Forty-eight rats were fed ad libitum, fasted 24 hr, rested 48 hr,and injected i.p. with 40 muCi of 14C-acetate/100 g body weight. Twenty-four rats had followed a progressive physical training program for 12 wk and 24 rats acted as their controls. Following this injection, the rats were sequentially sacrificed at 5-, 10-, 15-, and 20-min intervals and total cholesterol (TC), free cholesterol (FC), and triglyceride (TG) specific activity and concentrations were measured from serum, liver, triceps, and heart tissue. Curves relating specific activity to the time point data were fitted by the method of least squares. Comparison of these curves revealed that serum, liver, and triceps TC and FC specific activity were significantly higher in the trained rats. In contrast, corresponding TC and FC concentrations for these three tissues varied. Liver TC level was significantly less for the trained group, probably due to a reduction in the esterified moiety, since liver FC measures were unchanged. Training resulted in significantly lower TC concentrations in the selected tissues studied even though specific activity curves appeared similar for both groups. Our conclusions are that lipid metabolic adaptation; studied in vivo, occurs in tissues with training, but that these adaptations are not uniform across tissues, lipid moieties, or measurement parameters.