This study was designed to measure the response of key enzymes of ketone body metabolism in heart, skeletal muscle, and liver to diet and exercise, two conditions known to influence ketone body utilization. A 3 (diet: control, high fat, or high carbohydrate) X 2 (kill condition: rested or exhausted) X 2 (training: trained or untrained) factorial design was used to estimate main experimental effects as well as identify significant interactions of the variables. Physical training (treadmill running) was associated with a doubling of the activity of skeletal muscle 3-oxoacid CoA transferase, a key enzyme in extrahepatic ketone body utilization. The activity of the rate-limiting enzyme of liver ketone body production, hydroxymethylglutaryl CoA synthetase (HMG CoA synthetase), was not greatly influenced by training or exhuastive exercise indicating that the metabolic control of the ketosis of exercise may more likely be a function of the supply of fatty acids to the liver rather than the activity of HMG CoA synthetase. Feeding a high fat diet, on the other hand, significantly increased the activity of liver HMG CoA synthetase, indicating that the ketosis of fat feeding may be of a different nature than that of exercise. The results of this study indicate that physical training is associated with biochemical adaptations in ketone body metabolism as well as fatty acid oxidation, and that trained individuals are metabolically better endowed to benefit from the ketosis of exercise than untrained individuals.