When the oxygen supply to the myocardium is compromised, glycolysis may provide the critical energy necessary for the heart's survival. We investigated whether insulin, through its effects on myocardial glycolysis, influences the performance and the metabolism of the perfused rat heart during anoxia. Hearts in a modified Langendorff apparatus were perfused for 30 minutes with anoxic media containing glucose (50–500 mg/100 ml) with and without insulin (100 munits/ml) or with anoxic media containing 200 mg glucose/100 ml and varying insulin concentrations (0.1–100 munits/ml). Hearts were paced, and left ventricular pressure, maximum rate of rise of left ventricular pressure, and lactate production were monitored. Increasing glucose concentration alone progressively enhanced performance and lactate generation in the beating anoxic heart. Addition of insulin resulted in significant increases in left ventricular performance and lactate production at all levels of glucose concentration. The myocardial content of high energy intermediates (creatine phosphate, adenosine triphosphate, adenosine diphosphate, and adenosine monophosphate) after 30 minutes of anoxia was not altered by varying concentrations of glucose or insulin. To assess the effects of insulin and glucose in the absence of contraction, hearts were arrested with media containing a high potassium concentration (26 mEq/liter). Although lactate production was lower in arrested hearts than it was in beating hearts, it was enhanced by insulin. Insulin also produced significant increases in high-energy intermediates in arrested hearts. It was concluded that insulin increases the utilization of glucose and thus causes the enhancement of ventricular performance in the anoxic heart.