The goal of this proposal is to identify whether increasing cardiac glycolysis at its rate-limiting step can mitigate diabetic cardiomyopathy (DC) and improve whole body glucose tolerance. DC is a major public health issue that arises in both type 1 and type 2 diabetes and is mediated by numerous factors. Chief amongst them is the loss of metabolic flexibility, which is the capacity of the heart to take up and metabolize available circulating nutrients. The healthy heart primarily uses fatty acids, but it can shift to glucose metabolism in response to feeding. However, with diabetes the heart relies almost exclusively on fatty acid oxidation and if chronic, this leads to mitochondrial dysfunction, oxidative stress, and ultimately DC. While restoring proper cardiac metabolism has therapeutic potential, there are currently no treatments to normalize metabolic inflexibility. We posit that increasing glycolysis can normalize metabolic inflexibility and mitigate DC. We have been testing this hypothesis using mice that have enhanced cardiac glucose metabolism (GlycoHi mice) via the expression of a constitutively active form of the glycolytic regulator, phosphofructokinase-2 (PFK-2). We found that: GlycoHi mice are resistant to diet-induced cardiac diastolic dysfunction; GlycoHi heart mitochondria have an enhanced capacity to use pyruvate, indicative of increased metabolic flexibility; and female GlycoHi mice have improved systemic glucose tolerance and are resistant to HFD effects. This supports our hypothesis that increasing cardiac PFK-2 activity can mitigate DC and have beneficial effects on whole body glucose regulation. Our first Aim is to test the hypothesis that increasing cardiac glycolysis improves metabolic flexibility in response to HFD or type 1 diabetes. Control and GlycoHi mice will be subjected to HFD or induced with type 1 diabetes. We will determine cardiac function and metabolic profile by both proteomics and metabolomics. Metabolic flexibility will be measured in adult cardiomyocytes using a radiolabeled assay. Aim 2 will test the hypothesis that increasing cardiac glycolysis sustains mitochondrial function under diabetic conditions. We will interrogate mitochondrial function in diabetic (T1D and T2D) control GlycoHi, and PFK-2 knockout mice. We will also determine how the increase in glycolysis is able to sustain pyruvate dehydrogenase activity. Aim 3 will determine the mechanisms by which increasing cardiac glycolysis improves whole body glucose tolerance in diabetic GlycoHi mice. We will discern between increased energy expenditure, using metabolic cages, and increased insulin sensitivity in heart, skeletal muscle, and adipose tissue. We will also test the hypothesis that the effects are mediated through changes in adipocyte differentiation and bioenergetics. The occurrence of diabetes continues to increase, and heart disease and heart failure are leading causes of death in this population. It is not known whether increasing cardi...