A Non-cholinergic Transmitter, Pituitary Adenylate Cyclase-activating Polypeptide, Utilizes a Novel Mechanism to Evoke Catecholamine Secretion in Rat Adrenal Chromaffin Cells

Pituitary adenylate cyclase-activating polypeptide (PACAP) is the most potent non-cholinergic neurotransmitter to stimulate catecholamine secretion from rat chromaffin cells; however, the mechanism of action is not clear. We used amperometric detection of exocytosis and indo-1 monitoring of [Ca2+]i to identify PACAP actions in cultured chromaffin cells. PACAP (100 nM) required external Ca2+ to evoke secretion. However, unlike nicotine and KCl which caused immediate and relatively brief secretion, PACAP has a latency of 6.8 +/- 0.96 s to the first secretory response and secretion continued for up to 2 min. PACAP elevation of [Ca2+]i showed similar latency and often remained above base line for several minutes following a brief exposure. ZnCl2 (100 microM) selectively inhibited PACAP-stimulated secretion and [Ca2+]i with little effect on nicotine-evoked responses. Nifedipine (10 microM) had little effect on PACAP-evoked secretion but inhibited nicotine-evoked secretion by more than 80%, while omega-conotoxin (100 nM) failed to affect either agonist. PACAP-stimulated cAMP levels required 5 s to significantly increase, consistent with the latency of exocytotic and Ca2+ responses. Forskolin (10 microM) caused responses similar to PACAP. PACAP-evoked exocytosis was blocked by the protein kinase A inhibitor adenosine 3'5'-cyclic monophosphorothioate Rp-diastereomer (Rp-cAMPS). These data showed that PACAP stimulates exocytosis by a mechanism distinctly different from cholinergic transmitters that appears to involve cAMP-mediated Ca2+ influx. Differences in receptor coupling mechanisms and pharmacology of Ca2+ entry stimulated by cholinergic and peptidergic agonists support the idea that the peptidergic system maintains catecholamine secretion under conditions where the cholinergic system desensitizes or otherwise fails.