Primary structure and functional expression of a cortical collecting duct K(ir) channel

Maintenance of a negative membrane potential in the cortical collecting duct (CCD) principal cell depends on a small-conductance, inward-rectifying basolateral membrane K⁺ (K(ir)) channel. In the present study, a candidate cDNA encoding this K⁺ channel, CCD-IRK₃, was isolated from a mouse collecting duct cell line, M1. CCD-IRK₃ shares a high degree of homology with a human brain inward-rectifier K⁺ channel (K(ir) 2.3). By Northern analysis, CCD-IRK₃ transcript (2.9 kb) was readily detected in M1 CCD cells but not in Madin-Darby canine kidney, LLC-PK₁, Chinese hamster ovary, or monkey kidney fibroblast cell lines. CCD-IRK₃-specific reverse transcription-polymerase chain reaction confirmed bonafide expression in the kidney. Functional expression studies in Xenopus oocytes revealed that CCD- IRK₃ operates as strongly inward-rectifying K⁺ channel. The cation selectivity profile of CCD-IRK₃[ionic permeability values (P(K)/P(i)), Tl ≤ Rb ≤ K⁺ >> NH₄ > Na; inward-slope conductance (G(K)/G(i)), Tl ≤ K⁺ >> NH₄ > Na > Rb] is similiar to the macroscopic CCD basolateral membrane K⁺ conductance (G(K)/G(i), K⁺ >> NH₄ > Rb; P(K)/P(i), Rb ≃ K⁺ >> NH₄). CCD- IRK₃ also exhibits the pharmacological features of the native channel. Patch-damp analysis reveals that CCD-IRK₃ functions as a high open probability, voltage-independent, small-conductance channel (14.5 pS), consistent with the native channel. Based on these independent lines of evidence, CCD-IRK₃ is a possible candidate for the small-conductance basolateral K(ir) channel in the CCD.