Lipid signal transduction /oscillatory insulin secretion

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DESCRIPTION (provided by applicant): Insulin secretion oscillates, possibly due to oscillatory metabolism of glycolysis, mitochondrial energy production, anaplerosis and lipolysis. These oscillations generate important signals underlying oscillations in electrical activity, ion fluxes and, ultimately, secretion. We hypothesize that FFA may participate as coupling factors to synchronize islet metabolic signal transduction and insulin secretion and to provide communication between peripheral fat cells and islets. This may be achieved by recruiting beta cells, increasing synchrony among beta cells or enhancing responsiveness of beta-cells to glucose. We further presume that FFA and their release by hormone sensitive lipase (HSL) generate important signals that regulate insulin secretion in response to fuel stimuli and elevation of cAMP. We predict that animal models deficient in PFK-M (phosphofructokinase-M) or HSL will demonstrate the critical importance of glycolytic and lipolytic pathways in insulin secretion. Aim 1 will determine the effect of FFA on glucose-induced mitochondrial metabolic oscillations of individual cells within the intact islet. The role of endogenous FFA as a signaling molecule will be studied by monitoring the synchronized metabolic activity in the intact HSL deficient islet. Aim 2 will determine the relationship between oscillatory changes in lipid partitioning and lipolysis in beta cells and islets and GSIS and the effect of FFA on these parameters. Intracellular distribution and movement in real time of fluorescent FFA will be monitored using confocal imaging of beta cells and islets. In addition FFA and glycerol release from stimulated cells will be temporally correlated with insulin secretion and measurements made in Aim 1. Aim 3 will test the hypothesis that FFA promote secretion via direct modulation of voltage-gated L-type calcium channels. Aim 4 will test the hypothesis that oscillatory FFA release from fat cells via HSL-mediated lipolysis plays a role in synchronized insulin secretion from islets. Sequential co-perifusion of isolated fat cells and pancreatic islets will be undertaken. Aim 5 will test the hypothesis that regulation of HSL in the R-cell occurs via oscillatory inhibition by long chain-CoA. Aim 6 will study the mechanism whereby HSL-deficiency in beta-cell results in altered GSIS. Aim 7 will test mechanisms of secretory and electrical synchronization in the intact perfused pancreas and its relationship to islet and single cell studies.