Multiple protein kinase pathways are involved in gastrin-releasing peptidereceptor-regulated secretion

Gastrin-releasing peptide (GRP) and its amphibian homolog, bombesin, are po tent secretogogues in mammals. We determined the roles of intracellular fre e Ca2+ ([Ca2+](i)), protein kinase C (PKC), and mitogen-activated protein k inases (MASK) in GRP receptor (GRP-R)-regulated secretion. Bombesin induced either [Ca2+](i) oscillations or a biphasic elevation in [Ca2+](i). The bi phasic response was associated with peptide secretion. Receptor-activated s ecretion was blocked by removal of extracellular Ca2+, by chelation of [Ca2 +](i), and by treatment with inhibitors of phospholipase C, conventional PK C isozymes, and MAPK kinase (MEK). Agonist-induced increases in [Ca2+](i) w ere also inhibited by dominant negative MEK-1 and the MEK inhibitor, PD8905 9, but not by an inhibitor of PKC. Direct activation of PKC by a phorbol es ter activated MAPK and stimulated peptide secretion without a concomitant i ncrease in [Ca2+](i). Inhibition of MEK blocked both bombesin- and phorbol la-myristate 13-acetate induced secretion. GRP-R-regulated secretion is ini tiated by an increase in [Ca2+](i); however, elevated [Ca2+](i) is insuffic ient to stimulate secretion in the absence of activation of PKC and the dow nstream MEK/MAPK pathways. We demonstrated that the activity of MEK is impo rtant for maintaining elevated [Ca2+](i) levels induced by GRP-R activation , suggesting that MEK may affect receptor-regulated secretion by modulating the activity of Ca2+-sensitive PKC.