1 mM) and the NO donor spermine NONOate (Sp-NO, 1 mM) were evaluated on neurotransmission failure during nonfatiguing and fatiguing contractions of the rat diaphragm under hypoxic (PO2 approximately 5.8 kPa) and hyperoxic conditions (PO2 approximately 64.0 kPa). Hypoxia impaired force generated by both muscle stimulation at 40 HZ (P40M) and by nerve stimulation at 40 HZ (P40N). The effect of hypoxia in the latter was more pronounced. L-NNA increased P40N whereas Sp-NO decreased P40N during hypoxia. In contrast, neither L-NNA nor Sp-NO affected P40N during hyperoxia. L-NNA only slightly reduced neurotransmission failure during fatiguing contractions under hyperoxic conditions. Consequently, neurotransmission failure assessed by comparing force loss during repetitive nerve simulation and superimposed direct muscle stimulation was more pronounced in hypoxia, which was alleviated by L-NNA and aggravated by Sp-NO. These data provide insight in the underlying mechanisms of hypoxia-induced neurotransmission failure. This is important as respiratory muscle failure may result from hypoxia in vivo.     

Guidelines for the treatment of autoimmune neuromuscular transmission disorders

Important progress has been made in our understanding of the cellular and molecular processes underlying the autoimmune neuromuscular transmission (NMT) disorders; myasthenia gravis (MG), Lambert–Eaton myasthenic syndrome (LEMS) and neuromyotonia (peripheral nerve hyperexcitability; Isaacs syndrome). To prepare consensus guidelines for the treatment of the autoimmune NMT disorders. References retrieved from MEDLINE, EMBASE and the Cochrane Library were considered and statements prepared and agreed on by disease experts and a patient representative. The proposed practical treatment guidelines are agreed upon by the Task Force: (i) Anticholinesterase drugs should be the first drug to be given in the management of MG (good practice point). (ii) Plasma exchange is recommended as a short‐term treatment in MG, especially in severe cases to induce remission and in preparation for surgery (level B recommendation). (iii) Intravenous immunoglobulin (IvIg) and plasma exchange are equally effective for the treatment of MG exacerbations (level A Recommendation). (iv) For patients with non‐thymomatous autoimmune MG, thymectomy (TE) is recommended as an option to increase the probability of remission or improvement (level B recommendation). (v) Once thymoma is diagnosed TE is indicated irrespective of the severity of MG (level A recommendation). (vi) Oral corticosteroids is a first choice drug when immunosuppressive drugs are necessary in MG (good practice point). (vii) In patients where long‐term immunosuppression is necessary, azathioprine is recommended together with steroids to allow tapering the steroids to the lowest possible dose whilst maintaining azathioprine (level A recommendation). (viii) 3,4‐diaminopyridine is recommended as symptomatic treatment and IvIg has a positive short‐term effect in LEMS (good practice point). (ix) All neuromyotonia patients should be treated symptomatically with an anti‐epileptic drug that reduces peripheral nerve hyperexcitability (good practice point). (x) Definitive management of paraneoplastic neuromyotonia and LEMS is treatment of the underlying tumour (good practice point). (xi) For immunosuppressive treatment of LEMS and NMT it is reasonable to adopt treatment procedures by analogy with MG (good practice point).     

Efficacy of obidoxime in human organophosphorus poisoning: Determination by neuromuscular transmission studies

Six patients with organophosphorus compound intoxications developed an intermediate syndrome (weakness and fasciculations) and obidoxime was given on eight occasions. The efficacy of the acetylcholinesterase (AChE) reactivator was monitored electrophysiologically by neuromuscular transmission studies using single and repetitive nerve stimulation (20 and 50 Hz) and the activity of the serum (butyryl) cholinesterase (ChE). Dramatic electrophysiologic improvement was seen when obidoxime was given early within 12 h in 3 patients, although evidence of AChE inhibition did not subside completely. When administration of obidoxime was delayed 26 h or more after intoxication on five occasions, electrophysiologic improvement was mild or absent. In one case, 66 h after intoxication with oxydemeton-S-methyl, the neuromuscular block worsened, indicating that aging of the AChE had been completed. The electrophysiologic improvement was always accompanied with better motor function but not necessarily with improvement of the overall clinical status. Serum ChE did not predict the oxime effect at the motor endplate. In humans, the efficacy of oximes in AChE reactivation can be determined rapidly using electrophysiologic studies.© 1995 John Wiley &Sons, Inc.     

