Differential motion dynamics of synaptic vesicles undergoing spontaneous and activity-evoked endocytosis

Synaptic vesicle exo- and endocytosis are usually driven by neuronal activity but can also occur spontaneously. The identity and differences between vesicles supporting evoked and spontaneous neurotransmission remain highly debated. Here we combined nanometer-resolution imaging with a transient motion analysis approach to examine the dynamics of individual synaptic vesicles in hippocampal terminals under physiological conditions. We found that vesicles undergoing spontaneous and stimulated endocytosis differ in their dynamic behavior, particularly in the ability to engage in directed motion. Our data indicate that such motional differences depend on the myosin family of motor proteins, particularly myosin II. Analysis of synaptic transmission in the presence of myosin II inhibitor confirmed a specific role for myosin II in evoked, but not spontaneous, neurotransmission and also suggested a functional role of myosin II-mediated vesicle motion in supporting vesicle mobilization during neural activity.