Evidence for two distinct processes in the final stages of neurotransmitter release as detected by binomial analysis in calcium and strontium solutions

The statistical parameters underlying acetylcholine (ACh) release were studied using Ca²⁺ and Sr²⁺ ions to promote ACh secretion. Experiments were performed at frog neuromuscular junctions using electrophysiological recording techniques. Increases in asynchronous ACh release, reflected as the frequency of occurrence of miniature end-plate potentials (MEPP_f), were evoked by high potassium depolarization in either Ca²⁺ or Sr²⁺ solutions. Increases in MEPP_f mediated by Ca²⁺ were of very low probability and well-described by a Poisson distribution whilst similar MEPP_f increases mediated by Sr²⁺ were best described as a simple binomial distribution. From the binomial distribution in Sr²⁺ solutions, values for the average probability of release (p) and the number of releasable ACh quanta (n) may be determined (whereby mean MEPP_f = np). In Sr²⁺ solutions, values of p were independent of both bin width and of the value of n, suggesting that both n and p were stationary. Calculations of p using the simple binomial distribution in Sr²⁺ solutions gave theoretical values for the third moment of the mean which were indistinguishable from the experimental distribution. These results, in conjunction with Monte Carlo simulations of the data, suggest that spatial and temporal variance do not measurably affect the analysis. Synchronous ACh release evoked by nerve impulses (end-plate potentials, EPPs) follow a simple binomial distribution in both Ca²6+ and Sr²⁺ solutions. Similar mean levels of synchronous ACh release (m, where m = np) were produced by lower values of P and higher values of n in Ca²⁺ as compared to Sr²⁺ The statistical analyses suggest the presence of two different Ca²⁺-dependent steps in the final stages of neurotransmitter release. The results are discussed in accordance with (i) statistical models for quantal neurotransmitter release, (ii) the role of Sr²⁺ as a partial agonist for evoked ACh release, and (iii) the specific loci that may represent the sites of Ca²⁺ and Sr²⁺ sensitivity.