Glucose-induced insulin secretion from purified beta-cells. A role for modulation of Ca2+ influx by cAMP- and protein kinase C-dependent signal transduction pathways.

Abstract

The effects of activation of cAMP- and protein kinase C-dependent signal transduction pathways were investigated on intracellular Ca2+ concentration ([Ca2+]i), cAMP content and insulin secretion from beta-cells purified by fluorescence-activated cell sorting from normal rat islets. The secretion of insulin from suspensions of purified beta-cells was dependent on glucose concentration and hormonal signals, including cAMP and activators of protein kinase C. Microfluorimetric measurement of [Ca2+]i with the fluorescent Ca2+ indicator fura-2 indicated that beta-cells differed immensely in their individual responsiveness to glucose stimulation. An increase in [Ca2+]i occurred in approximately 70% of beta-cells, whereas approximately 30% of beta-cells were nonresponsive to a glucose stimulus. Elevation of cAMP levels by theophylline or glucagon transformed nonresponsive beta-cells into cells which displayed marked increases in [Ca2+]i, and beta-cells which exhibited glucose-induced changes in [Ca2+]i showed further increases in [Ca2+]i and in the amplitude of Ca2+ oscillations. Carbachol and 12-O-tetradecanoylphorbol-13-acetate, activators of protein kinase C, did not induce any alterations in intracellular cAMP levels; nonetheless, these agents increased both the number of beta-cells which exhibited glucose-induced changes in [Ca2+]i and the amplitude of oscillations. The ability of cAMP or activators of protein kinase C to increase [Ca2+]i in single beta-cells was directly correlated with the ability of beta-cell suspensions to secrete insulin in response to a glucose stimulus. These results suggest that both cAMP- and protein kinase C-dependent pathways may regulate Ca2+ entry into beta-cells, possibly via voltage-dependent Ca2+ channels. Thus, this may represent a common mechanism whereby these different signal transduction pathways potentiate glucose-induced insulin secretion from beta-cells.