Vesicle-associated proteins and quantal release at single active zones of amphibian (Bufo marinus) motor-nerve terminals

A study was made to determine the disposition of vesicle-associated protein s (syntaxin, SV2, SNAP-25) and calcium channels with respect to the spatial extent of spontaneous and evoked quantal release within regions of amphibi an motor-nerve terminal branches delineated by FM1-43 stained vesicle clust ers (blobs). Discrete concentrations of vesicles revealed similar to 2 mu m apart along the length of terminal branches through FM1-43 staining were i dentical in size and spacing to those identified along terminal branches wi th SV2 antibody (AbSV2). Fluorescent antibodies to syntaxin 1 (AbS), SNAP-2 5 (AbS25) and the calcium channel alpha 1B subunit (Ab alpha 1B) were found in relatively high concentrations coincident with the AbSV2 blobs. Three e xtracellular recording electrodes were placed in the vicinity of individual FM1-43 blobs, and an algorithm was used to determine the spatial origin of miniature endplate potentials (MEPPs) and EPPs together with their relativ e amplitudes. MEPPs and EPPs originated throughout the region stained by FM 1-43 but not elsewhere; amplitude-frequency distributions of MEPPs and EPPs were similar for all FM1-43 blobs with average coefficients of variation o f no less than 0.28. A linear relationship existed between the size of an F M1-43 blob, measured as the integrated extent of FM1-43 staining of a blob, and the frequency of MEPPs as well as the probability of EPPs from the blo b. There was a proximo-distal gradient in the size of FM1-43 blobs along th e length of single terminal branches, suggesting a gradient in release prob ability along the branches. The frequency distribution of the distances bet ween blobs was approximately Gaussian, whereas the frequency distribution o f the size of blobs was highly skewed and was best fitted with a gamma dist ribution. It is concluded that there are correlations among the extent of l abeling of SNAP-25, syntaxin and calcium channels at a release site, the st ore of vesicles to be found there, and the probability of spontaneous and e voked quantal release.