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  • Title: Relationships between cytosolic [ATP], [ATP]/[ADP] and ionic fluxes in the perfused rat heart: A 31P, 23Na and 87Rb NMR study.
    Author: Stewart LC, Deslauriers R, Kupriyanov VV.
    Journal: J Mol Cell Cardiol; 1994 Oct; 26(10):1377-92. PubMed ID: 7869398.
    Abstract:
    In order to assess the relationship between cytosolic [ATP] or [ATP]/[ADP] and the intracellular Na+ concentration ([Na+]i), we have used the phosphate trap 2-deoxy-D-glucose (DG) to alter the high energy phosphate levels in rat cardiomyocytes. Pyruvate-perfused rat hearts were treated with 2 mM DG in the presence of 10IU/l of insulin for 28 min, followed by perfusion with DG without insulin for 60 min. The DG + insulin treatment resulted in dramatic changes in the 31P NMR spectra: phosphocreatine (PCr) and total ATP decreased (to 15 and 35%, respectively) and deoxyglucose-6-phosphate accumulated, with little change in either inorganic phosphate or intracellular pH. These changes corresponded to a decrease in cytoplasmic [ATP] (from 7.6 to 1.8 mM), [ATP]/[ADP] (from 494 to 24) and ATP affinity [A(ATP), by 8.9 kJ/mol] and an increase in [ADP] (five-fold) and free [Mg2+] (two-fold). Subsequent perfusion with DG--insulin resulted in slow recovery of PCr, [ATP]/[ADP] and A(ATP) such that the "low energy" state lasted an additional 16 min during which ATP remained low and constant. There were no detectable changes in the intracellular Na+ content as assessed by shift reagent-aided 23Na NMR at the end of DG + insulin treatment (98 +/- 18%, 28-36 min of the protocol). In addition, there was no change in the Rb+ influx rate as measured by 87Rb NMR at the beginning of insulin washout which was achieved by replacing 20% of the KCl with RbCl ([K+] = 3.76 mM, [Rb+] = 0.94 mM). During DG + insulin treatment the pressure-rate product (PRP) decreased by half and was restored upon insulin washout to 80% of its initial value both in the presence and in the absence of the shift reagent [5 mM Dy (triethylenetetraminehexaacetate)3-]. These data imply that unfavorable thermodynamic [low A(ATP)] and kinetic (low [ATP] and [ATP]/[ADP]) conditions induced by DG treatment do not inhibit Na+, K(+)-ATPase activity. We speculate that during anoxia when changes in [ATP]/[ADP] are comparable to those induced by DG treatment, the observed increase in [Na+]i is not due to inhibition of the Na+ pump by reduced [ATP] or [ATP]/[ADP].
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