Energy requirements for the Na+ gradient in the oxygenated isolated heart: effect of changing the free energy of ATP hydrolysis

Abstract

This study tests the hypothesis that a decrease of the free energy of ATP hydrolysis (Δ GATP) below a threshold value will inhibit Na+-K+-ATPase (Na+ pump) activity and result in an increase of intracellular Na+ concentration ([Na+]i) in the heart. Conditions were designed in which hearts were solely dependent on ATP derived from oxidative phosphorylation. The only substrate supplied was the fatty acid butyrate (Bu) at either low, 0.1 mM (LowBu), or high, 4 mM (HighBu), concentrations. Escalating work demand reduced the Δ GATP of the LowBu hearts. 31P, 23Na, and 87Rb NMR spectroscopy measured high-energy phosphate metabolites, [Na+]i, and Rb+ uptake. Rb+ uptake was used to estimate Na+ pump activity. To measure [Na+]i using a shift reagent for cations, extracellular Ca2+ was reduced to 0.85 mM, which eliminated work demand Δ GATP reductions. Increasing extracellular Na+ ([Formula: see text]) to 200 mM restored work demand Δ GATP reductions. In response to higher [Na+]e, [Na+]i increased equally in LowBu and HighBu hearts to ∼8.6 mM, but Δ GATP decreased only in LowBu hearts. At lowest work demand the LowBu heart Δ GATP was –53 kJ/mol, Rb+ uptake was similar to that of HighBu hearts, and [Na+]i was constant. At highest work demand the LowBu heart Δ GATP decreased to –48 kJ/mol, the [Na+]i increased to 25 mM, and Rb+ uptake was 56% of that in HighBu hearts. At the highest work demand the HighBu heart Δ GATP was –54 kJ/mol and [Na+]i increased only ∼10%. We conclude that a Δ GATP below –50 kJ/mol limits the Na+ pump and prevents maintenance of [Na+]i homeostasis.