AB STRA CT This study was designed to determine the effect of nitroglycerin upon transmural distribution of myocardial blood flow in the awake dog during normal conditions and in the presence of ischemia-induced coronary vasodilation. Studies were performed in chronically prepared dogs with electromagnetic flowmeters and hydraulic occluders on the left circumflex coronary artery. Regional myocardial blood flow was estimated by using radionuclide-labeled microspheres, 7-10 im in diameter, injected into the left atrium. During control conditions endocardial flow (0.86±SEMI 0.05 ml/min per g) slightly exceeded epicardial flow (0.72±0.03 ml/min per g, P < 0.05), and this distribution of flow was not significantly altered by nitroglycerin. After a 5-s coronary artery occlusion, reactive hyperemia occurred with excess inflow of arterial blood effecting 360+15 15% repayment of the blood flow debt incurred during occlusion. When arterial inflow was limited to the preocclusion rate during coronary vasodilation after a 5-s total coronary artery occlusion, flow to the subepicardial myocardium was increased at the expense of underperfusion of the subendocardial myocardium, and the delayed reactive hyperemia was markedly augmented (mean blood flow debt repayment = 775±105%, P < 0.01). These data suggested that subendocardial underperfusion during the interval of coronary vasodilation in the presence of a flow-limiting