Creatine kinase (CK) release in response to excessive electrically stimulated contractile activity has been studied in isolated rat soleus muscles. The exacerbation of CK release induced by contractile activity was found to be directly related to the length of time for which the muscle was stimulated and indirectly related to the recovery of force following the end of stimulation. 31P-NMR studies were undertaken using a recirculating superfused muscle preparation and demonstrated that muscles subjected to two different stimulation protocols (stimulation for 0.5 s every 2 s in oxygenated medium or for 1.5 s every 2 s in anoxic medium) had similar falls in ATP content and pH despite a substantially greater release of CK from the muscles stimulated under anoxia. However, stimulated muscles under anoxia showed a more rapid fall and reduced recovery of phosphocreatine and a greater sustained elevation of inorganic phosphate than muscles in oxygenated medium. It is concluded that only part of the increased loss of CK from muscles stimulated in anoxic medium can be explained by release from cells which have lost energy supplies and therefore that other mechanisms must exist which allow release of CK and other cytosolic enzymes from muscle cells.