Ultrafast endocytosis (UFE) is a clathrin-independent endocytic pathway triggered by sustained exocytosis that recovers membrane and vesicular proteins in tens of milliseconds at neuronal synapses (Miller Watanabe et al., 2013). Following exocytosis, uncoated endocytic pits localize to the boundaries of periactive zones (PAZs) at nerve terminals (Watanabe et al., 2013). The endocytic structures have characteristic size, quite different to synaptic vesicles. How this fastest of all endocytic pathways is activated, how these endocytic structures are generated, and the origin of their size and spatial localization remains unexplained. Here, we show UFE is activated by directed membrane flow driven by membrane tension gradients. Synaptic release from nerve terminals occurs at specialized ∼200 nm active zones where synaptic vesicles are docked, primed and fused with the plasma membrane. Crucially, active zones lack actin cortex, whereas the surrounding PAZ has high actin cortex density (Ogunmowo et al., 2023). Using computational modeling we find synaptic vesicle fusion generates membrane tension gradients that drive membrane flow which is directed to the actin-rich PAZ by membrane-cortex adhesion. At the PAZ boundary, accumulated membrane lowers tension and triggers a membrane instability that within ∼20 ms nucleates endocytic structures which mature into uncoated pits ∼1.5-fold bigger than synaptic vesicles within ∼40 ms. The localization, the nucleation and maturation times and the endocytic structure sizes are all in quantitative agreement with experiments in optogenetically stimulated mouse hippocampal neurons (Watanabe et al., 2013). When exocytic stimulation is sustained, we find continued membrane fusion in the active zone activates a reservoir of multiple uncoated endocytic pits in the PAZ. This is consistent with the reservoir of endocytic pits observed at neuromuscular junctions following fusion of multiple synaptic vesicles in the active zone (Miller and Heuser, 1984).
Huang et al. (Sun,) studied this question.