Dantrolene and the effect of temperature on the spontaneous release of transmitter at the frog neuromuscular junction

Summary The characteristic effect of temperature on m.e.p.p. frequency at the amphibian neuromuscular junction is unaltered by the presence of Dantrolene (an agent that is believed to reduce the efflux of Ca²⁺ from intracellular stores) or by changes in [Ca²⁺]_o. It is concluded that temperature affects the release system directly, with a transition temperature at about 16°C.     

Evolution in time of transmitter release at frog neuromuscular junction]

It has previously been shown that the unitary quantal end-plate potentials observed at synapses blocked by low Ca++ high Mg++ ringer, belong to distinct clusters according to their amplitude, time to peak and latency characteristics. These clusters correspond probably to distinct releasing units dispersed along the presynaptic terminal branches. The distribution versus time of the occurence of unitary potentials belonging to one latency cluster has been studied over long lasting evoked nerve activity (stimulation frequencies: 1 to 10 Hz). It was observed that transmitter release at one releasing site is statistically periodic with emitting periods separated by rest periods. At a given end-plate the period of emitting activity seems to be independent from one emitting site to another.     

Changes in transmitter release at frog neuromuscular junction induced by 4-aminopyridine]

4-aminopyridine (4-AP) at micromolar concentrations, increases the end-plate potential amplitude in curarized preparations and the mean quantal content in every preparation tested, but the spontaneous release is not modified by 4-AP. These results can explain the anticurare activity observed in the wole animal or in vitro. 4-AP prolongs the falling phase of the muscle action potential without change in the muscle membrane potential.     

Activity increases quantal size at the frog neuromuscular junction

Two h of nerve stimulation at 10 Hz or of elevated spontaneous release in hypertonic solution increased the size of miniature end-plate potentials (m.e.p.p.'s) and currents at the frog neuromuscular junction, probably by increasing the amount of acetylcholine in a quantum. Increases in quantal size may modulate synaptic transmission.     

Activity increases quantal size at the frog neuromuscular junction

Summary Two h of nerve stimulation at 10 Hz or of elevated spontaneous release in hypertonic solution increased the size of miniature end-plate potentials (m.e.p.p.'s) and currents at the frog neuromuscular junction, probably by increasing the amount of acetylcholine in a quantum. Increases in quantal size may modulate synaptic transmission.Acknowledgments. Supported by grant 10320 from the NINCDS and by a grant from the MDA. We are grateful to Dr I.S. Cohen for participating in some of the experiments and for helpful criticism, and to Drs C. Clausen and G. Baldo for help with the computer. Some of the results were presented at a meeting of the Physiological Society.     

Morphological evidence for tracer uptake at the active zones of stimulated frog neuromuscular junction

Lanthanum or ferritin added to the fixative were found in small and non-coated vesicles located at active zones in nerve-muscle preparations stimulated by potassium during cold aldehyde fixation. The presence of labeled vesicles at active zones supports the hypothesis that a double process of exo-endocytosis might occur under moderate stimulation conditions.     

The effect of charatoxin, 4-methylthio-1,2-dithiolane,on the frog sartorius neuromuscular junction

Summary The insecticide 4-methylthio-1,2-dithiolane, named charatoxin, blocks the frog muscular twitches elicited through the neuromuscular junction. The activity level and the course of inhibition is comparable to that of 4-dimethyl-amino-1,2-dithiolane, nereistoxin.The authors are grateful to Dr N. Jacobsen, Cheminova A/S for synthesizing the toxins and the 2 analogues.     

The effect of charatoxin, 4-methylthio-1,2-dithiolane, on the frog sartorius neuromuscular junction

The insecticide 4-methylthio-1,2-dithiolane, named charatoxin, blocks the frog muscular twitches elicited through the neuromuscular junction. The activity level and the course of inhibition is comparable to that of 4-dimethyl-amino-1,2-dithiolane, nereistoxin.     

