Expression and permeation properties of the K(+) channel Kir7.1 in the retinal pigment epithelium.
Bovine Kir7.1 clones were obtained from a retinal pigment epithelium (RPE)-subtracted cDNA library. Human RPE cDNA library screening resulted in clones encoding full-length human Kir7.1. Northern blot analysis indicated that bovine Kir7.1 is highly expressed in the RPE. Human Kir7.1 channels were expressed in Xenopus oocytes and studied using the two-electrode voltage-clamp technique. The macroscopic Kir7.1 conductance exhibited mild inward rectification and an inverse dependence on extracellular K+ concentration ([K+]o). The selectivity sequence based on permeability ratios was K+ (1.0) approximately Rb+ (0.89) > Cs+ (0.013) > Na+ (0.003) approximately Li+ (0.001) and the sequence based on conductance ratios was Rb+ (9.5) >> K+ (1.0) > Na+ (0.458) > Cs+ (0.331) > Li+ (0.139). Non-stationary noise analysis of Rb+ currents in cell-attached patches yielded a unitary conductance for Kir7.1 of approximately 2 pS. In whole-cell recordings from freshly isolated bovine RPE cells, the predominant current was a mild inwardly rectifying K+ current that exhibited an inverse dependence of conductance on [K+]o. The selectivity sequence based on permeability ratios was K+ (1.0) approximately Rb+ (0.89) > Cs+ (0.021) > Na+ (0.003) approximately Li+ (0.002) and the sequence based on conductance ratios was Rb+ (8.9) >> K+ (1.0) > Na+ (0.59) > Cs+ (0.23) > Li+ (0.08). In cell-attached recordings with Rb+ in the pipette, inwardly rectifying currents were observed in nine of 12 patches of RPE apical membrane but in only one of 13 basolateral membrane patches. Non-stationary noise analysis of Rb+ currents in cell-attached apical membrane patches yielded a unitary conductance for RPE Kir of approximately 2 pS. On the basis of this molecular and electrophysiological evidence, we conclude that Kir7.1 channel subunits comprise the K+ conductance of the RPE apical membrane.
W. K. Kellogg Eye Center, Department of Ophthalmology and Visual Sciences, University of Michigan, Ann Arbor, MI 48105, USA.
M
M
Shimura
Y
Y
Yuan
J T
JT
Chang
S
S
Zhang
P A
PA
Campochiaro
D J
DJ
Zack
B A
BA
Hughes
D000818:Animals; D016477:Artifacts; D015152:Blotting, Northern; D002417:Cattle; D002462:Cell Membrane / Q000378:metabolism; D003001:Cloning, Molecular; D004553:Electric Conductivity; D006801:Humans; D009865:Oocytes / Q000378:metabolism; D018408:Patch-Clamp Techniques; D010539:Permeability; D010857:Pigment Epithelium of Eye / Q000378:metabolism*; D015221:Potassium Channels / Q000378:metabolism* / Q000502:physiology; D024661:Potassium Channels, Inwardly Rectifying; D014018:Tissue Distribution; D014981:Xenopus
D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying