The importance of the BAR domain of endophilin in synaptic vesicle endocytosis was tested in presynaptic microinjection experiments in the lamprey giant synapse. Antibodies as well as Fab fragments directed to the BAR domain caused a stimulus-dependent decrease in the number of synaptic vesicles along with an accumulation of shallow clathrin coated pits in the periactive zone. Moreover, the isolated BAR domain protein also caused an accumulation of shallow-coated pits in the periactive zone, in addition to appearance of narrow tubules in synaptic regions. The BAR domain of endophilin is thus required for efficient progression of the synaptic vesicle cycle. Synapse 64:556–560, 2010. © 2010 Wiley-Liss, Inc.