Formation of an endophilin-Ca²⁺ channel complex is critical for clathrin-mediated synaptic vesicle endocytosis

A tight balance between synaptic vesicle exocytosis and endocytosis is fundamental to maintaining synaptic structure and function. Calcium influx through voltage-gated Ca²⁺ channels is crucial in regulating synaptic vesicle exocytosis. However, much less is known about how Ca ²⁺ regulates vesicle endocytosis or how the endocytic machinery becomes enriched at the nerve terminal. We report here a direct interaction between voltage-gated Ca²⁺ channels and endophilin, a key regulator of clathrin-mediated synaptic vesicle endocytosis. Formation of the endophlin-Ca²⁺ channel complex is Ca²⁺ dependent. The primary Ca²⁺ binding domain resides within endophilin and regulates both endophilin-Ca²⁺ channel and endophilin-dynamin complexes. Introduction into hippocampal neurons of a dominant-negative endophilin construct, which constitutively binds to Ca²⁺ channels, significantly reduces endocytosis-mediated uptake of FM 4-64 dye without abolishing exocytosis. These results suggest an important role for Ca ²⁺ channels in coordinating synaptic vesicle recycling by directly coupling to both exocytotic and endocytic machineries.