The patterns of use of substrate in reperfused myocardium are not yet well elucidated, and their delineation is essential for adequate interpretation of results of analyses performed after positron emission tomography with labeled substrates to differentiate normal from abnormal heart muscle. Accordingly, in open-chest, anesthetized dogs we measured glucose and fatty acid utilization in normal, ischemic, and reperfused myocardium and assessed the contributions of metabolism of each substrate to overall oxidative metabolism. Intracoronary [3H]glucose and [14C]palmitate were administered in five control dogs, eight dogs subjected to 1 h of coronary occlusion, and nine dogs subjected to reperfusion after 1 h of ischemia. Regional coronary venous blood samples were assayed sequentially. In reperfused myocardium, utilization of glucose (463 +/- 88 nmol X g-1 X min-1) was 103% greater than that in ischemic and 273% greater than in normal myocardium (P less than 0.05 for each). Utilization of fatty acid during reperfusion (55 +/- 10 nmol X g-1 X min-1), although greater than that in ischemic myocardium, was significantly less than that in normal myocardium (48% of control, P less than 0.05) despite restoration of flow to 80% of control values. Although glucose constituted 70% of the substrate oxidized in ischemic myocardium, its contribution to overall oxidative metabolism in reperfused myocardium was only 25%. In contrast, despite diminished net uptake of fatty acid, oxidation of fatty acid accounted for 63% of total oxygen consumption in reperfused myocardium. These studies indicate that canine myocardium reperfused after 1 h of ischemia exhibits enhanced uptake of glucose and impaired utilization of palmitate.(ABSTRACT TRUNCATED AT 250 WORDS)