Ca²⁺ influx through voltage-gated channels initiates the exocytotic fusion of synaptic vesicles to the plasma membrane. Here we show that RIM binding proteins (RBPs), which associate with Ca²⁺ channels in hair cells, photoreceptors, and neurons, interact with α_1D (L type) and α_1B (N type) Ca²⁺ channel subunits. RBPs contain three Src homology 3 domains that bind to proline-rich motifs in α₁ subunits and Rab3-interacting molecules (RIMs). Overexpression in PC12 cells of fusion proteins that suppress the interactions of RBPs with RIMs and α₁ augments the exocytosis triggered by depolarization. RBPs may regulate the strength of synaptic transmission by creating a functional link between the synaptic-vesicle tethering apparatus, which includes RIMs and Rab3, and the fusion machinery, which includes Ca²⁺ channels and the SNARE complex.