The effects of 4 weeks of voluntary exercise (rotating wheel) on serum and liver lipid levels were studied in 6-week-old male Wistar rats. The hypotriglyceridemic effect of exercise was not influenced by the dietary fat levels (4.5 and 14.5%), whereas the hypocholesterolemic effect was higher in a low-fat diet than in a high-fat diet. Although the serum cholesterol-lowering effect of exercise was marginal, the beneficial effect on hepatic lipids was observed only in rats fed a high-cholesterol diet. The hepatic contribution to the serum lipid-lowering action of exercise was examined in the isolated perfused livers of rats fed a low fat diet. The reduction (34%) of serum triglyceride by exercise corresponded to the decrease (32%) in the hepatic triglyceride secretion rate, indicating a significant role of the liver in the hypotriglyceridemic effect of exercise. In contrast, the decrease (31 %) in serum total cholesterol was considerably greater than that (23%) in the hepatic secretion, suggesting an extra-hepatic contribution. On the other hand, the hepatic production of ketone body was increased by exercise. These results indicate that the altered hepatic partition of long-chain free fatty acids between esterification and oxidation is one of the causative factors for the serum lipid-lowering effect of exercise.