N-type Ca²⁺ channels bind directly to the synaptic core complex of VAMP/synaptobrevin, syntaxin, and SNAP-25. Peptides containing the synaptic protein interaction ("synprint") site caused dissociation of N-type Ca²⁺ channels from the synaptic core complex. Introduction of synprint peptides into presynaptic superior cervical ganglion neurons reversibly inhibited synaptic transmission. Fast EPSPs due to synchronous transmitter release were inhibited, while late EPSPs arising from asynchronous release following a train of action potentials were increased and paired-pulse facilitation was increased. The corresponding peptides from L-type Ca²⁺ channels had no effect, and the N-type peptides had no effect on Ca²⁺ currents through N-type Ca²⁺ channels. These results are consistent with the hypothesis that binding of the synaptic core complex to presynaptic N-type Ca²⁺ channels is required for Ca²⁺ influx to elicit rapid, synchronous neurotransmitter release.