Glucose-induced insulin secretion is defective in type II diabetes mellitus, and the events which couple glucose recognition to insulin secretion by pancreatic islet beta cells are incompletely understood. In isolated islets, glucose induces hydrolysis of membrane phospholipid, as reflected by a rise in the mass of unesterified arachidonate, a decline of arachidonate esterified to phospholipids, accumulation of arachidonate metabolites, and accumulation of inositol-1,4,5-trisphosphate. The hypothesis that mediator substance derived from phospholipid hydrolysis participate in signal transduction in islets is the principal focus of this proposal, in which the temporal profile of secretagogue induced appearance of the mass of phospholipid-derived mediators will be examined quantitatively and in which the biochemical actions of these compounds will be investigated. The analytic approach in most of the quantitative studies involves addition of a heavy isotope-labelled standard of the compound of interest to terminate islet incubations, liquid chromatographic isolation, conversion to a halogenated derivative, and analysis by highly sensitive and specific negative ion chemical ionization mass spectrometric methods. Biochemical events initiated by phospholipid hydrolysis which will be examined include: 1.) Secretagogue-induced accumulation of endogenous diacylglycerol, its fatty acid composition, and it possible biochemical effects; 2.) Accumulation of unesterified arachidonate and its temporal relationship to insulin secretion under conditions where one or both of these processes are perturbed; 3.) The time course of secretagogue-induced accumulation of arachidonate metabolites and their influence on insulin secretion and on biochemical events involved in secretion; 4.) Secretagogue induced changes in the phospholipids of intact islets and subcellular organelles including the appearance of novel phospholipids with potential mediator functions; and 5.) Accumulation by mass of endogenous inositol phosphates and their cyclic analogs and the possible involvement of glucose recognition factors in this process. These studies may help to clarify the mechanisms whereby glucose is recognized by islets and coupled to the secretion of insulin.
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