Although the calcium/calmodulin-activated phosphatase calcineurin may dephosphorylate many endocytic proteins, it is not considered a key molecule in mediating the major forms of endocytosis at synapses-slow, clathrin-dependent and the rapid, clathrin-independent endocytosis. Here we studied the role of calcineurin in endocytosis by reducing calcium influx, inhibiting calmodulin with pharmacological blockers and knockdown of calmodulin, and by inhibiting calcineurin with pharmacological blockers and knock-out of calcineurin. These manipulations significantly inhibited both rapid and slow endocytosis at the large calyx-type synapse in 7- to 10-d-old rats and mice, and slow, clathrin-dependent endocytosis at the conventional cultured hippocampal synapse of rats and mice. These results suggest that calcium influx during nerve firing activates calcium/calmodulin-dependent calcineurin, which controls the speed of both rapid and slow endocytosis at synapses by dephosphorylating endocytic proteins. The calcium/calmodulin/calcineurin signaling pathway may underlie regulation of endocytosis by nerve activity and calcium as reported at many synapses over the last several decades.