The mixing of gastric and pancreatic juice subjects the jejunum to unique ionic conditions with high luminal CO2 tension and HCO3 (-) concentration. We investigated the role of the small intestinal apical anion exchangers PAT-1 (Slc26a6) and DRA (Slc26a3) in basal and CO2/HCO3 (-)-stimulated jejunal fluid absorption. Single pass perfusion of jejunal segments was performed in anaesthetised wild type (WT) as well as in mice deficient in DRA, PAT-1, Na+/H+ exchanger 3 (NHE3) or NHE2, and in carbonic anhydrase II (CAII). Unbuffered saline (pH 7.4) perfusion of WT jejunum resulted in fluid absorption and acidification of the effluent. DRA-deficient jejunum absorbed less fluid than WT, and acidified the effluent more strongly, consistent with its action as a Cl-/HCO3 (-) exchanger. PAT-1-deficient jejunum also absorbed less fluid but resulted in less effluent acidification. Switching the luminal solution to a 5 % CO2/HCO3 (-) buffered solution (pH 7.4), resulted in a decrease in jejunal enterocyte pH(i) in all genotypes, an increase in luminal surface pH and a strong increase in fluid absorption in a PAT-1- and NHE3- but not DRA-, CAII, or NHE2-dependent fashion. Even in the absence of luminal Cl-, luminal CO2/HCO3 (-) augmented fluid absorption in WT, CAII, NHE2- or DRA-deficient, but not in PAT-1- or NHE3-deficient mice, indicating the likelihood that PAT-1 serves to import HCO3 (-) and NHE3 serves to import Na+ under these circumstances. The results suggest that PAT-1 plays an important role in jejunal Na+HCO3 (-) reabsorption, while DRA absorbs Cl- and exports HCO3 (-) in a partly CAII-dependent fashion. Both PAT-1 and DRA significantly contribute to intestinal fluid absorption and enterocyte acid/base balance but are activated by different ion gradients.