Clinical and experimental investigations have indicated that diabetes per se can lead to a deterioration of cardiac function that cannot be solely attributable to coronary artery disease. What is apparent from most models of heart failure is that ventricular myocytes undergo similar phenotypic and genotypic changes (defined as a cardiomyopathy) in response to various metabolic and systemic stresses. This investigation is designed to test the hypothesis that cellular responses to type I and type 2 diabetes converge early on in the diseases, and manifest in abnormal cardiomyocyte intracellular Ca2+ regulation. The applicants will test this hypothesis by determining intracellular signaling pathways that contribute to the development of abnormal excitation- contraction (E-C) coupling in two distinct models of early stages of diabetes: 1) culturing myocytes in a hyperglycemic milieu and 2) sucrose-feeding animals triggering the development of insulin resistance. Myocyte E-C coupling will be evaluated using biophysical measures (video-edge detection, electrophysiology and spectrofluorometry), and kinase activity and expression of specific proteins using biochemical and immunoblot assays. Pharmacological interventions (both in cell culture and in vivo) will be used to evaluate the roles of target kinases (i.e., PKC, PKA and CaMKII) in the pathogenesis of impaired E-C coupling. The working hypothesis is that changes in regulatory protein activity precede overt changes in gene expression of ion channels and membrane pumps, such that these changes are the initial cause of myocyte dysfunction rather than a compensatory phenotypic change induced to preserve contractile function. This investigation will focus on cellular mechanisms which contribute to abnormal E-C coupling in diabetes and are common to those in other models of heart failure (e.g., depressed K+ conductance and SR Ca2+ fluxes). The development of cellular insulin resistance is likely to be a key factor in the etiology of diabetic cardiomyopathy and may represent the recapitulation of fetal gene expression, as shown for hypertrophic myocardium.
