Delineation of the Chemomechanosensory Regulation of Gastrin Secretion Using Pure Rodent G Cells

Abstract

Gastrin is a key regulator of gastric acid secretion. We aimed to isolate pure G cells to identify the mechanistic basis of luminal- and strain-mediated regulation. Using gradient centrifugation and fluorescence-activated cell sorting, rat G cells were prepared and luminal, neural, hormonal, and mechanical activation of secretion and signaling pathways studied. Pure G-cell preparations (>97%) were isolated. Reverse-transcription polymerase chain reaction identified neural, hormonal, bacterial, and luminal G protein-coupled receptors, and immunostaining visualized specific sweet/bitter receptors and the tastant-associated G protein alpha-gustducin. Gastrin release was stimulated by forskolin (adenosine 3',5'-cyclic monophosphate [cAMP] inducer, 10 micromol/L; >3-fold), potentiated by 3-isobutyl-1-methylxanthine (IBMX; phosphodiesterase type 5 inhibitor and adenosine antagonist, 10 micromol/L) and phorbol myristate acetate (phorbol ester, 10 micromol/L), and inhibited by H-89 (protein kinase A inhibitor, 10 micromol/L), PD98059 (MEK1 inhibitor, 0.1 micromol/L), and wortmannin (phosphatidylinositol 3-kinase inhibitor, 1 nmol/L). Gastrin release was stimulated by neuronal G protein-coupled receptor ligands, pituitary adenylate cyclase-activating protein (20 pmol/L, >8-fold) and bombesin (0.1 micromol/L, 8-fold) through cAMP signaling. The tastants sucralose, glucose, caffeine, denatonium, and the vanilloid receptor activator capsaicin all stimulated secretion (>3-fold), as did bacterial lipopolysaccharides Salmonella enteritidis (0.24 nmol/L, 5-fold) greater than Helicobacter pylori (0.57 micromol/L, 3-fold). Secretion was associated with elevated cAMP levels (approximately 2-fold) and could be inhibited by H-89 and PD98059 and potentiated by IBMX and cholera toxin (250 microg/mL). Bacterially mediated secretion also involved activation of nuclear factor kappaB and the c-Jun-N-terminal kinase pathway. Mechanical strain stimulated (2-fold to 8-fold) gastrin release, and decreasing pH from 7.4 to 5.5 inhibited release. The adenosine receptor 2B antagonist MRS1754 inhibited mechanically induced gastrin release. G cells are luminal sampling chemomechanosensory cells whose secretion is regulated by neural, hormonal, luminal, and mechanical factors through protein kinase A activation, cAMP signaling, and mitogen-activated protein kinase phosphorylation.