Synapses are specialized intercellular junctions in which cell adhesion molecules connect the presynaptic machinery for neurotransmitter release to the postsynaptic machinery for receptor signalling. Neurotransmitter release requires the presynaptic co-assembly of Ca²⁺ channels with the secretory apparatus, but little is known about how synaptic components are organized. α-Neurexins, a family of >1,000 presynaptic cell-surface proteins encoded by three genes, link the pre- and postsynaptic compartments of synapses by binding extracellularly to postsynaptic cell adhesion molecules and intracellularly to presynaptic PDZ domain proteins. Using triple-knockout mice, we show that α-neurexins are not required for synapse formation, but are essential for Ca²⁺-triggered neurotransmitter release. Neurotransmitter release is impaired because synaptic Ca²⁺ channel function is markedly reduced, although the number of cell-surface Ca²⁺ channels appears normal. These data suggest that α-neurexins organize presynaptic terminals by functionally coupling Ca²⁺ channels to the presynaptic machinery.