Reversed electrogenic sodium bicarbonate cotransporter1 is the major acid loader during recovery from cytosolic alkalosis in mouse cortical astrocytes

Recovery of intracellular pH from cytosolic alkalosis has been attributed primarily to Cl- coupled acid loaders/base extruders such as Cl-/HCO3- or Cl-/OH- exchangers. We have studied this process in cortical astrocytes from wild-type and transgenic mouse models with gene deletion for the electrogenic sodium bicarbonate cotransporter1 (NBCe1) and for carbonic anhydrase (CA) isoform II. An acute cytosolic alkalosis was induced by the removal of either CO2/HCO3- or butyric acid, and the subsequent acid loading was analysed by monitoring changes in cytosolic H+ or Na+ using ion-sensitive fluorescent dyes. We have identified that NBCe1 reverses during alkalosis and contributes more than 70% to the rate of recovery from alkalosis by extruding Na+ and HCO3-. After CA inhibition or in CAII-knockout (KO) cells, the rate of recovery was reduced by 40%, and even by 70% in the nominal absence of CO2/HCO3-. Increasing the extracellular K+ concentration modulated the rate of acid loading in wild-type cells, but not in NBCe1-KO cells. Removing chloride had only a minor effect on the recovery from alkalosis. Reversal of NBCe1 by reducing pH/[HCO3-] was demonstrated in astrocytes and in Xenopus oocytes, in which human NBCe1 was heterologously expressed. The results obtained suggest that reversed NBCe1, supported by CAII activity, plays a major role in acid-loading cortical astrocytes to support recovery from cytosolic alkalosis.