Neurotransmitter release and its facilitation in crayfish

Abstract

Quantal synaptic currents were recorded at nerve terminations on the opener muscle of crayfish using a macropatch-clamp electrode, and the release was elicited by depolarizing current pulses applied to the terminal through the same electrode. After 2 ms depolarization pulses at low temperature, release started with about 2 ms delay after the onset of depolarization, and the maximum rate of release occurred at about 4 ms delay. Large variations in Ca inflow during the pulses were concluded from the facilitation of test EPSCs. The time course of release proved to be remarkably invariant in spite of large changes in release. If a conditioning train of depolarization pulses preceded the test pulse, release due to the test pulse was facilitated up to 60-fold, but the shapes of distributions of quantal delays were practically not affected by this facilitation. Facilitation by the conditioning trains must have raised the [Ca]_i level at the onset of the test pulse. The invariance of the time course of release with respect to the level of [Ca]_i cannot be explained by theories in which [Ca]_i alone controls the time course of release.

The time courses of reactions controlling release were explored by mathematical analysis and simulation. A reaction scheme in which the activation of “release sites” directly by depolarization had rate limiting control on the release reactions, in which rise of [Ca]_i only was a promoting cofactor, and in which a cooperative reaction involving the complex of release sites and Ca_i, (SCa_i) was one of the final steps eliciting release, was able to predict the delayed onset of release and the substantial latency between the end of the depolarization pulse and the maximum of the rate of release. Reaction schemes in which the direct effect of depolarization on release occurred at one or more steps following the entry of Ca could be excluded generally by showing conflict with the experimental findings.