The regulation of insulin secretion from pancreatic beta cells is a multifaceted, complex and coordinated process. In addition to glucose, the beta cell relies on the presentation of hormones, neurotransmitters and other fuels to adjust insulin secretory patterns and responses to insulin requirements. Its failure to properly analyze incoming information and to appropriately couple this to insulin secretion results in a failure of glucose homeostasis and, in the worst case, Type II non- insulin dependent diabetes mellitus (NIDDM). The transduction systems that allow the beta cell not only to anticipate insulin requirements but to respond to stimulation by various agonists with graded increments of hormone output have been the subject of intense investigation. Previous studies suggest that information flow in the phosphoinositide (PI) cycle occupies a particularly unique role in determining how the beta cell responds to different agonists. Our working hypothesis is that beta cell PI hydrolysis participates a) in the biphasic pattern of insulin secretion observed in fuel - or neurohumoral-stimulated islets, b) in the amplified secretory response noted with islets primed by both neurohumoral agonists such as acetylcholine or cholecystokinin and fuel agonists such as glucose or monomethylsuccinate, and c) in the process of suppression, desensitization or third phase release which characterizes the response of islets chronically exposed to several agonists including high glucose (glucose toxicity), monomethylsuccinate, acetylcholine or cholecystokinin. The activation of phospholipase C, protein kinase C and, in the case of the latter enzyme, its ability to phosphorylate its protein substrates assume importance in these responses. Freshly isolated islets, free from exocrine contamination will be employed. They display vigorous insulin secretory responses to fuel or neurohumoral stimulation comparable to those observed in vivo or with the perfused pancreas preparation. They can be subjected to further detailed biochemical analysis using monoclonal or polyclonal antibodies directed at isozymes of phospholpase C, protein kinase C, and several of its established protein substrates, and diacylglycerol kinase. Furthermore, and similar to findings made in vivo, beta cell sensitivity to stimulation can be either augmented or reversibly suppressed with the appropriate manipulation and the contribution of these enzymes to the responses observed can be assessed. Our goal is to elucidate the contribution of events proximal and distal to PI hydrolysis in the regulation of insulin secretion from pancreatic beta cells. Since a reversible failure of beta cells to appropriately sense glucose characterizes Type II diabetes, particular emphasis will be placed on establishing the contribution of this transduction pathway to disordered secretion observed from islets exposed to modest increments in the extracellular glucose concentration. Most important from a clinical perspective, we will establish the conditions which restrain the development of third phase release or, if established, manipulations which facilitate or accelerate its reversal with the restoration of normal beta cell chemosensitivity.
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