Guidelines for treatment of autoimmune neuromuscular transmission disorders

Background: Important progress has been made in our understanding of the autoimmune neuromuscular transmission (NMT) disorders; myasthenia gravis (MG), Lambert–Eaton myasthenic syndrome (LEMS) and neuromyotonia (Isaacs’ syndrome). Methods: To prepare consensus guidelines for the treatment of the autoimmune NMT disorders, references retrieved from MEDLINE, EMBASE and the Cochrane Library were considered and statements prepared and agreed on by disease experts. Conclusions: Anticholinesterase drugs should be given first in the management of MG, but with some caution in patients with MuSK antibodies (good practice point). Plasma exchange is recommended in severe cases to induce remission and in preparation for surgery (recommendation level B). IvIg and plasma exchange are effective for the treatment of MG exacerbations (recommendation level A). For patients with non‐thymomatous MG, thymectomy is recommended as an option to increase the probability of remission or improvement (recommendation level B). Once thymoma is diagnosed, thymectomy is indicated irrespective of MG severity (recommendation level A). Oral corticosteroids are first choice drugs when immunosuppressive drugs are necessary (good practice point). When long‐term immunosuppression is necessary, azathioprine is recommended to allow tapering the steroids to the lowest possible dose whilst maintaining azathioprine (recommendation level A). 3,4‐Diaminopyridine is recommended as symptomatic treatment and IvIG has a positive short‐term effect in LEMS (good practice point). Neuromyotonia patients should be treated with an antiepileptic drug that reduces peripheral nerve hyperexcitability (good practice point). For paraneoplastic LEMS and neuromyotonia optimal treatment of the underlying tumour is essential (good practice point). Immunosuppressive treatment of LEMS and neuromyotonia should be similar to MG (good practice point).     

The novel TrkB receptor agonist 7,8-dihydroxyflavone enhances neuromuscular transmission

Neurotrophin signaling at the neuromuscular junction modulates cholinergic transmission and enhances neuromuscular transmission via the tropomyosin-related kinase receptor subtype B (TrkB).A novel flavonoid, 7,8-dihydroxyflavone (7,8-DHF), selectively activates TrkB receptors. Using TrkB(F616A) mice that are susceptible to specific inhibition of TrkB activity by 1NMPP1, we show that neuromuscular transmission is enhanced by 7,8-DHF (∽32%) via activation of TrkB in diaphragm muscle. The small molecule 7,8-DHF may constitute a novel therapy to improve neuromuscular function.     

Alterations of spontaneous quantal transmitter release at the mammalian neuromuscular junction induced by divalent and trivalent ions

The present studies have demonstrated that Ca and Sr have a similar ability to support the release of spontaneous quanta of transmitter at the rat neuromuscular junction. The foreign ions Co, Pb, Zn and La have also been shown to support spontaneous release, although the time course of the increase in miniature end-plate potentials (m.e.p.p.s) caused by these ions is much slower than that caused by Ca and Sr. However, Sr, Co, Pb and Zn were unable to sustain m.e.p.p.s for prolonged periods. High extracellular K (10--15 mM) inhibited the steady state m.e.p.p. frequency in Ca and Sr concentrations above 1 mM, and also delayed the m.e.p.p. frequency increase induced by 1 mM Pb. It is suggested that K ions directly inhibit divalent ion entry into the nerve terminal.     

Neurotrophins improve neuromuscular transmission in the adult rat diaphragm

Neurotrophins modulate acute and sustained synaptic plasticity. In cultured Xenopus laevis neuromuscular junctions, neurotrophins improve neuromuscular transmission. Whether this influence exists at the mammalian neuromuscular junction is unknown. We hypothesized that neurotrophins improve neuromuscular transmission at neuromuscular junctions of adult rat diaphragm muscle fibers. A diaphragm muscle-phrenic nerve preparation was used to determine the effects of brain-derived neurotrophic factor (BDNF), neurotrophin-4 (NT-4) and K252a [tyrosine kinase (Trk) receptor inhibitor] on the extent of neuromuscular transmission failure induced by repetitive nerve stimulation. We found significant enhancement of neuromuscular transmission with BDNF or NT-4 treatment, whereas K252a treatment worsened neuromuscular transmission. In contrast, diaphragm muscle contractile and fatigue properties were unaffected by neurotrophin or K252a treatment. These results demonstrate that BDNF and NT-4 improve synaptic transmission in the adult rat diaphragm muscle, likely in a Trk-dependent fashion. Neurotrophins may constitute a novel therapeutic target to improve neuromuscular function in the diaphragm.     

