Islet Secretory Defect in Insulin Receptor Substrate 1 Null Mice Is Linked With Reduced Calcium Signaling and Expression of Sarco(endo)plasmic Reticulum Ca2+-ATPase (SERCA)-2b and -3

Islet Secretory Defect in Insulin Receptor Substrate 1 Null Mice Is Linked With Reduced Calcium Signaling and Expression of Sarco(endo)plasmic Reticulum Ca 2+ -ATPase (SERCA)-2b and -3 Rohit N. Kulkarni 1 , Michael G. Roper 2 , Gabriella Dahlgren 2 , David Q. Shih 3 , Lisa M. Kauri 2 , Jennifer L. Peters 2 , Markus Stoffel 3 and Robert T. Kennedy 2 1 Research Division, Joslin Diabetes Center, and Department of Medicine, Harvard Medical School, Boston, Massachusetts 2 Department of Chemistry, University of Florida, Gainesville, Florida 3 Laboratory of Metabolic Diseases, The Rockefeller University, New York, New York Address correspondence and reprint requests to Rohit N. Kulkarni, MD, PhD, Rm. 602, Joslin Diabetes Center, One Joslin Pl., Boston MA 02215. E-mail: rohit.kulkarni{at}joslin.harvard.edu Abstract Mice with deletion of insulin receptor substrate (IRS)-1 (IRS-1 knockout [KO] mice) show mild insulin resistance and defective glucose-stimulated insulin secretion and reduced insulin synthesis. To further define the role of IRS-1 in islet function, we examined the insulin secretory defect in the knockouts using freshly isolated islets and primary β-cells. IRS-1 KO β-cells exhibited a significantly shorter increase in intracellular free Ca 2+ concentration ([Ca 2+ ] i ) than controls when briefly stimulated with glucose or glyceraldehyde and when l -arginine was used to potentiate the stimulatory effect of glucose. These changes were paralleled by a lower number of exocytotic events in the KO β-cells in response to the same secretagogues, indicating reduced insulin secretion. Furthermore, the normal oscillations in intracellular Ca 2+ and O 2 consumption after glucose stimulation were dampened in freshly isolated KO islets. Semiquantitative RT-PCR showed a dramatically reduced islet expression of sarco(endo)plasmic reticulum Ca 2+ -ATPase (SERCA)-2b and -3 in the mutants. These data provide evidence that IRS-1 modulation of insulin secretion is associated with Ca 2+ signaling and expression of SERCA-2b and -3 genes in pancreatic islets and provides a direct link between insulin resistance and defective insulin secretion. 5-HT, 5-hydroxytryptamine [Ca2+]i, intracellular free Ca2+ concentration IRS, insulin receptor substrate KRB, Krebs-Ringer buffer PI, phosphatidylinositol SERCA, sarco(endo)plasmic reticulum Ca2+-ATPase Footnotes G.D, J.L.P., and R.T.K are currently affiliated with the Departments of Chemistry and Pharmacology, University of Michigan, A