All cellular metabolism, i.e. anabolism, catabolism and transport, is intimately coupled to the cellular energy level as expressed in the [ATP]/[ADP][Pi]. Mitochondrial oxidative phosphorylation is primarily responsible for maintaining the energy level and it thereby strongly influences all other metabolism. Our objective is to gain an understanding of the regulation of cellular metabolism and in this project we will concentrate on the role of oxygen and intracellular pH in metabolic homeostasis. In order to approach this general question we will: 1. Examine the oxygen dependence of mitochondrial oxidative phosphorylation in vivo as it is expressed in the cytoplasmic [ATP]/[ADP][Pi] and [NAD]/[NADH], the mitochondrial [NAD]/[NADH] and in the behavior of the energy requiring metabolic reactions such as gluconeogenesis, urea synthesis and protein synthesis. 2. Examine the proposal that mitochondrial oxidative phosphorylation is the primary oxygen sensor for regulation of tissue oxygen tension through control of local vascular resistance. 3. Measure the oxygen dependence of prostaglandin synthesis by cultured endothelial cells. 4. Establish the relationship of intracellular to extracellular pH in suspensions of isolated hepatocytes and then determine the pH dependence of gluconeogenesis, urea synthesis, respiration, [ATP]/[ADP][Pi], etc. The dependence of regulatory hormone function (norepinephyrine, glucagon) on intra and extracellular pH will also be measured. 5. Measure in vitro dependence of the apparent Km for oxygen on pH, [ATP]/[ADP][Pi] and reduction of cytochrome c using suspensions of isolated mitochondria.
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