mTORc2 in Distal Convoluted Tubule and Renal K+ Excretion during High Dietary K+ Intake

Abstract

Key Points High K stimulates mechanistic target of rapamycin complex 2 (mTORc2) in the distal convoluted tubule (DCT).Inhibition of mTORc2 decreased the basolateral Kir4.1/Kir5.1 and Na-Cl cotransporter in the DCT.Inhibition of mTORc2 of the DCT compromised kidneys' ability to excrete potassium during high K intake. Background Renal mechanistic target of rapamycin complex 2 (mTORc2) plays a role in regulating renal K+ excretion (renal-EK) and K+ homeostasis. Inhibition of renal mTORc2 causes hyperkalemia due to suppressing epithelial Na+ channel and renal outer medullary K+ (Kir1.1) in the collecting duct. We now explore whether mTORc2 of distal convoluted tubules (DCTs) regulates basolateral Kir4.1/Kir5.1, Na-Cl cotransporter (NCC), and renal-EK. Methods We used patch-clamp technique to examine basolateral Kir4.1/Kir5.1 in early DCT, immunoblotting, and immunofluorescence to examine NCC expression and in vivo measurement of urinary K+ excretion to determine baseline renal-EK in mice treated with an mTORc2 inhibitor and in DCT-specific rapamycin-insensitive companion of mTOR knockout (DCT-RICTOR-KO) mice. Results Inhibition of mTORc2 with AZD8055 abolished high-K+–induced inhibition of Kir4.1/Kir5.1 in DCT, high potassium–induced depolarization of the DCT membrane, and high potassium–induced suppression of phosphorylated Na-Cl cotransporter (pNCC) expression. AZD8055 stimulated the 40-pS inwardly rectifying K+ channel (Kir4.1/Kir5.1-heterotetramer) in early DCT in the mice on overnight high potassium intake; this effect was absent in the presence of protein kinase C inhibitors, which also stimulated Kir4.1/Kir5.1. AZD8055 treatment decreased renal-EK in animals on overnight high-potassium diet. Deletion of RICTOR in the DCT increased the Kir4.1/Kir5.1-mediated K+ currents, hyperpolarized the DCT membrane, and increased the expression of pWNK4 and pNCC. Renal-EK was lower and plasma K+ was higher in DCT-RICTOR-KO mice than corresponding control mice. In addition, overnight high-potassium diet did not inhibit Kir4.1/Kir5.1 activity in the DCT and failed to inhibit the expression of pNCC in DCT-RICTOR-KO mice. Overnight high potassium intake stimulated renal-EK in control mice, but this effect was attenuated in DCT-RICTOR-KO mice. Thus, overnight high potassium intake induced hyperkalemia in DCT-RICTOR-KO mice but not in control mice. Conclusions mTORc2 of the DCT inhibits Kir4.1/Kir5.1 activity and NCC expression and stimulates renal-EK during high potassium intake.