Metabolic enzyme activity mediating myocardial stunning

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DESCRIPTION (provided by applicant): Stimulating carbohydrate oxidation via pyruvate dehydrogenase (PDH) activation is known to irmprove contractile recovery of postischemic myocardium. We have determined that the benefits of activating PDH rely neither on glycolytic nor mitochondrial energy production. Instead, PDH-dependent changes in cytosolic redox state influence the recovery of the postischemic heart. Our results on PDH activation during the first hour of reperfusion in the in vivo heart of conscious pigs demonstrate reversal of early reperfusion injury that results in sustained improvement in contractility. Exciting new data show that stimulating pyruvate oxidation in reoxygenated cardiomyocytes improves contractile response to calcium, eliminates calcium overload, and improves contractile relaxation rate. Therefore, this study explores the hypothesis that PDH activation eliminates early reperfusion injury via favorable shifts in cytosolic redox balance that improve calcium homeostasis and mitochondrial function due to elimination of calcium overload and potentially, the integrity of the myofilaments. This hypothesis will be tested in both the single adult rat cardiomyocyte and in the isolated, perfused rat heart using a novel combination of techniques that include optical microscopy and 13C NMR spectroscopy. Specific aims are: 1) Determine the effects of cytosolic redox state on intracellular calcium levels and contractile response to calcium in reoxygenated cardiomyocytes; 2) a) Evaluate the response of SERCA2a activity to augmented carbohydrate oxidation as a mechanism of improved calcium handling in reperfused hearts; b) if SERCA2a activity is improved by PDH activation in reperfused hearts then determine whether SERCA2a over expression in vivo rat hearts produces similar benefits to contractile recovery; 3) Determine the relationship between calcium and induced shifts in cytosolic redox state in affecting contractile function during augmented carbohydrate oxidation in reperfused myocardium; 4) Elucidate the relationship/competition between the calcium activated mitochondrial dehydrogenase, alpha-ketoglutarate dehydrogenase, versus activity of cytosolic reducing equivalent transport (malate-aspartate shuttle) in response to calcium load in the reperfused myocardium; 5) Examine the potential for augmented carbohydrate oxidation to reverse stunning via reduced myofilament protein degradation and improved calcium sensitivity.