Existence of different populations of unitary evoked postsynaptic potentials at the frog neuromuscular junction]

The postsynaptic response to monoquantic evoked transmitter release (mean quantal content about 0.3) has been studied at temperatures from 10 to 23 degrees C. The delay between nerve stimulation (1 to 10 Hz) and the unitary postsynaptic potentials fluctuates by steps. The existence of preferential delay sites can always be detected (mean number 13.5 +/- 3.1). Identical delay unitary postsynaptic potentials often shows identical amplitude and identical time to peak. These results suggest that few emitting sites are preferentially activated along the nerve terminal at low level release during long lasting stimulation. The "single process" assumption used in statistical studies of transmitter release is probably oversimplified.     

Zinc antagonizes the effect of botulinum type A toxin at the mouse neuromuscular junction

Summary Zn²⁺ (10–100 M) elevated the frequency of miniature end-plate potentials (MEPPs) in the mouse diaphragm. The effect did not depend on external Ca²⁺. Botulinum type A toxin (BTX_A, 50 ng/ml) abolished MEPPs almost completely within 30 min. Zn²⁺ (100 M) restored MEPPs and increased their frequency after they had been abolished by BTX_A in Ca²⁺-free solutions. The antagonistic effect of Zn²⁺ in the Ca²⁺-free solution was reduced by exposing the diaphragm to the toxin in the Ca²⁺-free solutions containing high K⁺. Thus, the action of BTX_A is probably enhanced by depolarization of the motor nerve terminals.     

Zinc antagonizes the effect of botulinum type A toxin at the mouse neuromuscular junction

Zn2+ (10-100 microM) elevated the frequency of miniature end-plate potentials (MEPPs) in the mouse diaphragm. The effect did not depend on external Ca2+. Botulinum type A toxin (BTXA, 50 ng/ml) abolished MEPPs almost completely within 30 min. Zn2+ (100 microM) restored MEPPs and increased their frequency after they had been abolished by BTXA in Ca2+ -free solutions. The antagonistic effect of Zn2+ in the Ca2+ -free solution was reduced by exposing the diaphragm to the toxin in the Ca2+ -free solutions containing high K+. Thus, the action of BTXA is probably enhanced by depolarization of the motor nerve terminals.     

Spontaneous miniature activity and gradation of transmission at the neuromuscular junction

Zusammenfassung Aus den bisher veröffentlichten experimentellen Befunden über presynaptische, für die neuromuskuläre Erregungsübertragung massgeblich erscheinende Vorgänge wird geschlossen, dass die Förderung der Erregungsübertragung in direkter Beziehung zur Grösse des Aktionspotentials in den motorischen Nervenendigungen steht, während die Frequenz der spontanen Miniaturentladungen an sich kein eindeutiges Mass für die Intensität des Übertragungsvorganges von Nervimpulsen zum Muskel darstellt.     

Depletion of synaptic vesicles at the frog (Rana pipiens) neuromuscular junctions by tetraphenylboron

Summary At the frog neuromuscular junction tetraphenylboron produced a decrease in miniature endplate potential amplitude, accompanied by a decrease in the number but not the size of synaptic vesicles.This work was supported by NIH Grants Nos NS-07740 and NS-05707 to R. L. P. and was done during the tenure of a Research Fellowship of Muscular Dystrophy Associations of America, a Wellcome Research Travel Grant and a grant from the Carnegie Trust for the Universities of Scotland to I. G. M. The assistance of Dr.R. M. Schnitzler, W. G. Boldosser andP. M. Spannbauer is gratefully acknowledged.     

Depletion of synaptic vesicles at the frog (Rana pipiens) neuromuscular junctions by tetraphenylboron.

At the frog neuromuscular junction tetraphenylboron produced a decrease in miniature endplate potential amplitude, accompanied by a decrease in the number but not the size of synaptic vesicles.     

Presynaptic effect of the neuro-muscular transmitter

Zusammenfassung Nach transversaler Durchschneidung der Muskelfasern des isolierten Rattenzwerchfelles auf jeder Seite der Endplattenregion wurde die elektrische Aktivität im stimulierten Nervus phrenicus nach Zusatz von Prostigmin, DFP und zum Teil auch Acetylcholin studiert. Die orthodromen Nervenimpulse wurden von antidromer Aktivität im Nerv gefolgt, auch dann, wenn durch Demarkationsvorgänge das Ruhepotential des Muskels bis auf 20 mV heruntergebracht und die elektrische Aktivität im Muskel gelöscht war. Dies scheint für eine praesynaptische Wirkung des Acetylcholins zu sprechen.