Time-dependent effects of the neuropeptide PACAP on catecholamine secretion - Stimulation and desensitization

Pituitary adenylyl cyclase-activating polypeptide (PACAP) is a potent endog enous secretagogue for chromaffin cells. We previously reported that PACAP coupled to the PAC1 receptor to evoke dihydropyridine-sensitive early (15 t o 20 minutes) catecholamine secretion and cAMP response element binding pro tein-mediated trans-activation of the secretory protein chromogranin A prom oter 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 catecholamin e secretion. Initial secretory and chromogranin A trans-activation response s induced by PACAP were desensitized in a dose-dependent fashion after pree xposure of cells to PACAP, and the IC50 doses of PACAP for desensitization were approximate to 18- to approximate to 32-fold lower than the EC50 activ ating doses for-secretion or transcription. Desensitization of the initial secretion response was associated with decreased Ca2+ influx through L-type voltage-operated Ca2+ channels. Acute exposure to PACAP also triggered lon g-lasting (up to 3 hours), extracellular Ca2+-dependent, pertussis toxin-in sensitive catecholamine secretion; indeed, even after short-term (20 minute s) exposure to PACAP and removal of the secretagogue, PC12 cells continued to secrete norepinephrine up to 76.9+/-0.22% of cellular norepinephrine con tent after 3 hours. A phospholipase C-beta inhibitor (U-73122) blocked this extended secretory response, which was dependent on low-magnitude Ca2+ inf lux resistant to several L-, N-, P/Q-, or T-type Ca2+ channel antagonists, but sensitive to Zn2+, Ni2+, Cd2+, or to the store-operated Ca2+ channel bl ocker SKF96365. A less than additive effect of the sarcoendoplasmic reticul um Ca2+-ATPase inhibitor thapsigargin plus PACAP on this sustained secretio n also supported a contribution of store-operated Ca2+ entry to the sustain ed secretory response. We propose that PACAP-evoked secretion and transcrip tion are subject to homologous desensitization in PC12 cells; however, PACA P also induces long-lasting secretion, even under dose and time circumstanc es in which acute, dihydropyridine-sensitive secretion has been desensitize d. Although initial secretion is mediated by an L-type voltage-operated Ca2 + channel, extended secretion may involve a store-operated Ca2+ channel tha t is activated through a G(q/11)/phospholipase C-beta/phosphoinositide sign aling pathway.