Effects of lead on neuromuscular transmission in the frog

The acute effects of Pb²⁺ on synaptic transmission at the frog neuromuscular junction were measured using conventional microelectrode techniques. Experiments were performed on preparations bathed in high magnesium/low calcium Ringer solution in order to record subthreshold endplate potentials (EPPs). The effects of Pb²⁺ on the muscle membrane and postsynaptic membrane were minimal since relatively high doses of Pb²⁺ caused no significant change in the input resistance of the muscle fiber and in the amplitude of the acetylcholine (ACh) iontophoteric potential when the ACh micropipette was highly localized. However, when the ACh micropipette was moved away from the receptors, the resulting ACh potential was reduced significantly by Pb²⁺. Pb²⁺ is a potent blocker of the EPP. Extracellular recordings from motor nerve terminals showed that endplate currents (EPCs) were reduced by Pb²⁺ while the nerve terminal potentials were unaffected. Therefore, Pb²⁺ blocks evoked transmitter release at a step following the depolarization of the nerve terminal. The blocking effect on the EPP was overcome when [Ca²⁺]_o was raised. The log-log relationship between [Ca²⁺]_o (abscissa) and EPP amplitude was shifted to the right in the presence of 1 μM Pb²⁺; the x¯±S.E. slopes were 4.16 ± 0.12 (control) and 4.05 ± 0.13 (Pb²⁺). Reciprocal plots relating [Ca²⁺]_o⁻¹ to (EPP)^−1/5 confirmed that Pb²⁺ competitively antagonized the action of Ca²⁺. The dissociation constant between Pb²⁺ and the Ca²⁺ receptor site was found to be 0.99 μM. Pb²⁺ is about 3 × 10³ times more potent than is Mg²⁺, about 150 times more potent than is either Mn²⁺ or Co²⁺, and about 3 times more potent than Cd²⁺ is in blocking evoked release of ACh. After Pb²⁺ decreased the EPP, the MEPP frequency began to increase; this was probably the result of intracellular Pb²⁺ disrupting the Ca²⁺ sequestering activity of mitochondria and/or other intraterminal organelles. [Ca²⁺]_i was thereby increased and an increase in MEPP frequency followed. Decreased MEPP amplitudes were observed when the MEPP frequency had been increased by Pb²⁺. Pb²⁺ may affect most chemical synapses in a manner which is similar to its effects on the neuromuscular junction and that this may be one of its important neurotoxic effects.     

Autoantibodies in neuromuscular transmission disorders

It is a great pleasure to be asked to honour the memory of Dr. Baldev Singh by reviewing the field of autoantibodies in myasthenia gravis and other neurotransmission disorders. The neuromuscular junction (NMJ) is the site of a number of different autoimmune and genetic disorders, and it is also the target of many neurotoxins from venomous snakes, spiders, scorpions and other species. The molecular organization of the NMJ is graphically represented in Figure 1A, where different ion channels, receptors and other proteins are shown. Four of the ion channels or receptors are directly involved in autoimmune diseases. This brief review will not only concentrate on these conditions but also illustrate how their study is helping us to understand the etiology of rare but treatable neurological syndromes of the central nervous system.Open in a separate windowFigure 1(A) The neuromuscular junction showing the targets for antibodies in disease. Rapsyn is the cytoskeletal protein that clusters the acetylcholine receptors. (B) The neuromuscular junction in myasthenia gravis with AChR antibodies. The AChRs are reduced in number and there is morphological damage to the postsynaptic membrane. (C) The AChR viewed from above the membrane consists of five subunits, two alphas, one beta, one delta and either a gamma (fetal type) or epsilon (adult type). Bungarotoxin and acetylcholine bind to sites on the interfaces between the alpha subunit and adjacent subunits. Many, but not all, antibodies bind to a region known as the main immunogenic region on the alpha subunits. Mothers with babies who are born with arthrogryposis may have antibodies that bind to a gamma-subunit specific site     

Effects of levetiracetam on axon excitability and synaptic transmission at the crayfish neuromuscular junction

Levetiracetam (LEV) is a widely prescribed antiepileptic drug, but its actions on neuronal function are not fully characterized. Since this drug is believed to enter neurons by binding to a vesicular protein during endocytosis, we used motor axons of the crayfish opener neuromuscular junction to examine potential impacts of LEV on axon excitability. Two electrode current clamp from the inhibitory axon of the opener showed that LEV reduced action potential (AP) amplitude (AP_amp) and suppressed synaptic transmission, although the latter occurred with a longer delay than the reduction in AP_amp. Comparison of antidromic and orthodromic conducting APs in LEV suggested that this drug preferentially reduced excitability of the proximal axon, despite the expectation that it entered the axon at the terminals and should affect the distal branches first. Results presented here suggest that LEV modulates axonal excitability, which may in turn contribute to its antiepileptic effects.     

