Regulation by membrane potential of phosphatidylinositol hydrolysis in pancreatic islets

In pancreatic islets stimulated with carbamylcholine (carbachol), hydrolysis of both phosphatidylinositol (PtdIns) and phosphatidylinositol bisphosphate (Ptd-InsP2) occurs and can be measured as the inositol monophosphates Ins(1)P1, or Ins(4)P1, respectively (Biden, T. J., Prugue, M. L., and Davison, A. G. M. (1992) Biochem. J. 285, 541-549). Our current aim was to establish whether these two events were independently regulated. Rats islets were labeled with either [3H]inositol or [3H]arachidonic acid for measurement of InsP1s by high performance liquid chromatography or diacylglycerol by TLC, respectively. The rise in Ins(1)P1 due to carbachol (1 min) was inhibited by 50% by concomitantly raising extracellular KCl ([K+]e) from 6 to 30 mM, thereby depolarizing the islets. Similar results, obtained in the absence of extracellular Ca2+, exclude the involvement of voltage-gated Ca2+ channels. Conversely, hyperpolarization, by lowering [K+]e to 3 mM, increased by 30% the rise in Ins(1)P1. In fact, over the [K+]e range of 3 to 48 mM, stimulated Ins(1)P1 accumulation was directly proportional to the calculated membrane potential. In contrast, raising [K+]e from 6 to 48 mM exerted no significant effect on carbachol-stimulated Ins(4)P1 levels, and both Ins(1)P1 and Ins(4)P1 were unaffected in the absence of carbachol. The rises in Ins(1)P1 (but not Ins(4)P1) were also inhibited by depolarization with the sodium pump inhibitor, ouabain, or the K+ channel blocker, tolbutamide. Stimulated diacylglycerol accumulation and insulin secretion (20 min) showed a biphasic dependency on [K+]e, being less pronounced at 6 mM than at either 3 or 30 mM KCl. This reflects a selective potentiation of PtdIns and PtdInsP2 hydrolysis, due, respectively, to hyperpolarization and the gating of voltage-dependent Ca2+ channels. The differential regulation of these two hydrolytic events is probably important for independent control of the activation of protein kinase C and Ca2+ mobilization and might play a role in modulating the secretory response in vivo.