The purpose of this study was to determine whether exercise training would prevent the progressive functional decline in pump function of hearts from diabetic rats. Four groups were studied: sedentary control, trained control, sedentary diabetic, and trained diabetic. Trained rats were adapted to the treadmill prior to induction of diabetes in half of the group streptozotocin injected (50 mg/kg). Thereafter the duration, speed, and grade were then progressively increased until the trained rats could run for 60 min at 27 m/min, 5% grade (wk 8). Cardiac output and work were measured in isolated working hearts perfused at various left atrial filling pressures and with buffer containing the concentrations of glucose and fatty acids found in vivo. Sedentary diabetic rats had lowered body weight, elevated plasma glucose, triacylglycerol, and cholesterol. Exercise training of diabetic rats lowered plasma triacylglycerol levels. Training increased plantaris muscle cytochrome oxidase activity significantly in both the trained control and trained diabetic groups. Cardiac pump function was impaired in hearts from the sedentary diabetic rats perfused with either normal or diabetic substrate conditions, but the impairment was larger under diabetic conditions. Training of diabetic rats prevented this depression. Myocardial carnitine content was decreased in hearts from sedentary diabetic rats. Exercise training increased carnitine content in both control and diabetic rats. This training protocol did not affect cardiac pump function of the trained control group. These results suggest that exercise training may limit the myocardial contractile dysfunction associated with diabetes mellitus.