1. The effects of omega-conotoxin GVIA (conotoxin), a potent inhibitor of neuronal N-type Ca2+ channels, have been examined on responses to stimulation of noradrenergic, cholinergic and non-adrenergic, non-cholinergic (NANC) nerves in a range of isolated tissues to investigate the role of conotoxin-sensitive Ca2+ channels in neurotransmission. 2. Contractions elicited by field stimulation of noradrenergic nerves in rat and mouse anococcygeus muscles, rabbit ear artery and rat vas deferens (epididymal portion) were inhibited by conotoxin. Responses to noradrenaline, and to adenosine triphosphate in the vas deferens, were not affected. 3. Positive chronotropic responses to field stimulation of noradrenergic nerves were inhibited by conotoxin in rat and mouse atria, but responses to noradrenaline and tyramine were not affected. 4. The stimulation-induced release of noradrenaline was inhibited by conotoxin in the rabbit ear artery and in rat and mouse atria. 5. Relaxations in response to stimulation of the noradrenergic perivascular mesenteric nerves were reduced or abolished by conotoxin in rat and rabbit jejunum. The response to noradrenaline in rat jejunum was not affected. 6. Contractions elicited by stimulation of cholinergic nerves were inhibited by conotoxin in rat jejunum and mouse ileum (perivascular mesenteric nerves), and in guinea-pig taenia caeci (field stimulation). Responses to acetylcholine in rat jejunum and mouse ileum were not affected. 7. Contractions elicited by stimulation of the cholinergic plus NANC pelvic nerves were inhibited by conotoxin in rabbit colon, and to a lesser extent in guinea-pig colon. The stimulation-induced contraction of the guinea-pig colon was inhibited by conotoxin by a greater proportion in the presence than in the absence of atropine. Responses to acetylcholine were not affected in the rabbit colon but were slightly reduced in the guinea-pig colon. 8. Relaxations in response to field stimulation of NANC nerves were inhibited by conotoxin in guinea-pig taenia caeci and rat gastric fundus strips, and in rat anococcygeus muscle when the tone was raised by guanethidine but not when it was raised by carbachol. The relaxations produced by sodium nitroprusside in the rat gastric fundus and anococcygeus were not affected. 9. Contractions of the rat bladder elicited by stimulation of the peri-urethral nerves, which are NANC- and cholinergically mediated, were relatively insensitive to inhibition by conotoxin. The response were almost completely abolished by tetrodotoxin. 10. The conotoxin-induced inhibitions of responses to nerve stimulation developed slowly and persisted after removal of conotoxin. The responses were almost completely abolished by tetrodotoxin. 10. The conotoxin-induced inhibitions of responses to nerve stimulation developed slowly and persisted after removal of conotoxin. 11. The inhibitory effect of conotoxin was inversely proportional to the frequency of stimulation (in several preparations) and to the Ca2+ concentration in the bathing solution (in rat vas deferens). These observations suggest that the inhibition by conotoxin of the Ca2+ influx required for excitation-secretion coupling in autonomic nerve terminals is not absolute, and can be overcome by repeated stimulation or by raising the Ca2 + concentration.