Lack of Kir4.1 in the Distal Convoluted Tubule Causes ENaC Hyperactivity During K⁺ Restriction Leading to Hypokalemia

Loss-of-function mutations in KCNJ10, encoding Kir4.1, cause EAST/SeSAME syndrome, with renal salt-wasting tubulopathy and hypokalemia. We hypothesized that Kir4.1 deletion specifically in the distal convoluted tubule (DCT) stimulates ENaC activity via the mammalian target of rapamycin (mTOR)-dependent mechanisms, contributing to hypokalemia.

Metabolic cages, electrophysiology, immunoblotting, immunostaining, and in vivo diuretic response experiments were used to examine biochemical parameters, Kir4.1/Kir5.1 activity, NCC and ENaC function in the DCT-specific Kir4.1 knockout (DCT-Kir4.1 KO) mice under normal or K⁺ restriction conditions.

DCT-Kir4.1 KO mice exhibited impaired basolateral K⁺ channel and NCC activity, enhanced ENaC activity, and mild hypokalemia. Amiloride treatment induced similar natriuresis and kaliuresis in DCT-Kir4.1 KO and kidney-specific Kir4.1 KO mice, but had minimal effects in collecting system Kir4.1 KO mice, suggesting high ENaC activity following Kir4.1 deletion in the DCT. Notably, severe hypokalemia, along with upregulated ENaC expression and activity, was observed in DCT-Kir4.1 KO mice under dietary K⁺ restriction. Patch-clamp experiments further revealed elevated ENaC currents in the DCT2 of KO mice on a low-K⁺ diet, independent of aldosterone levels. Inhibition of mTOR with AZD8055 reduced SGK1/Nedd4-2 phosphorylation, cleaved α-ENaC expression, and DCT2 ENaC currents, suggesting a role for mTOR in ENaC hyperactivity in K⁺-restricted DCT-Kir4.1 KO mice. This notion was also supported by the upregulated Rictor expression observed in the isolated DCT of these KO mice.

We conclude that Kir4.1 deletion drives ENaC hyperactivity in the DCT via the mTORC2-dependent SGK1/Nedd4-2 signaling pathway, promoting low potassium diet-induced hypokalemia.