Voltage-dependent Na+-HCO3-cotransporter and Na+/H+ exchanger are involved in intracellular pH regulation of cultured mature rat cerebellar oligodendrocytes

Abstract

Intracellular pH (pH(i)) was measured at 37°C in mature rat cerebellar oligodendrocytes dissociated in culture by using the pH-sensitive probe BCECF. Cells were identified by anti-galactocerebroside antibody. The mean steady-state pH(i) was 7.02 in the absence of CO2/bicarbonate (Hepes- buffered solution) at an external pH of 7.40 and 7.04 in 5% CO2/25 mM bicarbonate-buffered solution at the same external phi this value was modified neither by the removal of external chloride nor by the addition of the chloride-coupled transport blocker DIDS. In both external solutions steady-state pH(i) values were strongly dependent on external pH. In Hepes- buffered solution pH(i) recovery following an acid load required external Na+ and was completely inhibited by amiloride, indicating the presence of a Na+/H+ exchanger. In CO2/bicarbonate-buffered solution amiloride partially reduced the phi recovery rate, indicating the presence of a bicarbonate- dependent phi regulating mechanism. Membrane depolarization induced by increasing external K+ concentration elicited an alkalinization only in the presence of external Na+ and bicarbonate. Analysis of the calculated HCO3 fluxes with respect to membrane potential indicated that these fluxes were mediated by a Na+-HCO3 cotransport with a stoichiometry of 1:3. These results demonstrate that a Na+/H+ exchanger and a Na+-HCO3 cotransporter are involved in phi regulation of mature oligodendrocytes.