Effects of temperature and extracellular pH on metabolites: kinetics of anaerobic metabolism in resting muscle by 31P- and 1H-NMR spectroscopy

Abstract

SUMMARYEnvironmental stress, such as low temperature, extracellular acidosis and anoxia, is known to play a key role in metabolic regulation. The aim of the present study was to gain insight into the combined temperature-pH regulation of metabolic rate in frog muscle, i.e. an anoxia-tolerant tissue. The rate of exergonic metabolic processes occurring in resting isolated muscles was determined at 15°C and 25°C as well as at extracellular pH values higher (7.9), similar (7.3) and lower (7.0) than the physiological intracellular pH. 31P and 1H nuclear magnetic resonance spectroscopy high-resolution measurements were carried out at 4.7 T in isolated frog (Rana esculenta) gastrocnemius muscle during anoxia to assess, by means of reference compounds, the concentration of all phosphate metabolites and lactate. Intra- and extracellular pH was also determined.In the range of examined temperatures (15-25°C), the temperature dependence of anaerobic glycolysis was found to be higher than that of PCr depletion (Q10=2.3). High-energy phosphate metabolism was confirmed to be the initial and preferential energy source. The rate of phosphocreatine hydrolysis did not appear to be affected by extracellular pH changes. By contrast, independent of the intracellular pH value, at the higher temperature(25°C) a lowering of the extracellular pH from 7.9 to 7.0 caused a depression in lactate accumulation. This mechanism was ascribed to the transmembrane proton concentration gradient. This parameter was demonstrated to regulate glycolysis, probably through a reduced lactate efflux, depending on the activity of the lactate-H+ co-transporter.The calculated intracellular buffer capacity was related to intra- and extracellular pH and temperature. At the experimental extracellular pH of 7.9 and at a temperature of 15°C and 25°C, calculated intracellular buffering capacity was 29.50 μmol g-1 pH unit-1 and 69.98 μmol g-1 pH unit-1, respectively.