Carbonic anhydrases (CA) comprise a large family of enzymes, which catalyze the equilibration of CO₂, H⁺ and HCO₃^-, thus accelerating pH changes associated with CO₂ and HCO₃^-. Most cells are equipped with both intracellular and extracellular CA activity, which appear to take over different roles for pH homeostasis in cells and tissues. We have used Xenopus oocytes to express extracellular CAIV and/or cytosolic CAII, and have measured intracellular pH (pH_i) shifts and pH transients at the cell surface (pH_s) using pH-sensitive microelectrodes to evaluate the effect of the different locations of CA activity for pH shifts induced by addition and removal of CO₂/HCO₃^-. CA expression and CA activity in oocytes were confirmed by immunocytochemistry, Western blot, and mass spectrometry, respectively. Both, CAII and CAIV accelerated the rate of intracellular pH change, but only CAIV also greatly increased the pH_s transients. All CA-related changes of pH were suppressed by the CA inhibitor ethoxyzolamide. Expressing the catalytically non-active mutants of CAII (CA-V143Y) and of CAIV (CAIV-V165Y) resulted in pH shifts similar to those recorded in native oocytes. Co-expressing the sodium-bicarbonate cotransporter (NBCe1) with the CAs had little effect on the pH_s transients, but modulated the intracellular pH shifts. Our results indicate that intra- and extracellular CA activities are crucial for the rate of pH change on either side of the cell membrane, and (thus) also for the rate of bicarbonate rise and fall in cells. In addition, pH_s measurements are suitable to detect extracellular, but not intracellular CA activity. Supported by the Deutsche Forschungsgemeinschaft, DE231/24-1