Kir4.1/Kir5.1 of distal convoluted tubule is required for short-term angiotensin-II-induced stimulation of Na-Cl cotransporter

Angiotensin-II (Ang-II) perfusion stimulates inwardly-rectifying potassium channels 4.1 and 5.1 (Kir4.1/Kir5.1) in distal convoluted tubule (DCT) and thiazide-sensitive Na-Cl cotransporter (NCC). The aim of the present study is to explore the role of Kir4.1/Kir5.1 in mediating the effect of Ang-II on NCC. We used immunoblotting and patch-clamp experiments to examine the effect of 1- or 7-day Ang-II perfusion on basolateral Kir4.1/Kir5.1 in the DCT and NCC using kidney-tubule-specific (Ks) angiotensin II type 1a receptor (AT1Ar)-knockout (KO), Ks-Kir4.1-knockout and the corresponding wild-type mice. Ang-II perfusion for 1 and 7 days increased phospho-NCC (pNCC) and total-NCC (tNCC) expression and the effect of Ang-II perfusion on pNCC and tNCC was abolished in Ks-AT1aR-KO. Ang-II perfusion for 1 day robustly stimulates Kir4.1/Kir5.1 in the late DCT (DCT2) and to a lesser degree in the early DCT (DCT1), an effect was absent in Ks-AT1aR-KO mice. However, Ang-II perfusion for 7 days did not further stimulate Kir4.1/Kir5.1 in the DCT2 and only modestly increased Kir4.1/Kir5.1-mediated K⁺ currents in DCT1. Deletion of Kir4.1 not only significantly decreased the expression of pNCC and tNCC, but also abolished the effect of 1-day Ang-II perfusion on the expression of phospho-with-no-lysine kinase-4 (pWNK4), phospho-ste-20-proline-alanine-rich kinase (pSPAK), Pncc, and tNCC. However, 7-day Ang-II perfusion was still able to significantly stimulate the expression of pSPAK, pWNK4, pNCC, and tNCC, and increased thiazide-induced natriuresis in Ks-Kir4.1-KO mice without obvious changes in K⁺ channel activity in the DCT. We conclude that short-term Ang-II-induced stimulation of pWNK4, pSPAK, and pNCC depends on Kir4.1/Kir5.1 activity. However, long-term Ang-II is able to directly stimulate pWNK4, pSPAK, and pNCC by a Kir4.1/Kir5.1 independent mechanism.<b>NEW & NOTEWORTHY</b> We investigated the role of Kir4.1/Kir5.1 in mediating the effect of short-term Ang-II on Na-Cl cotransporter (NCC) expression/activity. We demonstrated that Kir4.1/Kir5.1 in the distal convoluted tubule is required for short-term Ang-II-induced stimulation of with-no-lysine-kinase 4 (WNK4), ste20-proline-alanine-rich kinase (SPAK), and NCC. However, sustained Ang-II stimulation is expected to activate WNK4, SPAK, and NCC by Kir4.1/Kir5.1-independent mechanism.