Defective insulin secretion in response to glucose is an important component of the β cell dysfunction seen in type 2 diabetes. As mitochondrial oxidative phosphorylation plays a key role in glucose-stimulated insulin secretion (GSIS), oxygen-sensing pathways may modulate insulin release. The von Hippel-Lindau (VHL) protein controls the degradation of hypoxia-inducible factor (HIF) to coordinate cellular and organismal responses to altered oxygenation. To determine the role of this pathway in controlling glucose-stimulated insulin release from pancreatic β cells, we generated mice lacking Vhl in pancreatic β cells (βVhlKO mice) and mice lacking Vhl in the pancreas (PVhlKO mice). Both mouse strains developed glucose intolerance with impaired insulin secretion. Furthermore, deletion of Vhl in β cells or the pancreas altered expression of genes involved in β cell function, including those involved in glucose transport and glycolysis, and isolated βVhlKO and PVhlKO islets displayed impaired glucose uptake and defective glucose metabolism. The abnormal glucose homeostasis was dependent on upregulation of Hif-1α expression, and deletion of Hif1a in Vhl-deficient β cells restored GSIS. Consistent with this, expression of activated Hif-1α in a mouse β cell line impaired GSIS. These data suggest that VHL/HIF oxygen-sensing mechanisms play a critical role in glucose homeostasis and that activation of this pathway in response to decreased islet oxygenation may contribute to β cell dysfunction. IntroductionBlood glucose levels are normally tightly controlled by the regulation of insulin release from the pancreatic β cells. Glucose-stimulated insulin secretion (GSIS) is a complex metabolic process involving the uptake and phosphorylation of glucose via GLUT2 transporters and glucokinase (Gck), respectively, metabolism of glucose-6-phosphate via the glycolytic pathway, and subsequent activation of mitochondrial metabolism to produce coupling factors such as ATP (1). A rise in the cytoplasmic ATP/ADP ratio leads to closure of K ATP channels, depolarization of the plasma membrane, opening of voltage-sensitive Ca 2+ channels, and activation of Ca 2+ -dependent exocytotic mechanisms, resulting in insulin secretion (1). This metabolic sensing mechanism requires molecular oxygen for the quantitative generation of ATP from glucose. Understanding the complex physi-

Aims/hypothesis Activation of the G protein-coupled receptor (GPR)40 by long-chain fatty acids potentiates glucosestimulated insulin secretion (GSIS) from pancreatic beta cells, and GPR40 agonists are in clinical development for type 2 diabetes therapy. GPR40 couples to the G protein subunit Gα q/11 but the signalling cascade activated downstream is unknown. This study aimed to determine the mechanisms of GPR40-dependent potentiation of GSIS by fatty acids. Methods Insulin secretion in response to glucose, oleate or diacylglycerol (DAG) was assessed in dynamic perifusions and static incubations in islets from wild-type (WT) and Gpr40 −/− mice. Depolymerisation of filamentous actin (F-actin) was visualised by phalloidin staining and epifluorescence. Pharmacological and molecular approaches were used to ascertain the roles of protein kinase D (PKD) and protein kinase C delta in GPR40-mediated potentiation of GSIS.Results Oleate potentiates the second phase of GSIS, and this effect is largely dependent upon GPR40. Accordingly, oleate induces rapid F-actin remodelling in WT but not in Gpr40islets. Exogenous DAG potentiates GSIS in both WT and Gpr40 −/− islets. Oleate induces PKD phosphorylation atElectronic supplementary material The online version of this article