title: PAC1hop receptor activation facilitates catecholamine secretion selectively through 2-APB-sensitive Ca(2+) channels in PC12 cells.
abstract: PACAP is a critical regulator of long-term catecholamine secretion from the adrenal medulla in vivo, however the receptor or pathways for Ca(2+) entry triggering acute and sustained secretion have not been adequately characterized. We have previously cloned the bovine adrenal chromaffin cell PAC1 receptor that contains the molecular determinants required for PACAP-induced Ca(2+) elevation and is responsible for imparting extracellular Ca(2+) influx-dependent secretory competence in PC12 cells. Here, we use this cell model to gain mechanistic insights into PAC1hop-dependent Ca(2+) pathways responsible for catecholamine secretion. PACAP-modulated extracellular Ca(2+) entry in PC12 cells could be partially blocked with nimodipine, an inhibitor of L-type VGCCs and partially blocked by 2-APB, an inhibitor and modulator of various transient receptor potential (TRP) channels. Despite the co-existence of these two modes of Ca(2+) entry, sustained catecholamine secretion in PC12 cells was exclusively modulated by 2-APB-sensitive Ca(2+) channels. While IP3 generation occurred after PACAP exposure, most PACAP-induced Ca(2+) mobilization involved release from ryanodine-gated cytosolic stores. 2-APB-sensitive Ca(2+) influx, and subsequent catecholamine secretion was however not functionally related to intracellular Ca(2+) mobilization and store depletion. The reconstituted PAC1hop-expessing PC12 cell model therefore recapitulates both PACAP-induced Ca(2+) release from ER stores and extracellular Ca(2+) entry that restores PACAP-induced secretory competence in neuroendocrine cells. We demonstrate here that although bPAC1hop receptor occupancy induces Ca(2+) entry through two independent sources, VGCCs and 2-APB-sensitive channels, only the latter contributes importantly to sustained vesicular catecholamine release that is a fundamental characteristic of this neuropeptide system. These results emphasize the importance of establishing functional linkages between Ca(2+) signaling pathways initiated by pleotrophic signaling molecules such as PACAP, and physiologically important downstream events, such as secretion, triggered by them.
authors.affiliation: Section on Molecular Neuroscience, Laboratory of Cellular and Molecular Regulation, National Institutes of Mental Health, Bethesda, Maryland 20892, United States.
authors.forename: Tomris
authors.initials: T
authors.lastname: Mustafa
authors.forename: James
authors.initials: J
authors.lastname: Walsh
authors.forename: Maurizio
authors.initials: M
authors.lastname: Grimaldi
authors.forename: Lee E
authors.initials: LE
authors.lastname: Eiden
mesh_terms: D000818:Animals; D001896:Boron Compounds / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000378:metabolism*; D020746:Calcium Channels, L-Type / Q000378:metabolism; D020013:Calcium Signaling; D002417:Cattle; D004095:Dihydropyridines / Q000494:pharmacology; D007295:Inositol Phosphates / Q000378:metabolism; D009638:Norepinephrine / Q000378:metabolism*; D016716:PC12 Cells; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide / Q000494:pharmacology; D020033:Protein Isoforms / Q000378:metabolism; D051381:Rats; D051237:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide, Type I / Q000378:metabolism*; D012433:Ryanodine / Q000494:pharmacology
chemical_list: D001896:Boron Compounds; D002121:Calcium Channel Blockers; D015220:Calcium Channels; D020746:Calcium Channels, L-Type; D004095:Dihydropyridines; D007295:Inositol Phosphates; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D020033:Protein Isoforms; D051237:Receptors, Pituitary Adenylate Cyclase-Activating Polypeptide, Type I; D012433:Ryanodine; C038806:1,4-dihydropyridine; C109986:2-aminoethoxydiphenyl borate; D002118:Calcium; D009638:Norepinephrine
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