Abnormalities of energy metabolism are suspected to contribute to progression of myocardial dysfunction in the failing heart. We have observed that heart failure is associated with decreased myocardial ATP content and increased free ADP, as well as a reduction of myocardial oxygen consumption (MVO2). Recent studies have demonstrated that nitric oxide (NO) can compete with oxygen at cytochrome C oxidase to limit mitochondrial respiration. Although endothelial NO synthase (eNOS) activity is decreased in the failing heart, inducible NO synthase (iNOS) expression has been reported in the failing heart. In preliminary studies we found that selective iNOS inhibition resulted in an increase of MVO2 in failing hearts but not in normal hearts. Consequently, studies are proposed to test the hypothesis that NO acts to limit MVO2 and ATP production in pacing-induced heart Failure in dogs. Use of both the selective iNOS inhibitor 1400W, the selective nNOS inhibitor vinyI-L-NIO, as well as nonselective NOS inhibition with nitro-L-arginine, will demonstrate whether iNOS or nNOS is the principle source of NO in the failing heart. Because NO reacts with superoxide (02-) to form peroxynitrite, which might also impair mitochondrial respiration, studies will determine whether scavenging 02- or supplying a peroxynitrite decomposition catalyst can increase MVO2 in the failing heart. Since inhibition of mitochondrial respiration would impair ATP synthesis, 31P nuclear magnetic resonance (NMR) spectroscopy will be used to assess the effect of selective iNOS and nonselective NOS blockade, as well as degrading peroxynitrite, on MVO2 and myocardial [ADP]. If inhibition of mitochondrial respiration by NO, O2- or peroxynitrite impairs ATP production, resulting in an increase of free cytosolic ADP, then blocking NO synthesis, scavenging 02- or decreasing peroxynitrite production with an 02- scavenger and/or iNOS inhibitor would cause a decrease of [ADP] with a simultaneous increase of MVO2. Consequently, 31P NMR spectroscopy will be used to test the hypothesis that NOS inhibition results in a decrease of cytosolic [ADP] at the same time that MVO2 is increased. The use of selective iNOS, nNOS and nonselective NOS inhibitors in vivo and in vitro (skinned fibers and isolated mitochondria) will demonstrate whether this NO effect is mediated by iNOS, nNOS or mitochondrial NOS. Additional studies will determine whether mRNA and protein expression and activity for eNOS, nNOS, iNOS, CuZn-SOD, Mn-SOD and Ec-SOD are altered in coronary vessels and myocardium by the presence of CHF.
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