Cytoplasmic pH and free Mg2+ in lymphocytes

Abstract

Measurements have been made of cytoplasmic pH, (pHi) and free Mg2+ concentration (Mg2+), in pig and mouse lymphocytes, pHi was measured in four ways: by a digitonin null-point technique; by direct measurement of the pH of freeze-thawed cell pellets; from the 31p nuclear magnetic resonance (NMR) spectrum of intracellular inorganic phosphate; and by the use of a newly synthesized, intracellularly-trappable fluorescent pH indicator. In HEPES buffered physiological saline with pH 7.4 at 37°C, pHi was close to 7.0. Addition of physiological levels of HCO3− and CO2 transiently acidified the cells by ~0.1 U. Mitogenic concentrations of concanavatin A (Con A) had no measurable effect on pH in the first hour. Mg2+- was assessed in three ways: (a) from the external Mg2+ null-point at which the ionophore A23187 produced no net movement of Mg2+ or H+; (b) by Mg-sensitive electrode measurements in freezethawed pellets; and (c) from the 31p nuclear magnetic resonance spectrum of the γ,-phosphate of intracellular ATP. Total cell Mg2+ was ~12 mmol per liter cell water. The NMR data indicated [Mg2+], >0.5 mM. The null-point method gave Mg2+-0.9 nM. The electrode measurements gave 1.35 raM, which was thought to be an overestimate. Exposure to mitogenic doses of Con A for 1 h gave no detectable change in total or free Mg2+. © 1982, Rockefeller University Press., All rights reserved.