Variations of intracellular pH in human erythrocytes via K+(Na+)/H+ exchange under low ionic strength conditions

The change of intracellular pH of erythrocytes under different experimental conditions was investigated using the pH-sensitive fluorescent dye BCECF a nd correlated with (ouabain + bumetanide + EGTA)-insensitive K+ efflux and Cl- loss. When human erythrocytes were suspended in a physiological NaCl so lution (pH(o) = 7.4), the measured pH(i) was 7.19 +/- 0.04 and remained con stant for 30 min. When erythrocytes were transferred into a low ionic stren gth (LIS) solution, an immediate alkalinization increased the pH(i) to 7.70 +/- 0.15, which was followed by a slower cell acidification. The alkaliniz ation of cells in LIS media was ascribed to a band 3 mediated effect since a rapid loss of approximately 80% of intracellular Cl- content was observed , which was sensitive to known anion transport inhibitors. In the case of c ellular acidification, a comparison of the calculated H+ influx with the me asured unidirectional K+ efflux at different extracellular ionic strengths showed a correlation with a nearly 1:1 stoichiometry. Both fluxes were enha nced by decreasing the ionic strength of the solution resulting in a H+ inf lux and a K+ efflux in LIS solution of 108.2 +/- 20.4 mmol (1(cells) hr)(-1 ) and 98.7 +/- 19.3 mmol (1(cells) hr)(-1), respectively. For bovine and po rcine erythrocytes, in LIS media, H+ influx and K+ efflux were of comparabl e magnitude, but only about 10% of the fluxes observed in human erythrocyte s under LIS conditions. Quinacrine, a known inhibitor of the mitochondrial K+(Na+)/H+ exchanger, inhibited the K+ efflux in LIS solution by about 80%. Our results provide evidence fur the existence of a K+(Na+)/H+ exchanger i ll the human erythrocyte membrane.