Distinguishing HCO 3 − from CO 3 2− transport by NBCe1‐A

Using a pH electrode to monitor extracellular pH in brain slices, Grichtchenko et al. observed pH changes that they at first attributed to HCO 3 − transport and therefore expected to be reduced by an inhibitor of carbonic anhydrase (CA). However, they found just the opposite and hypothesized that an electrogenic Na + ‐HCO 3 − cotransporter (NBCe) carries CO 3 2− rather than HCO 3 − . In the present study, we measure surface pH (pH S ) while voltage‐clamping a Xenopus oocyte expressing both NBCe1‐A (stoichiometry, 1 Na + : 2 HCO 3 − equivalents) and CA IV (catalytic domain on the cell surface). If NBCe1‐A moves HCO 3 − into the cell, CA IV will replenish the lost HCO 3 − at the cell's outer surface and as a consequence produce H + . If instead NBCe1‐A moves CO 3 2− into the cell, CA IV will consume H + . Because CA IV would produce H + in one case and consume it in the other, we can distinguish HCO 3 − from CO 3 2− transport by blocking CA IV. We found that shifting the holding potential (V h ) elicited predictable changes in pH S and current. Blocking CA IV with acetazolamide (ACZ) accentuated all pH S changes. When we expressed NBCe1‐A but not CA IV, the pH S changes were already large and unaffected by ACZ. In experiments on oocytes injected with H 2 O rather than cRNA, we did not observe any pH S changes or current changes in response to sudden shifts in V h . In conclusion, our findings are consistent with a model in which NBCe1‐A transports CO 3 2− rather than HCO 3 −