Fatigue and abnormal neuromuscular transmission in Kennedy's disease

We describe a patient with Kennedy's disease (X-linked bulbospinal neuronopathy) who experienced leg muscle fatigue with long-distance running. The patient also reported muscle twitching involving the face and extremities and long-standing muscle cramps. Aside from mild facial and tongue weakness (and fasciculations), his examination was normal, including completely preserved muscle strength in the extremities. Electrodiagnostic evaluation revealed evidence for a chronic motor axonopathy/neuronopathy and abnormal sensory nerve action potentials. In addition, repetitive nerve stimulation studies were normal, but neuromuscular jitter tested in the same muscle was markedly abnormal. The normal clinical strength and repetitive nerve stimulation studies in a muscle showing markedly increased neuromuscular jitter suggested a mechanism for this patient's symptoms of muscle fatigue, related to failure of neuromuscular transmission at a critical number of endplates during extremes of physical activity.     

Depression of neuromuscular transmission in methylmercury-poisoned rats: a glass microelectrode and single fiber electromyography study

Changes in neuromuscular transmission were examined in methylmercury (MeHg)-poisoned rats, given a total oral dose of 60 mg CH₃HgCl at 5 mg/kg/day. A microelectrode study was done on the 21st day. The mean quantal content and mean values of the immediately available pool of ACh in the MeHg-poisoned rats were reduced as compared to those in the control rats, but the mean values for the release probability of ACh did not differ significantly. Stimulation single fiber electromyography (SFEMG) was done on the biceps femoris muscle at 1,5, 10 and 20 Hz on the 28th day. Both a significant and consistent increase in jitter were found at increasing stimulation rates in the MeHg-poisoned rats. The SFEMG findings suggest presynaptic involvement due to accelerated depletion of ACh. We confirmed that neuromuscular transmission is depressed in MeHg-poisoned rats in vivo and in vitro.     

Incomplete recovery of endplate potential amplitude while intermittently activating rat soleus neuromuscular junctions in situ

Studies dealing with neuromuscular transmission efficacy typically employ continuous patterns of activation to demonstrate decrements in endplate potential (epp) amplitude. Recent evidence from rat diaphragm muscle has shown that including periods of quiescence to the stimulation protocol allows epp amplitude to recover between series of contractions. Whether similar recovery occurs in rat hindlimb muscle is unknown. In this study, we have measured declines in epp amplitude in rat soleus muscle during trains of stimulation evoked either continuously (10 s) or intermittently (400 ms repeated every second), using an in situ approach. As in diaphragm, we found that rest periods within the intermittent trains significantly improved neuromuscular transmission efficacy. However, unlike the diaphragm, epp amplitude recovery was incomplete even by the second train in the intermittent protocols, recovery being frequency-dependent and ranging from 40 to 50%. The results suggest that the kinetics of epp amplitude rundown and recovery may be muscle-specific, and should be considered when evaluating situations in which neuromuscular transmission efficacy may be altered.     

Myotonia and neuromuscular transmission in the mouse.

The role of neuromuscular transmission and acetylcholine receptors in the phenotypic expression of hereditary myotonia was reinvestigated in two mutants of the mouse, ADR (adr/adr) and MTO (adrmto/adrmto). Three neuromuscular blockers, curare, flaxedil, and alpha-bungarotoxin, did not prevent mechanical myotonia of EDL and soleus muscles from the two mutants. Furthermore, electrical myotonia was demonstrated in isolated ADR muscle fibers devoid of nerve endings. We conclude that neither release nor reception of acetylcholine are important for the mechanism of myotonia in mouse mutants. The previously described suppression of myotonic aftercontractions by high concentrations of curare (Muscle & Nerve 1987;10:293-298) could not be reproduced; rather, a prolongation of aftercontractions was found. The other drugs had no significant effect on myotonic aftercontractions. Because neuromuscular transmission is not involved in human myotonias, this result supports the use of myotonic mice as a model, at least for recessive generalized myotonia (Becker).     

Evaluation of neuromuscular transmission by using monopolar needle electrode

OBJECTIVE: To evaluate the value of single-fibre electromyography (SFEMG) with monopolar electrode (MNPE) in revealing neuromuscular transmission dysfunction. MATERIAL AND METHODS: We examined the extensor digitorum communis muscle by using single-fibre electrode (SFE) and MNPE sequentially, in randomly assigned 20 healthy volunteers and in 17 patients with known myasthenia gravis (MG). The high-pass filter setting was 3 kHz for MNPE. Ten individual jitter values were calculated for each electrode in every muscle. Repetitive nerve stimulation (RNS) test on trapezius muscle was performed on 15 patients. RESULTS: In controls, the mean jitter values were 27 +/- 9 (10-59) micro s with SFE, and 21 +/- 7.2 (9-56) micro s with MNPE (P = 0.001). In the MG group, the mean jitter values were 52.4 +/- 38 (12-221) micro s with SFE, and 51.8 +/- 34.7 (12-179) micro s with MNPE. Both electrodes identified junction dysfunction in 14 patients. RNS revealed decrement in four patients but 11. CONCLUSION: SFEMG with SFE is still the gold standard; however, SFEMG with MNPE is superior to RNS like SFEMG with SFE.