SummaryThe relation between the energies of ion movement and ATP hydrolysis is unknown in tissues with widely varying electric potentials. Consequently, we measured the concentration of the nine major inorganic ions in the extra-and intracellular phases in heart, liver, and red cells with resting electrical potentials, E N , of −86, −28, and −6 mV, respectively, under six different physiological conditions. We calculated the Nernst electric potential and the energy of ion movement between the phases. We found that the energy of ATP hydrolysis was essentially constant, between −54 and −58 kJ/mol, in all tissues and conditions. In contrast, as E N decreased, the energies of the Na + and K + gradients decreased, with slopes approximating their valence. The difference between the energies of Na + and K + gradients remained constant at 17 kJ/mol, which is approximately one third of the energy of ATP hydrolysis, demonstrating near-equilibrium of the Na + /K + ATPase in all tissues under all conditions. All cations, except K + , were pumped out of cells and all anions, except Cl − in liver and red cell, were pumped into cells. We conclude that the energy of ATP was expressed in Na + /K + ATPase and its linked inorganic ion transporters to create a Gibbs-Donnan near-equilibrium system, an inherent part of which was the electric potential.

From 31P NMR measurements made in vitro at 38 degrees C, I = 0.25, pH 5. 75-8.5, and calculated free [Mg2+] from 0 to 5 mM, we show that, within the physiological range of cytosolic free [Mg2+] from 0.25 to 1.5 mM, the chemical shift difference between the alpha- and beta-ATP resonances, deltaalphabeta, changes by only 0.6 ppm. Consequently, we developed new formalisms from known acid and Mg2+ dissociation constants by which the observed chemical shift of Pi, deltaPi, and the peak height ratio of the beta- and alpha-ATP resonances, hbeta/alpha, could be related to free [Mg2+] by simultaneous solution of: [equation: see text] We found that hbeta/alpha changed 2.5-fold as free [Mg2+] varied from 0.25 to 1.5 mM, providing a more sensitive and accurate measure of free cytosolic [Mg2+]. In working rat heart perfused with glucose, free [Mg2+] was 1.0 +/- 0.1 from hbeta/alpha and 1.2 +/- 0.03 from measured [citrate]/[isocitrate] but 0.51 +/- 0.1 from deltaalphabeta. Addition of ketone bodies to the perfusate decreased free [Mg2+] estimated from hbeta/alpha to 0.61 +/- 0.02 and 0.74 +/- 0.11 by [citrate]/[isocitrate] but the estimate from deltaalphabeta was unchanged at 0.46 +/- 0.04 mM. Such differences in estimated free [Mg2+] alter the apparent Keq of the creatine kinase reaction and hence the estimated cytosolic free [SigmaADP].

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