T-cell mediated acute inflammation of the ileum may occur during Crohn's disease exacerbations. During ileal inflammation, absorption of nutrients and electrolytes by villus cells is decreased with a concomitant increase in crypt and/or villus fluid secretion. These alterations lead to fluid accumulation and the subsequent diarrhoea. Net intestinal fluid secretion consists of HCO3−-rich plasma-like fluid. However, the regulation and mechanisms of HCO3− secretion in normal and acutely inflamed ileum are not clearly understood. To study this phenomenon, anti-CD3 monoclonal antibody (mAb)- induced in vivo ileal inflammatory mouse models was used for in vitro functional studies with Ussing chamber and pH stat techniques. Three hours after anti-CD3 mAb injection, ileal mucosa stripped of muscular and serosal layers showed a significant increase in short circuit current (Isc) (0.58 ± 0.07 μEq h−1 cm2versus 1.63 ± 0.14 μEq h−1 cm2). The cAMP-stimulated Isc component was sensitive to glibenclamide but not to DIDS, suggesting that a cystic fibrosis transmembrane conductance regulator (Cftr)-mediated anion conductance was responsible. Basal Cl−-dependent HCO3− secretion, measured using a pH stat technique, was decreased significantly in anti-CD3-injected mice, with a simultaneous increase in Cl−-independent HCO3− secretion that was also inhibited by glibenclamide. Experiments using Cftr−/− mice showed neither an increase in Isc nor an increase in HCO3− secretion, confirming the role for Cftr protein in stimulating anion secretion following anti-CD3 treatment. Western blot analysis indicated that Cftr protein levels were unaltered by anti-CD3 treatment, at least acutely. Finally, an immunoassay for cAMP showed significant increases in intracellular cAMP in villus cells, but not in crypt cells. These studies therefore suggest a shift from a predominantly electroneutral Cl−HCO3− exchange in normal mice, to a predominantly electrogenic anion secretion including HCO3− that occurs via functional Cftr during anti-CD3-mediated acute inflammation.