While autophagy is induced by myocardial ischemia/reperfusion, it is unclear whether autophagy is detrimental or beneficial to myocardial survival during ischemia/reperfusion. Isolated rat hearts were subjected to 30 min regional ischemia followed by 2 h of reperfusion. Autophagy was determined by the ratio of LC3 -II to LC3-I with Western blotting. Autophagy was prominent upon reperfusion but not during ischemia in rat hearts, indicating that autophagy may play a role during reperfusion phase. Ischemia or reperfusion did not enhance Beclin 1 expression, suggesting that Beclin 1 may not be critical for the formation of autophagy in isolated rat hearts. 3-methyladenine (3-MA), a classical inhibitor of autophagy, suppressed reperfusion-induced autophagy and reduced the infarct size when introduced at reperfusion. NECA, an agonist of adenosine receptors, and morphine also reduced the formation of autophagy as well as the infarct size when introduced at reperfusion. These data suggest that autophagy may play a detrimental role during reperfusion and that modulation of autophagy may prevent.