Forward [creatine phosphate (CP)----adenosine 5'-triphosphate (ATP)] and reverse (ATP----CP) fluxes of myocardial creatine kinase (CK) measured by using 31P nuclear magnetic resonance (NMR) and conventional saturation transfer (CST) methods are unequal; this is a paradoxical result because during steady state fluxes into and out of the CP pool must be the same. These measurements, however, treat the CK reaction as a two-site exchange problem and ignore the presence of the ATP gamma in equilibrium Pi exchange involving the ATPases. We have applied a method [Uğurbil, K. (1985) J. Magn. Reson. 64, 207] based on the saturation of multiple resonances, by which a single unidirectional rate constant can be measured unequivocally in the presence of multiple exchanges, to the measurement of CK fluxes in isovolumic rat hearts perfused under three different conditions; two of the three perfusion conditions showed a large discrepancy in the CK fluxes determined by CST, and one did not. In contrast, when the effect of the ATP gamma in equilibrium Pi exchange on the CK rate measurements was eliminated, multiple saturation transfer (MST) measurements on the same hearts yielded equal forward and reverse fluxes in all cases. The rate constant for the ATP gamma----CP conversion measured by MST was larger than the value obtained by the conventional methodology whereas both methods gave the same rate constant in the CP----ATP direction. These results demonstrate that the cause of the paradoxical data obtained by CST measurements of CK kinetics is the ATP gamma in equilibrium Pi exchange and that CK rates when determined rigorously are consistent with the CK reaction being in equilibrium.(ABSTRACT TRUNCATED AT 250 WORDS)

The mechanism by which mitochondrial respiration is coupled to ATP consumption in intact tissues is unclear. We determined the relationship between high-energy phosphate levels and oxygen consumption rate in rat hearts operating over a range of workloads and perfused with different substrates. With pyruvate +glucose perfusion, ADP levels were in general very low, and varied with MVO2 yielding an apparent Km of 25 +/- 5 microM, suggesting regulation of oxidative phosphorylation through availability of ADP. In contrast, with glucose perfusion in the presence or absence of insulin, ADP levels, ADP/ATP ratio or the phosphate potential were relatively constant over the workload range examined and generally not correlated with alterations in MVO2; it is suggested that under these conditions, carbon substrate delivery to the mitochondria may control mitochondrial respiration. The common feature of both of the suggested regulatory mechanisms is substrate limitation which, however, is exercised at different metabolic points depending on the carbon substrate available to the myocardium.

High resolution 'H NMR spectra of perfused rat hearts have been obtained under normoxic, ischemic and hypoxic conditions. Several myocardial metabolites including taurine, carnitine, lactate and tissue glycerides are detected in the 'H NMR spectra. Changes in oxygen availability induce perturbations in the levels of some metabolites, in particular, lactate. Experiments with fasted rats and with substrate-free perfusion suggest that the glycerides detected in 'H spectra are metabolically mobilizable but have a slow rate of turnover. These results demonstrate that utility of 'H NMR in monitoring myocardial metabolism. 'H-NMRPerfused rat heart Myocardial metabolism