Hearts from normal male rats were perfused with Krebs-Ringer bicarbonate buffer by the Langendorff method for periods ranging from 0–4 hr following an initial 15-min equilibration period. Spontaneous heart rate began to decrease by 30 min, whereas changes in coronary flow were not evident until after approximately 2 hr of perfusion. A reduction in isometric systolic tension was evident by 1 hr; after 4 hr of perfusion, tension was 35% of control. Cardiac glycogen concentrations became significantly different from control after 3 hr of perfusion, and at the end of 4 hr had decreased to 33% of control. Adenosine-5′-triphosphate content of hearts was not significantly different from control until perfused for 4 hr. In contrast, creatine phosphate concentrations decreased approximately 50% during the first hour of perfusion but remained constant thereafter. Prolonged perfusion (4 hr) caused no alteration in cardiac phosphorylase activity nor was there evidence of gross or microscopic lesions in the heart.