A readily retrievable pool of synaptic vesicles

Overview & Synopsis

Although clathrin-mediated endocytosis (CME) is thought to be the predominant mechanism of synaptic vesicle (SV) recycling, it seems to be too slow for fast recycling. Therefore, it was suggested that a pre-sorted and pre-assembled pool of SV proteins on the presynaptic membrane might support a first wave of fast CME. In this study we monitored the temporal dynamics of such a 'readily retrievable pool' of SV proteins in rat hippocampal neurons using a novel probe. Applying cypHer 5, a new cyanine dye-based pH-sensitive exogenous marker, coupled to antibodies against luminal domains of SV proteins we could reliably monitor SV recycling and for the first time demonstrate the preferential recruitment of a surface pool of SV proteins upon stimulated endocytosis. Using fluorescence nanoscopy of surface labeled synaptotagmin 1, we could resolve the spatial distribution of the surface pool at the periactive zone in hippocampal boutons, which represent putative sites of endocytosis.