CELLULAR CA2+ ION HOMEOSTASIS AND THE IMPACT OF AGING

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This project constitutes an investigation into mechanism whereby cells achieve the homeostasis of cytosolic free Ca2+ concentrations ((Ca2+)c), and allow perturbations in (Ca2+)c in response to hormones and neurotransmitters. Further, it addressed derangements in these control mechanisms which may occur in old-age. This year, we have asked the following questions. (1) What is the mechanism whereby the hormone glucagon leads to an increase in (Ca2+)c in hepatocytes? We have investigated the phosphorylation of proteins in a cellular fraction enriched in plasma membrane, on treatment with either cyclic-AMP or exogenous protein kinase A, in a procedure which mimics exposure of the cell o glucagon. Direct correlation between such phosphorylation events and changes in plasma membrane permeability to Ca2+ is not yet possible, however. (2) Are there correlates at the level of protein phosphorylation of the previously described decreased responsiveness of contractility to catecholamines in the aging heart? We have quantitated a decreased phosphate content of the proteins troponin- I and C-protein, on exposure of isolated cardiac myocytes to norepinephrine, when cells from senescent animals are compared to those from young adults. This is linked to an inferred decreased net formation of cyclic-AMP, in cells from the senescent animals. This may be expected to have an impact also on sarcolemmal and sarcoplasmic reticulum proteins responsible for Ca2+ transport. (3) Does ATP, which is released on sympathetic stimulation together with norepinephrine, have a positive inotropic effect, linked to mobilization of (Ca2+)c, when added at appropriate levels to isolated cardiac myocytes? Micromolar concentrations of exogenous ATP were found to be as potent a positive inotrope as norepinephrine at the same concentrations, and to give rise t an enhanced transient in (Ca2+)c on electrical stimulation. The pharmacology of the response was studied with other purine nucleotides and analogs.