Morphofunctional changes at the active zone during synaptic vesicle exocytosis

Abstract

The fusion of synaptic vesicles (SVs) with the plasma membrane (PM) proceeds through intermediate steps that remain poorly resolved. Additionally, the effect of persistent high or low exocytosis activity on intermediate steps remains unknown. Through time-resolved cryo-electron tomography, we ordered events into a sequence. Following stimulation, proximal tethered SVs rapidly form additional tethers with the PM. Simultaneously, fusion initiation occurs by membrane curvature ( buckling) of the SV and PM. It is followed by the formation of a fusion pore, and the collapse of SV membrane. At this time, membrane-proximal, but not membrane-distal, vesicles lose their interconnections, allowing them to move towards the PM. Two mutations of SNAP-25 that arrests or disinhibit spontaneous release, respectively, both caused a loss of interconnectors, while the disinhibiting mutant also caused a loss of membrane proximal multiple-tethered SVs. Overall, tether formation and connector dissolution is triggered by stimulation and respond to the spontaneous fusion rate. These morphological observations likely correspond to the transition of SVs from one functional pool to another.