The effect of independently varying heart rate, left ventricular systolic pressure, coronary blood flow or coronary artery perfusion pressure on the transmural distribution of myocardial blood flow was determined in the isolated supported dog heart preparation by use of radioactive microspheres. The distribution of radioactivity was expressed as the epicardial/endocardial blood flow ratio (epi/endo). At a constant coronary blood flow, increases in heart rate (111-250 beats/min) or decreases in coronary artery perfusion pressure (100-46 mm Hg) resulted in an elevation of epi/endo (0.64 to 1.12 and 0.71 to 1.41, respectively). Increasing coronary artery perfusion pressure (40-130 mm Hg) and coronary blood flow from low values resulted in a decreased epi/endo (1.28 to 0.76). The absolute level of coronary blood flow had no direct effect on epi/endo but acted through changes in coronary artery perfusion pressure. The major physiological determinants of intramyocardial blood flow distribution in the isolated dog heart are coronary artery perfusion pressure and heart rate, whereas left ventricular systolic pressure and coronary blood flow are of minor importance. An equation was developed relating the mean epi/endo of the left ventricle to hemodynamic factors studied, and a highly significant correlation was found between predicted and experimental ratios. These results indicate the potential usefulness of this model for predicting epicardial-endocardial blood flow ratios based upon drug-induced changes in hemodynamic parameters.