The mechanisms of chloride transport within the connecting tubule (CNT), a segment interposed between the distal convoluted tubule and the collecting tubule, is presently indetermined. We investigated basolateral chloride channels in the mouse CNT, using the cell-attached and inside-out variants of the patch-clamp method. Microdissected CNTs were isolated from collagenase-treated kidneys (Lourdel et al. 2003) after mice were humanely killed. The pipette solution contained (mM): 145 NaCl, 1 CaCl₂, 1 MgCl₂, 10 glucose and 10 Hepes at pH 7.4. The bath solution was similar except that it contained 5 mM KCl and 140 NaCl. In cell-attached patches, we found a channel with a conductance of 10.6 ± 0.9 pS (mean ± S.E.M.; n = 8 ) which is chloride-selective : the reversal potential significantly changed from 1.9 ± 4.0 mV in the control (n = 8), to 25.7 ± 1.2 mV in the presence of 100 mM Na-gluconate (n = 4), whereas there was no significant change (-1.1 ± 2.0 mV, n = 8) with 140 mM NMDG-chloride. We estimated the influence of extracellular Ca²⁺ and pH on channel activity by measuring the NP_o on separate patches. The NP_o dramatically increased from 23 ± 6 (n = 11) with 1 mM Ca²⁺ in the pipette to 101 ± 12 (n = 10) with 5 mM Ca²⁺. The NP_o also significantly increased with extracellular pH, from 26 ± 7 (n = 13) at pH 7.0 to 71 ± 12 at pH 7.8 (n = 12). The anionic selectivity was investigated quantitatively in the excised configuration and showed the relative permeability sequence Cl^– > Br^– ≈ NO₃^– > F^– (n = 4-6). The properties of the channel appear similar to a previously reported Cl^– channel in the DCT (Lourdel et al. 2003) and are compatible with the currently known properties of the ClC-K channels (Estevez et al. 2001). The unpaired Student’s t test was used. P < 0.05 was considered significant.