Dehydrogenase regulation of metabolite oxidation and efflux from mitochondria in intact hearts

To test how alpha-ketoglutarate dehydrogenase (alpha-KGDH) activity influences the balance between oxidative flux and transmitochondrial metabolite exchange, we monitored these rates in isolated mitochondria and in perfused rabbit hearts at an altered kinetics (K-m) of alpha-KGDH for alpha-ketoglutarate (alpha-KG). In isolated mitochondria, relative K-m dropped from 0.23 mM at pH = 7.2 to 0.10 mM at pH 6.8 (P < 0.05), and alpha-KG efflux decreased from 126 to 95 nmol min-l mg-l. In intact hearts, K-m was reduced with low intracellular pH, while matching control workload and respiratory rate with increased Ca2+ (pH(i) = 7.20, perfusate CaCl2 = 1.5 mM; pH(i) = 6.89, perfusate CaCl2 = 3 +/- 1 mM). Sequential C-13 nuclear magnetic resonance spectra from hearts oxidizing [2-C-13]acetate provided tricarboxylic acid cycle flux and the exchange rate between alpha-KG and cytosolic glutamate (F-1). Tricarboxylic acid cycle flux was 10 mu mol.min(-1).g(-1) in both groups, but F-1 fell from a control of 9.3 +/- 0.6 to 2.8 +/- 0.4 mu mol.min(-1).g(-1) at low K-m. The results indicate that increased activity of alpha-KGDH occurs at the expense of alpha-KG efflux during support of normal workloads.