The Temperature Dependence of Intracellular pH in Isolated Frog Skeletal Muscle: Lessons Concerning the Na+-H+ Exchanger

We used ³¹P NMR to investigate the temperature-dependence of intracellular pH (pH _i ) in isolated frog skeletal muscles. We found that lnH⁺ _i ] is a linear function of 1/T _abs paralleling those of neutral water (i.e., H⁺= OH⁻) and of a solution containing the fixed pH buffers of frog muscle cytosol. This classical van't Hoff relationship was unaffected by inhibition of glycolysis and was not dependent upon the pH or Na⁺] in the bathing solution. Insulin stimulation of Na⁺-H⁺ exchange shifted the intercept in the alkaline direction but had no effect on the slope. Acid loading followed by washout resulted in an amiloride-sensitive return to the (temperature dependent) basal pH _i . These results show that the temperature dependence of activation of Na⁺-H⁺ exchange is similar to that of the intracellular buffers, and suggest that constancy of H⁺]/OH⁻] with changing temperature is achieved in the short term by intracellular buffering and in the long term by the set-point of the Na⁺-H⁺ exchanger. Proton activation of the exchanger has an apparent standard enthalpy change (ΔH°) under both control and insulin-stimulated conditions that is similar to the ΔH° of the intracellular buffers and approximately half of the ΔH° for the dissociation of water. Thus, the temperature-dependent component of the standard free-energy change (ΔF°) is unaffected by insulin stimulation, suggesting that changes in Arrhenius activation energy (E _a ) may not be a part of the mechanism of hormone stimulation. Received: 12 February 1997/Revised: 1 October 1997