Selective NADH communication from α-ketoglutarate dehydrogenase to mitochondrial transhydrogenase prevents reactive oxygen species formation under reducing conditions in the heart

In heart failure, a functional block of complex I of the respiratory chain provokes superoxide generation, which is transformed to H(2)O(2) by dismutation. The Krebs cycle produces NADH, which delivers electrons to complex I, and NADPH for H(2)O(2) elimination via isocitrate dehydrogenase and nicotinamide nucleotide transhydrogenase (NNT). At high NADH levels, α-ketoglutarate dehydrogenase (α-KGDH) is a major source of superoxide in skeletal muscle mitochondria with low NNT activity. Here, we analyzed how α-KGDH and NNT control H(2)O(2) emission in cardiac mitochondria. In cardiac mitochondria from NNT-competent BL/6N mice, H(2)O(2) emission is equally low with pyruvate/malate (P/M) or α-ketoglutarate (α-KG) as substrates. Complex I inhibition with rotenone increases H(2)O(2) emission from P/M, but not α-KG respiring mitochondria, which is potentiated by depleting H(2)O(2)-eliminating capacity. Conversely, in NNT-deficient BL/6J mitochondria, H(2)O(2) emission is higher with α-KG than with P/M as substrate, and further potentiated by complex I blockade. Prior depletion of H(2)O(2)-eliminating capacity increases H(2)O(2) emission from P/M, but not α-KG respiring mitochondria. In cardiac myocytes, downregulation of α-KGDH activity impaired dynamic mitochondrial redox adaptation during workload transitions, without increasing H(2)O(2) emission. In conclusion, NADH from α-KGDH selectively shuttles to NNT for NADPH formation rather than to complex I of the respiratory chain for ATP production. Therefore, α-KGDH plays a key role for H(2)O(2) elimination, but is not a relevant source of superoxide in heart. In heart failure, α-KGDH/NNT-dependent NADPH formation ameliorates oxidative stress imposed by complex I blockade. Downregulation of α-KGDH may, therefore, predispose to oxidative stress in heart failure. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (10.1007/s00395-020-0815-1) contains supplementary material, which is available to authorized users.