Coupling of Ca2+-triggered unclamping and membrane fusion during neurotransmitter release

Neurotransmitter (NT) release is accomplished by a machinery that unclamps fusion in response to calcium and then fuses the synaptic vesicle and plasma membranes. These are often thought of as distinct tasks assigned to non-overlapping components. Vesicle release rates have a power law dependence on [Ca2+] with an exponent of 3-5, long taken to indicate that 3-5 Ca2+ ions bind the calcium sensor Synaptotagmin to trigger release. However, dependencies at low [Ca] are inconsistent with simple sequential binding to a single Ca2+ sensor followed by a final fusion step. Here we developed coarse-grained molecular dynamics simulations of the NT release machinery accounting for Synaptotagmin-mediated unclamping and SNARE-mediated fusion. Calcium-triggered unclamping and SNARE-mediated fusion emerged from simulations as contemporaneous, coupled processes. Increasing cytosolic [Ca2+], the instantaneous fusion rate increased as SNAREpins were progressively and reversibly released by dissociation of Synaptotagmin-SNAREpin complexes. Simulations reproduced the observed dependence of release rates on [Ca2+], but the power law was unrelated to the number of Ca2+ ions required. Action potential-evoked vesicle release probabilities depended on the number of transiently unclamped SNAREpins, explaining experimental dependencies of release probabilities on both unclamping and membrane-fusing machinery components. These results describe a highly cooperative NT release machinery with intrinsically inseparable unclamping and membrane-fusing functionalities.