Protein Kinase A-Dependent Stimulation of Exocytosis in Mouse Pancreatic β-Cells by Glucose-Dependent Insulinotropic Polypeptide

The mechanisms by which glucose-dependent insulinotropic polypeptide (GIP) stimulates insulin secretion were investigated by measurements of whole-cell Ca²⁺ currents, the cytoplasmic Ca²⁺ concentration, and cell capacitance as an indicator of exocytosis in individual mouse pancreatic β-cells maintained in short-term culture. GIP produced a 4.2-fold potentiation of depolarization-induced exocytosis. This stimulation of exocytosis was not associated with a change in the whole-cell Ca²⁺-current, and there was only a small increase (30%) in the cytoplasmic Ca²⁺ concentration [intercellular free Ca²⁺([Ca²⁺]_i)]. The stimulatory effect of GIP on exocytosis was blocked by pretreatment with the specific protein kinase A (PKA) inhibitor Rp-8-BrcAMPS. Glucagon-like peptide-I(7–36) amide (GLP-I) stimulated exocytosis (90%) in the presence of a maximal GIP concentration (100 nmol/1). Replacement of GLP-I with forskolin produced a similar stimulatory action on exocytosis. These effects of GLP-I and forskolin in the presence of GIP did not involve a change in the whole-cell Ca²⁺-current or [Ca²⁺]_i. GIP was ineffective in the presence of both forskolin and the phosphodiesterase inhibitor isobutylmethylxanthine (IBMX). Under the same experimental conditions, the protein kinase C (PKC)-activating phorbol ester 4-phorbol 12-myristate 13-acetate (PMA) stimulated exocytosis (60%). Collectively, our data indicate that the insulinotropic hormone GIP stimulates insulin secretion from pancreatic β-cells, through the cAMP/PKA signaling pathway, by interacting with the secretory machinery at a level distal to an elevation in [Ca²⁺]_i.