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B.A. Hughes, A. Swaminathan; Kir7.1 Potassium Channels Are Directly Regulated by Intracellular pH . Invest. Ophthalmol. Vis. Sci. 2005;46(13):3327.
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Purpose: Potassium secretion by Kir7.1 channels in the RPE apical membrane supports Na+/K+ ATPase activity and regulates the concentration of K+ in the subretinal space. Our previous patch–clamp studies on bovine RPE cells indicated that Kir7.1 channels are regulated by intracellular pH (pHi). The purpose of this study was to determine whether sensitivity to pHi is an inherent property of the Kir7.1 channel subunit. Methods: cRNA was synthesized from plasmid containing either wild type or mutant human Kir7.1 and injected into Xenopus oocytes for electrophysiological recording 1–4 days later. For intact oocytes, currents were recorded via the two–electrode voltage–clamp technique and pHi was acidified by the addition of butyrate to the bath. Recordings from excised inside–out macro–patches were used to test directly the effect of changes in pHi on Kir7.1 conductance. Results: In intact oocytes, Kir7.1 conductance was enhanced by mild intracellular acidification but inhibited by strong acidification. Mild acidification of the solution bathing the cytosolic face of inside–out membrane patches enhanced Kir7.1 conductance, but strong acidification or alkalinization inhibited it. Site–directed mutagenesis of histidine residues in the N– and C–terminal domains of the Kir7.1 subunit followed by functional analysis led to the identification of a histidine in the N–terminus that is crucial for the acid–induced activation of Kir7.1 channels in the physiological pHi range. Conclusions: The results indicate that pHi sensitivity is an inherent property of Kir7.1 channels and that it depends on a histidine residue in the N–terminus of the Kir7.1 channel subunit. We conclude that intracellular protons directly interact with Kir7.1 channels to modulate their activity and that this may provide an important mechanism by which K+ secretion into the subretinal space is controlled.
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