Time-Dependent Effects of the Neuropeptide PACAP on Catecholamine Secretion: Stimulation and Desensitization

Pituitary adenylyl cyclase-activating polypeptide (PACAP) is a potent endogenous secretagogue for chromaffin cells. We previously reported that PACAP coupled to the PAC1 receptor to evoke dihydropyridine-sensitive early (15 to 20 minutes) catecholamine secretion and cAMP response element binding protein-mediated trans-activation of the secretory protein chromogranin A promoter in PC12 pheochromocytoma cells. In this report, we studied whether the secretory and transcriptional responses elicited by PACAP were subject to desensitization. We found that PACAP evoked distinct immediate (initial, 0 to 20 minutes) and long-lasting (20 to 180 minutes) effects on catecholamine secretion. Initial secretory and chromogranin A trans-activation responses induced by PACAP were desensitized in a dose-dependent fashion after preexposure of cells to PACAP, and the IC50 doses of PACAP for desensitization were ≈18- to ≈32-fold lower than the EC50 activating doses for secretion or transcription. Desensitization of the initial secretion response was associated with decreased Ca influx through L-type voltage-operated Ca channels. Acute exposure to PACAP also triggered long-lasting (up to 3 hours), extracellular Ca-dependent, pertussis toxin-insensitive catecholamine secretion; indeed, even after short-term (20 minutes) exposure to PACAP and removal of the secretagogue, PC12 cells continued to secrete norepinephrine up to 76.9±0.22% of cellular norepinephrine content after 3 hours. A phospholipase C-β inhibitor (U-73122) blocked this extended secretory response, which was dependent on low-magnitude Ca influx resistant to several L-, N-, P/Q-, or T-type Ca channel antagonists, but sensitive to Zn, Ni, Cd, or to the store-operated Ca channel blocker SKF96365. A less than additive effect of the sarcoendoplasmic reticulum Ca-ATPase inhibitor thapsigargin plus PACAP on this sustained secretion also supported a contribution of store-operated Ca entry to the sustained secretory response. We propose that PACAP-evoked secretion and transcription are subject to homologous desensitization in PC12 cells; however, PACAP also induces long-lasting secretion, even under dose and time circumstances in which acute, dihydropyridine-sensitive secretion has been desensitized. Although initial secretion is mediated by an L-type voltage-operated Ca channel, extended secretion may involve a store-operated Ca channel that is activated through a Gq/11/phospholipase C-β/phosphoinositide signaling pathway.