We investigated the properties of basolateral Cl^– channels in the early distal convoluted tubule (DCT), using the cell-attached and excised configurations of the patch-clamp technique. Microdissected DCTs were isolated from collagenase-treated kidneys after mice were humanely killed. The pipette and bath solutions contained (mM): 140 NaCl, 4.8 KCl, 1.2 MgCl₂, 1 CaCl₂, 10 glucose and 10 Hepes (pH 7.4). We recorded one channel of 9.4 ± 0.3 pS (n = 23, mean ± S.E.M.). In the conditions specified above, the reversal potential (E_r) of the I-V relationship is close to zero (2.0 ± 2.2 mV). Replacing 100 mM NaCl with sodium gluconate shifted E_r by 30.2 ± 3.7 mV (n = 4) towards the right, whereas substituting N-methyl-D-glucamine for Na⁺ had no effect (-1 ± 1.9 mV, n = 5). This indicates Cl^– selectivity. We estimated the influence of extracellular Ca²⁺ and pH on channel activity by measuring the averaged current and NP_o on separate patches under various conditions of Ca²⁺ and pH. For this purpose, as the channel is inhibited at acid intracellular pH (Marvao & Teulon, 2000), the closed current level was assessed on the cell by inhibiting the channels via sodium acetate superfusion. The NP_o measured with zero Ca²⁺ in the pipette was 7.6 ± 1.6 (n = 13). It significantly increased to 33.2 ± 5.6 (n = 10) with 5 mM Ca²⁺ in the pipette. The influence of external pH was dramatic since NP_o significantly increased from 2.8 ± 0.1 (n = 9) at pH 6.4 to 44.7 ± 8.8 at pH 8.0 (n = 10). In contrast, the percentages of patches with active chloride channels were similar under the four conditions (71 and 62 % active patches at 0 and 5 mM Ca²⁺; 58 and 54 % active patches at pH 6.4 and 8.0). Upon excision, it was possible to assess quantitatively the anionic selectivity of the channel in the early DCT despite frequent channel rundown. The channel had the relative permeability sequence Cl^– > Br^– ~ NO₃^– ~ I^– > F^– (n = 4-5). The properties of the channel described here are compatible with the currently known properties of the ClC-K channels.

Student’s unpaired t test was used. P < 0.05 was considered significant.

All procedures accord with current National guidelines.