Changes in creatine kinase (CK) activity and CK isoenzyme profiles in plasma after exercise were studied in rats in order to establish the source of the exercise-induced rise in CK activity. Male and female rats ran on a treadmill for 2 h and blood samples, taken before and after exercise, were assayed for total CK, CK isoenzymes and aminoaspartate transaminase (AST) activity. These enzymes were also assayed in homogenates of liver and several muscles. We found that the isoenzyme composition of liver, plasma and muscle did not differ between the sexes. However, the exercise-induced CK and AST responses did differ: CK and AST increased after exercise in males (101% and 15% resp.), but much less in females (47% and 1%). Although the isoenzyme profiles in rest did not differ, significant differences were observed after running: in males CK-MM increased with 678%, but females only showed a 114% increase. In contrast, CK-BB showed a small increase that was about the same for both sexes (males 41%, females 35%). We conclude that both males and females show a small and similar increase in CK-BB activity after exercise, and that a large release of CK-MM from skeletal muscle, observed only in males, accounts for sex-linked differences reported earlier.