Bicarbonate permeability of epithelial chloride channels has been studied using the patch-clamp technique. The experiments were performed in excised insideout oriented membrane patches from three different cultured cell types" (a) HT29 colon carcinoma cell line, (b) Ts4 colon carcinoma cell line, and (c) respiratory epithelial cells (REC) in primary culture. In all three preparations we observed outwardly rectifying chloride channels with similar conductances with 145 mmol/1 NaC1 solution in the pipette and in the bath (Cl-pipette/ Cl-bath). When C1-was replaced by HCOs in the bath (C1-/HCO~-) the conductance of the channel at negative clamp voltages was reduced significantly by 40% for HT29 (n = 6), 39% for T84 (r/= 7), and 38% for REC (n = 6). Similarly, the zero-current potential (VI = o) was shifted from 0 mV (C1-/C1-) to negative values (C1-/ HCOs revealing permeability ratios Pca/PHco3 of 2.4 _+ 0.1 for HT29 (n = 6), 2.0 __ 0.1 for T84 (n = 7), and 1.8 _+ 0.1 for REC (n = 7). With NaHCO3 as the pipette solution and NaC1 in the bath, the Vx : o was positive and a Pcl/P~co~ value of 2.3 + 0.1 was determined for HT29 (n = 5). Replacement of C1 in the bath by HCO~ reduced V~ = o to values close to 0 inV. In another series of experiments, the pipette was filled with 145 retool/1 NaC1 and the bath contained 35 retool/1 NaC1 to which 35 mmol/1 NaHCOa were added. We found that neither the conductance for the inward current nor V1 = o was changed significantly with the additon of NaHCOa (HT29, n = 6). We conclude that the HCO~ permeability and HCO~ conductance of these channels is about half of that for C1-.