To study the relation among mitochondrial energy supply, cardiac performance, and energy transfer through creatine kinase (CK), two acute models of inhibition of ATP synthesis were compared in the isovolumic acetate-perfused rat heart. Similar impairments of mechanical performance (rate-pressure product, RPP) were achieved by various stepwise decreases in O(2) supply (PO(2) down to 20% of control) or by infusing CN (0.15-0.25 mM). The forward CK flux measured by saturation-transfer (31)P NMR spectroscopy was 6.1 +/- 0. 4 mM/s in control hearts. Only after severe hypoxia (PO(2) < 40% of control) did CK flux drop (to 1.9 +/- 0.2 mM/s at PO(2) = 25% of control) together with impaired systolic activity and a rise in end-diastolic pressure. In contrast, in mild hypoxia CK flux remained constant and similar to control (5.3 +/- 0.5 mM/s, not significant) despite a twofold reduction in systolic activity. Similarly in all CN groups, constant CK flux was maintained for a threefold reduction in RPP, showing the absence of a relation between cardiac performance and global NMR-measured CK flux during mild ATP synthesis inhibition.