Expression and permeation properties of the K+ channel Kir7.1 in the retinal pigment epithelium

1. Bovine Kir7.1 clones were obtained from a retinal pigment epithelium (RP E)-subtracted cDNA library. Human RPE cDNA library screening resulted in cl ones encoding full-length human Kir7.1. 2. Northern blot analysis indicated that bovine Kir7.1 is highly expressed in the RPE. 3. Human Kir7.1. channels were expressed in Xenopus oocytes and studied usi ng the two-electrode voltage-clamp technique. 4. The macroscopic Kir7.1 conductance exhibited mild inward rectification a nd an inverse dependence on extracellular K+ concentration ([K+](o)). The s electivity sequence based on permeability ratios was K+ (1.0) approximate t o Rb+ (0.89) > Cs+ (0.013) > Na+ (0.003) approximate to Li+ (0.001) and the sequence based on conductance ratios was Rb+ (9.5) much greater than K+ (1 .0)> Na+ (0.458) > Cs+ (0.331) > Li+ (0.139). 5. Non-stationary noise analysis of Rbf currents in cell-attached patches y ielded a unitary conductance for Kir7.1. of similar to2 pS. 6. In whole-cell recordings from freshly isolated bovine RPE cells, the pre dominant current was a mild inwardly rectifying K+ current that exhibited a n inverse dependence of conductance on [K+](o). The selectivity sequence ba sed on permeability ratios was K+ (1.0) approximate to Rb+ (0.89) > Cs+ (0. 021) > Na+ (0.003) approximate to Li+ (0.002) and the sequence based on con ductance ratios was Rb+ (8.9) much greater than K+ (1.0) > Na+ (0.59) > Cs (0.23) > Li+ (0.08). 7. 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. 8. Non-stationary noise analysis of Rbf currents-in cell-attached apical me mbrane patches yielded a unitary conductance for RPE Kir of similar to2 pS. 9. On the basis of this molecular and electrophysiological evidence, we con clude that Kir7.1 channel subunits comprise the K+ conductance of the RPE a pical membrane.