In enteric synaptosomes of the rat, the role of voltage-dependent Ca²⁺ channels in K⁺-induced VIP release and nitric oxide (NO) synthesis was investigated. Basal VIP release was 39 ± 4 pg/mg, and cofactor-substituted NO synthase activity was 7.0 ± 0.8 fmol · mg¹ · min¹. K⁺ depolarization (65 mM) stimulated VIP release Ca²⁺ dependently (basal, 100%; K⁺, 172.2 ± 16.2%; P < 0.05, n = 5). K⁺-stimulated VIP release was reduced by blockers of the P-type (-agatoxin-IVA, 3 × 10⁸ M) and N-type (-conotoxin-GVIA, 10⁶ M) Ca²⁺ channels by ~50 and 25%, respectively, but not by blockers of the L-type (isradipine, 10⁸ M), Q-type (-conotoxin-MVIIC, 10⁶ M), or T-type (Ni²⁺, 10⁶ M) Ca²⁺ channels. In contrast, NO synthesis was suppressed by -agatoxin-IVA, -conotoxin-GVIA, and isradipine by ~79, 70, and 70%, respectively, whereas Ni²⁺ and -conotoxin-MVIIC had no effect. These findings are suggestive of a coupling of depolarization-induced VIP release primarily to the P- and N-type Ca²⁺ channels, whereas NO synthesis is presumably dependent on Ca²⁺ influx not only via the P- and N- but also via the L-type Ca²⁺ channel. In contrast, none of the Ca²⁺ channel blockers affected VIP release evoked by exogenous NO, suggesting that NO induces VIP secretion by a different mechanism, presumably involving intracellular Ca²⁺ stores.