MOLECULAR MECHANISMS OF BETA CELL DYFUNCTION IN DIABETES

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The focus of this Program Project Application is on the molecular mechanisms associated with insulin secretion. Particular emphasis will be upon the identification of the molecular defects relevant to the understanding and treatment of abnormal beta cell function in patients with early IDDM. The Program consists of 4 projects and 2 cores and will carry out a multidisciplinary investigation of the insulin secretory process from a number of standpoints including the basic molecular biology of genes whose products play an important role in insulin secretion, to the study of the regulation of beta cell gene expression in normal and diabetic animals, to the electrophysiologic correlates of beta cell secretory dysfunction and an investigation of the role of the small GTP-binding proteins in the regulation of insulin secretion. Project 1 aims to clone and characterize genes expressed in the beta cell which are involved in the regulation of insulin secretion through effects on intracellular concentrations of Ca2+ and K+ including beta cell calcium channels, voltage and ATP-sensitive K+ channels and the IP3 receptors. Project 2 aims to study the molecular physiology of insulin secretion and its pathophysiology in diabetes by investigating factors responsible for the normal oscillatory pattern of insulin secretion. The regulation of pancreatic beta cell gene expression will be studied in normal and diabetic rates and in rats in which beta cell dysfunction has been induced by high dose glucose infusion. The pharmacologic control of insulin secretion will be investigated in a model beta cell line, the betaTC3 cell line which has been demonstrated to increase its insulin responsiveness to glucose, after incubation in low glucose when compared to high glucose. Project 3 aims to study the electrophysiologic correlates of the reduced beta cell function in diabetes by investigating beta cells from normal and diabetic or prediabetic animals with the whole cell and single channel patch clamp techniques and fura-2 fluorimetry with digital imaging to study patterns of Ca2+ and K+ signalling. Project 4 aims to define the role of small GTP-binding proteins (including Rab 1,2,6 and 3) in directing the intracellular traffic of proteins destined for packaging in beta granules. This will be accomplished by studying the intracellular localization of these proteins and the effects of their overproduction and altered function on proinsulin biosynthesis, processing and secretion from beta cell lines. The Animal and Cell Culture Core will provide investigators with whole animal models of diabetes and beta cell dysfunction, isolated islets and dispersed beta cells, model beta cell lines and heterologous systems for expression of cloned genes.