Background: CF patients suffer from a variety of gastrointestinal problems, which may all be directly or indirectly linked to the inability of intestinal mucosa to secrete HCO₃^– and to inhibit Na⁺/H⁺ mediated fluid absorption and proton secretion. This has led to an intense search for alternative modes of intestinal anion transport, and expressions of a variety of potential alternative intestinal anion transporters have been reported. Aim: To delineate the dependency of different modes of intestinal HCO₃^– secretion on CFTR expression. Methods and Results: We studied acid-, agonist- and HCO₃^– stimulated as well as Cl^–-dependent HCO₃^– secretion in the CFTR^tm1cam and WT murine duodenum in vivo. NHE3 and Slc26a6-deficient mice were used for selected questions. Luminal acid, forskolin, heat-stable E. coli enterotoxin (STa), PG E2, carbachol, and an increase of blood HCO₃^– all stimulated duodenal HCO₃^– secretion in anesthetized (10 µl/g I.P. haloperidol/midazolam/fentanyl cocktail (haloperidol 12.5 mg/kg, fentanyl 0.325 mg/kg and midazolam 5 mg/kg body weight)) WT but not in CFTR^tm1cam mice. Pharmacological inhibition or genetic ablation of NHE3 resulted in a significantly higher basal HCO₃^– secretory rate, which was electroneutral and therefore due to an unmasking of apical Cl^–/HCO₃^– exchange activity. Accordingly, Slc26a6 ablation attenuated S1611-induced J_HCO3^–. Removal of luminal Cl^– reverted basal HCO₃^– secretion to H⁺ secretion, but surprisingly, forskolin was able to elicit a full HCO₃^– secretory response. In the absence of CFTR, electroneutral NaCl absorptive rates were similar to WT rates, but S1611 induced virtually no increase in HCO₃^– secretion. Conclusion: This indicates that the apical anion exchangers Slc26a6 and Slc26a3 need proton recycling via NHE3 to operate in the Cl^– absorptive mode, and Cl^– exit via CFTR to operate in the HCO₃^– secretory mode. In addition, Cl^– independent HCO₃^– secretion can be stimulated by cAMP increase, likely from the crypt reagion, which expresses high levels of CFTR but none of the anion exchangers.