Methods to Study Roles of β-Arrestins in the Regulation of Pancreatic β-Cell Function

Abstract

Novel findings reveal important functional roles for β-arrestin 1 and β-arrestin 2 in the regulation of insulin secretion, β-cell survival, and β-cell mass plasticity not only by glucose but also by G-protein-coupled receptors, such as the glucagon-like peptide-1 (GLP-1) and the pituitary adenylate cyclase-activating polypeptide (PACAP) receptors or GPR40, or tyrosine kinase receptors, such as the insulin receptor. Here, we describe experimental protocols to knock down β-arrestins by small interference RNA, to follow subcellular localization of β-arrestins in the cytosol and nucleus of the insulinoma INS-1E rat pancreatic β-cell line, and to analyze β-arrestin protein expression by Western blot using INS-1E cells and isolated mouse or human pancreatic islets. We also provide details on how to genotype β-arrestin 2 knockout (Arrb2−/−) mice and to evaluate β-arrestin-mediated roles in β-cell mass plasticity and β-cell signaling using immunocytochemistry on pancreatic sections or on primary dispersed β-cells from wild-type mice and Arrb2−/− mice.