The generation of climbing fibre responses in cerebellar Purkinje cells has been analysed in co-cultured slices of rat cerebellum and inferior olive. Complex spikes were evoked in Purkinje cells by climbing fibre activation or by intrasomatic injection of depolarizing current pulses. Microfluorometric measurements of cytosolic free calcium ([Ca2+]i) by means of intracellularly injected fura-2 combined with intracellular recordings revealed that both types of complex spikes were accompanied by a transient rise in [Ca2]i, which was most prominent at dendritic locations. Synaptically induced Ca2+ transients were completely and reversibly abolished by 6-cyano-7-nitroquinoxaline-2–3-dione (CNQX, 5 μM), an antagonist of the ionotropic action mediated by non-N-methyl-d-aspartate excitatory amino acid receptors. Ca2+ transients evoked by injections of depolarizing current pulses were not affected by CNQX. These observations indicate that Ca2+ transients induced by climbing fibre activity are generated by voltage-gated Ca2+ channels, which are activated by a CNQX-sensitive synaptic depolarization.