Effects of perfusion pressure in a range from 50 to 140 cmH2O on intracellular Ca2+ concentration ([Ca2+]i) were evaluated along with cardiac function, energy metabolism, and left ventricular geometry in a concentration of 2 or 4 mM of extracellular Ca2+ ([Ca2+]o) in isovolumic perfused rat hearts. [Ca2+]i was evaluated with a surface fluorometry technique in hearts loaded with indo-1/AM. The systolic and diastolic values and the amplitude (difference between systolic and diastolic values) of indo-1 fluorescence ratio (an index of [Ca2+]i) were linearly related to perfusion pressure. Changes in the fluorescence ratio were harmonious with rapid changes in left ventricular pressure and stabilized within 30-40 s after changes in perfusion pressure. Developed pressure and O2 consumption were closely, linearly correlated with the fluorescence ratio irrespective of [Ca2+]o. Left ventricular end-diastolic wall thickness, measured by 2-dimensional echocardiography, paralleled perfusion pressure and showed a good correlation with the fluorescence ratio. Diastolic myocardial adenosine 3',5'-cyclic monophosphate significantly decreased only at the lowest perfusion pressure. The ln[phosphocreatine]/[Pi] also changed with altered perfusion pressure. In conclusion, perfusion pressure modulates [Ca2+]i, which in turn regulates myocardial contraction and associated O2 utilization.