Project Summary Islet glucose-stimulated somatostatin (Sst) secretion is lost in patients with type-2 diabetes (T2D) and in animal models of the disease, which contributes to disrupted glucagon and insulin secretion. It is generally accepted that Sst secretion from -cells occurs in response to elevated intracellular Ca2+, which primarily results from endoplasmic reticulum (ER) Ca2+ (Ca2+ER) release. However, the mechanisms that control -cell Ca2+ER handling and how they are altered in T2D are largely unknown. Data from our lab finds that the islet-enriched two-pore- domain K+ channel, TALK-1, is an ER localized channel in that provides a countercurrent for -cell Ca2+ER release and Ca2+ER leak. TALK-1-mediated augmentation of the electrochemical driving force for -cell Ca2+ER leak con- strains Ca2+ER storage, which limits glucose-stimulated Ca2+ER release and Sst secretion. Further data show that -cell Ca2+ER release and Sst secretion are amplified by glucose-induced allosteric activation of -cell Ca2+-sens- ing receptors (CaSRs). Finally, our preliminary data provide the first evidence that diabetic conditions diminish -cell Ca2+ER storage, which contributes to perturbations in glucose-stimulated Ca2+ handling and Sst secretion under diabetic conditions. Based on these exciting preliminary data, the overall objective of this proposal is to elucidate how -cell Ca2+ER is controlled and becomes disrupted during the pathogenesis of diabetes. This project will test the central hypothesis that glucose-stimulated -cell Sst secretion is amplified by CaSR-mediated Ca2+ER release, which is controlled by TALK-1 channel constraint of Ca2+ER storage. The rationale that underlies this project is that understanding how CaSR and TALK-1 control -cell Ca2+ER handling and Sst secretion will expose novel therapeutic targets for restoring glucose-stimulated Sst secretion and islet hormone secretion in T2D. This project will be accomplished with the following two specific aims: 1) Determine how -cell CaSR controls Ca2+ER handling, Sst secretion, and islet hormone secretion; and 2) Determine how TALK-1 channel control of Ca2+ER release modulates -cell function and dysfunction. Under the first aim, transgenic mice with -cell ablation of CaSR as well as human pseudoislets with ShRNA knockdown of -cell CaSR will be utilized to assess the roles of the Ca2+-sensing receptor during secretagogue modulation of -cell Ca2+ handling and Sst secretion. Aim1 will also determine how depletion of -cell Ca2+ER stores under diabetic conditions impacts CaSR signaling and Sst secretion. Under the second aim, the function TALK-1 channels on -cell Ca2+ER handling and function will be determined in mice with -cell specific ablation of TALK-1 and in human pseudoislets containing either -cells with knockdown of TALK-1 or expressing dominant negative TALK-1 channel subunits. Furthermore, Aim2 will determine how TALK-1 augmentation of -cell Ca2+ER depletion under the stressf...