Welling P A
Department of Physiology, University of Maryland School of Medicine, Baltimore 21201, USA.
Am J Physiol. 1997 Nov;273(5):F825-36. doi: 10.1152/ajprenal.1997.273.5.F825.
Maintenance of a negative membrane potential in the cortical collecting duct (CCD) principal cell depends on a small-conductance, inward-rectifying basolateral membrane K+ (Kir) 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 (Kir 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 (PK/Pi), Tl > or = Rb > or = K+ >> NH4 > Na; inward-slope conductance (GK/Gi), Tl > or = K+ >> NH4 > Na > Rb] is similar to the macroscopic CCD basolateral membrane K+ conductance (GK/Gi, K+ >> NH4 > Rb; PK/Pi, Rb approximately equal to K+ >> NH4). CCD-IRK3 also exhibits the pharmacological features of the native channel. Patch-clamp 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 Kir channel in the CCD.
皮质集合管（CCD）主细胞中负膜电位的维持依赖于一种小电导、内向整流的基底外侧膜钾离子（Kir）通道。在本研究中，从一种小鼠集合管细胞系M1中分离出了编码该钾离子通道的候选互补DNA（cDNA），即CCD - IRK3。CCD - IRK3与人脑内向整流钾离子通道（Kir 2.3）具有高度同源性。通过Northern分析，在M1 CCD细胞中很容易检测到CCD - IRK3转录本（2.9 kb），但在Madin - Darby犬肾细胞、LLC - PK1细胞、中国仓鼠卵巢细胞或猴肾成纤维细胞系中未检测到。CCD - IRK3特异性逆转录 - 聚合酶链反应证实其在肾脏中真实表达。在非洲爪蟾卵母细胞中的功能表达研究表明，CCD - IRK3作为一种强内向整流钾离子通道发挥作用。CCD - IRK3的阳离子选择性特征[离子通透性值（PK/Pi），铊≥铷≥钾离子>>铵离子>钠离子；内向斜率电导（GK/Gi），铊≥钾离子>>铵离子>钠离子>铷]与宏观的CCD基底外侧膜钾离子电导（GK/Gi，钾离子>>铵离子>铷；PK/Pi，铷约等于钾离子>>铵离子）相似。CCD - IRK3也表现出天然通道的药理学特征。膜片钳分析表明，CCD - IRK3作为一种高开放概率、电压不依赖的小电导通道（14.5 pS）发挥作用，这与天然通道一致。基于这些独立的证据线索，CCD - IRK3可能是CCD中小电导基底外侧Kir通道的候选者。
