P/Q-type Ca2+ channels, which are postulated to play major roles in synaptic transmission, are regulated in a variety of ways. Ca2+ currents through P/Q-type Ca2+ channels (Cav2.1/β1a/a2δ) heterologously expressed in mammalian cells were recorded using the whole-cell patch clamp method. The oxidant H2O2 increased the current amplitude and the effect was reversed by the reducing agent dithiothreitol (DTT). The stimulatory effect of H2O2 on the Ca2+ current was mimicked by the NO donors, SNAP, and diethylamine NONOate, and reversed by the reducing agent DTT. The presence of a soluble guanylate cyclase inhibitor did not abolish the ability of SNAP to increase the Ca2+ current. Adenovirus-mediated overexpression of nitric oxide synthase in combination with application of the Ca2+ ionophore A23187 also increased the Ca2+ current amplitude and the effect was again reversed by DTT. The NOS inhibitor L-NAME abolished the stimulatory effect of A23187, and A23187 did not change the Ca2+ currents in the cells treated with control adenovirus particles. The time course of the decline of the Ca2+ current, but not of the Ba2+ current, in response to repeated epolarization was markedly slowed by adenovirus-mediated overexpression of nitric oxide synthase. The results demonstrate that nitric oxide enhances the channel activity by promoting oxidation and suggest that Ca2+, nitric oxide synthase, and nitric oxide could constitute a positive feedback loop for regulation of voltage-gated P/Q-type Ca2+ channels.