ROLE OF CA IN THE REGULATION OF ENERGY METABOLISM

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This project is designed to assess the physiological importance of calcium-ions in the regulation of energy metabolism. Specifically, we have exposed isolated cardiac myocytes to a variety of conditions expected to alter cytosolic free calcium ion concentration and have measured the amount of the active form of pyruvate dehydrogenase ([PDHA]) which results, as well as estimating calcium concentration with fluorescent chelating agents. Protocols have included progressive plasma membrane depolarization with increasing KCl, and the use of veratridine. Further, we have used interventions which enforce quiscence on the cells, viz. loading with chelating agents and the use of the inhibitor ryanodine, to separate the effect of calcium on pyruvate dehydrogenase from the effects of a decrease in the ATP/ADP ratio, which is normally a sequel to increased calcium availability in muscle. These studies have shown that calcium is quantitatively an important signal in controlling pyruvate dehydrogenase activity in heart cells. We have also investigated the role of calcium in the control of pyruvate dehydrogenase in isolated hepatocytes. The hormones glucagon, vasopressin and the alpha1-adrenergic agonist phenylephrine all led to an increase in cytosolic free calcium concentration, an increase in PDHA content and an increase in mitochondrial NADH content: the latter is indicative of increased mitochondrial dehydrogenase activity in general. Further, phorbol esters, which activate protein kinase C, were found to prevent increases in calcium due to glucagon or phenylephrine, but not that due to vasopressin. In every case, there was a parallelism between effects on cytosolic free calcium concentration and effects on PDHA content, in strong support of the thesis that the former exercises a large degree of control over the latter.