Abstract

Spontaneous miniature end-plate potentials (m.e.p.p.s) were recorded intracellularly in rat diaphragm muscle preparations bathed in calcium-free Ringer's solution containing 0.5 mM Mg⁺⁺ and 10 mM ethylene glycol bis (β-aminoethyl ether)-N, N'-tetraacetic acid for 1 hour and then exposed to batrachotoxin (BTX). At 37°c, BTX increased m.e.p.p frequency from 0.83 to about 150 sec^-1 and partially depolarized the muscle membrane within 15 minutes. At 28°C, BTX also produced membrane depolarization but increased the spontaneous m.e.p.p. frequency only to 5 sec^-1. In the calcium-free Ringer's solution, veratridine increased the spontaneous m.e.p.p. frequency significantly at 37°C but had little effect on m.e.p.p. frequency at 28°C. Ouabain also increased spontaneous m.e.p.p. frequency at 37°C and caused only a slight postsynaptic membrane depolarization. High extracellular concentrations of potassium (20 mM) had little or no effect on the spontaneous m.e.p.p. frequency in the calcium-free Ringer's solution at either 28 or 37°C. The effects of BTX and veratridine were due to an increase in sodium permeability (P_Na) as they were antagonized by tetrodotoxin. In phrenic nerve-diaphragm muscle preparations pretreated with 15 mM Mg⁺⁺-Ringen's solution at 34°C, BTX abruptly blocked the end-plate potential in 23 minutes, while the nerve action potential remained unaffected for over 1 hour. When the end-plate potential was blocked the frequency of spontaneous m.e.p.p.s was either normal of increased. It appears that the invasion of the nerve terminal by the action potential is prevented by a certain critical degree of depolarization of the presynaptic membrane that is less than the degree of depolarization required to induce an increase in m.e.p.p. frequency. The drug-elicited increase in PNa appears to be only an initial step in excitation-secretion coupling, since at 28°C BTX caused only membrane depolarization while at 37°C BTX caused both membrane depolarization and increased m.e.p.p. frequency. These results are compatible with the proposal that under calcium-free conditions, spontaneous transmitter release depends on an internal calcium pool, presumably located in the presynaptic membrane, which may be activated at a critical temperature by drug-elicited increases in internal sodium concentration.