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Am J Physiol Cell Physiol 274: C1552-C1562, 1998;
0363-6143/98 $5.00
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Vol. 274, Issue 6, C1552-C1562, June 1998

Glucagon-mediated Ca2+ signaling in BHK cells expressing cloned human glucagon receptors

Lars H. Hansen1, Jesper Gromada2, Pierre Bouchelouche3, Ted Whitmore4, Laura Jelinek4, Wayne Kindsvogel4, and Erica Nishimura1

1 Department of Molecular Signaling, Hagedorn Research Institute, DK-2820 Gentofte; 2 Islet Cell Physiology, Novo Nordisk A/S, DK-2100 Copenhagen; 3 Department of Clinical Biochemistry, Roskilde County Hospital, DK-4600 Køge, Denmark; and 4 ZymoGenetics Inc., Seattle, Washington 98102

From video imaging of fura 2-loaded baby hamster kidney (BHK) cells stably expressing the cloned human glucagon receptor, we found the Ca2+ response to glucagon to be specific, dose dependent, synchronous, sensitive to pertussis toxin, and independent of Ca2+ influx. Forskolin did not elicit a Ca2+ response, but treatment with a protein kinase A inhibitor, the Rp diastereomer of 8-bromoadenosine-3',5'-cyclic monophosphothioate, resulted in a reduced glucagon-mediated Ca2+ response as well as Ca2+ oscillations. The specific phospholipase C inhibitor U-73122 abolished the Ca2+ response to glucagon, and a modest twofold increase in inositol trisphosphate (IP3) production could be observed after stimulation with glucagon. In BHK cells coexpressing glucagon and muscarinic (M1) acetylcholine receptors, carbachol blocked the rise in intracellular free Ca2+ concentrations in response to glucagon, whereas glucagon did not affect the carbachol-induced increase in Ca2+. Furthermore, carbachol, but not glucagon, could block thapsigargin-activated increases in intracellular free Ca2+ concentration. These results indicate that, in BHK cells, glucagon receptors can activate not only adenylate cyclase but also a second independent G protein-coupled pathway that leads to the stimulation of phospholipase C and the release of Ca2+ from IP3-sensitive intracellular Ca2+ stores. Finally, we provide evidence to suggest that cAMP potentiates the IP3-mediated effects on intracellular Ca2+ handling.

phospholipase C; adenosine 3',5'-cyclic monophosphate; G proteins; inositol trisphosphate; intracellular calcium stores; baby hamster kidney cells


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