1. Archive
2. Vol.51, No.4

An ATP-Regulated and pH-Sensitive Inwardly Rectifying K⁺ Channel in Cultured Human Proximal Tubule Cells

Abstract: Although renal K⁺ channels along the nephron have been explored in various animal species, little is known about the K⁺ channels in human proximal tubule cells. Using the patch-clamp technique, we investigated the properties of an inwardly rectifying K⁺ channel present in the surface membrane of cultured human proximal tubule cells of normal kidney origin. This channel was the most frequently observed K⁺ channel in cell-attached patches, and cytoplasmic ATP was required to maintain channel activity in inside-out patches. Its single channel conductance was about 42 pS for inward currents and 7 pS for outward currents under the symmetrical K⁺ condition. The ATP effect on channel activity was dose-dependently stimulatory within a range of 0.1 to 10 mM, and a nonhydrolyzable ATP analog, AMP-PNP (3 mM), had no effect on channel activity in either the presence or absence of ATP (1 mM). The channel activity observed in cell-attached patches was reduced to 30 to 50% of controls by a membrane-permeable nonspecific protein kinase inhibitor, K252a (1 M), or a potent protein kinase A inhibitor, KT5720 (500 nM). In contrast, a membrane-permeable cAMP analog, 8Br-cAMP (100 M), induced a twofold increase in channel activity. The addition of a catalytic subunit of protein kinase A (PKA-CS, 100 U/ml) to the bath in inside-out patches stimulated channel activity in the presence of 1 mM ATP. Furthermore, the channel activity maintained with 1 mM ATP in inside-out patches was suppressed by internal acidification and enhanced by alkalization. These results suggest that the activity of the inwardly rectifying K⁺ channel in cultured human proximal tubule cells was ATP-dependent and regulated at least in part by cAMP/PKA-mediated phosphorylation processes and intracellular pH.

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