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

1 Nuclear Magnetic Resonance Center and 2 Positron-Emission Tomography Laboratory, Department of Radiology, Massachusetts General Hospital, Harvard Medical School, Boston, Massachusetts 02129; and 3 Department of Molecular and Cellular Physiology, Pennsylvania State University Medical School, Hershey, Pennsylvania 17033 To test how -ketoglutarate dehydrogenase ( -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 -KGDH for -ketoglutarate ( -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 -KG efflux decreased from 126 to 95 nmol · min 1 · mg 1 . In intact hearts, K m was reduced with low intracellular pH, while matching control workload and respiratory rate with increased Ca 2+ (pH i = 7.20, perfusate CaCl 2 = 1.5 mM; pH i = 6.89, perfusate CaCl 2 = 3 ± 1 mM). Sequential 13 C nuclear magnetic resonance spectra from hearts oxidizing [2- 13 C]acetate provided tricarboxylic acid cycle flux and the exchange rate between -KG and cytosolic glutamate ( F 1 ). Tricarboxylic acid cycle flux was 10 µmol · min 1 · g 1 in both groups, but F 1 fell from a control of 9.3 ± 0.6 to 2.8 ± 0.4 µmol · min 1 · g 1 at low K m . The results indicate that increased activity of -KGDH occurs at the expense of -KG efflux during support of normal workloads. metabolic regulation; tricarboxylic acid cycle; myocardium; nuclear magnetic resonance