Intracellular pH regulation in primary rat astrocytes and C6 glioma cells

Publisher Website
Google Scholar
Abstract
We used the pH‐sensitive fluorescent dye BCECF to study intracellular pH (pHi) regulation in primary cultures of rat astrocytes and C6 glioma cells. Both cell types contain three pH‐regulating transporters: (1) alkalinizing Na+/H+ exchange; (2) alkalinizing Na+ + HCO3 −/Cl− exchange; and (3) acidifying Cl−/HCO3− exchange. Na+/H+ exchange was most evident in the absence of CO2; recovery from acidification was Na+ dependent and amiloride sensitive. Exposure to CO2 caused a cell alkalinization that was inhibited by DIDS, dependent on external Na+, and inhibited 75% in the absence of Cl− (thus mediated by Na+ + HCO3−/Cl− exchange). When pHi was increased above the normal steady‐state pHi, a DIDS‐inhibitable and Na+ ‐independent acidifying recovery was evident, indicating the presence of Cl− /HCO3− exchange. Astrocytes, but not C6 cells, contain a fourth pH‐regulating transporter, Na+ −HCO3− cotransport; in the presence of CO2, depolarization caused an alkalinization of 0.12 +− 0.01 (n = 8) and increased the rate of CO2‐induced alkalinization from 0.23 ± 0.02 to 0.42 ± 0.03 pH unit/min. Since C6 cells lack the Na+ ‐HCO3+ cotransporter, they are an inferior model of pHi regulation in glia. Our results differ from previous observations in glia in that: (1) Na+ /H+ exchange was entirely inhibited by amiloride; (2) Na+ + HCO3−/Cl− exchange was present and largely responsible for CO2−induced alkalinization; (3) Cl− /HCO3− exchange was only active at pHi values above steady state; and (4) depolarization‐induced alkalinization of astrocytes was seen only in the presence of CO2.