Mitochondrial-Cytosolic Interactions in Perfused Rat Heart ROLE OF COUPLED TRAKSAMINATION IN REPLETION OF CITRIC ACID CYCLE INTERMEDIATES*

The kinetics of changes in the tissue content of citric acid cycle intermediates and free amino acids have been investigated in isolated perfused hearts during the transition from substrate-free perfusion to perfusion with medium containing 5 mM glucose and 5 s 10m3 units of insulin per ml. Glycolytic flux increased within 1 min to about 300 pmoles of glucose equivalents per g (dry weight) per hour, and oxygen uptake increased by 18 to 28% from a basal rate of 3000 patoms per g (dry weight) per hour. The ratio of lactate to pyruvate in the perfusion fluid showed a transient rise from a value of 0.4 to a peak of 18 after 2 min and reached a final steady state value of 4. The tissue contents of oxalacetate and aspartate decreased by ‘76 and 59%, respectively, within the first 3 min, whereas the glutamate content increased reciprocally by 230%. The tissue contents of malate, citrate, and alanine increased more gradually over a 12-min period by 4.3-, 8.6-, and 4.4-fold, respectively. The a-ketoglutarate content initially increased rapidly and then fell before rising again to final values about Z-fold greater than the initial. The changes in the tissue contents of amino acids other than glutamate, aspartate, and alanine were minor. Net accumulation of citric acid cycle intermediates in the heart following addition of glucose can be accounted for by double transamination between alanine and aspartate aminotransferases. Creatine-P levels increased by 14 pmoles per g (dry weight), whereas tissue inorganic phosphate levels decreased by 12. pmoles per g (dry weight). These changes together with the observed increase of the ATP :ADP ratio from 10.8 to 14.2 indicate a higher energy state of the adenine nucleotide system after glucose addition. From the measured rates of glucose metabolism, oxygen uptake, and changes of the aspartate and alanine contents, together with the previously verified assumption that glycolytically generated NADH is transported to the mitochondria via the malate-aspartate cycle, calculations were made of flux through steps of the citric acid cycle and the anion exchange reactions across the mitochondrial membrane.