Inhibition of mTORc1 in the distal-convoluted-tubule (DCT) augments renal K ⁺ -excretion and decreases plasma K ⁺ concentration (hypokalemia)

Aim: Mechanistic-target-of-rapamycin-complex-1 (mTORc1) plays a role in maintaining K + homeostasis. Rapamycin, an inhibitor of mTORc1 which is approved for treatment of cancer and for use in preventing rejection after organ transplantation, has been shown to decrease plasma K + concentrations (hypokalemia). Hypothesis: We now test the hypothesis that mTORc1 of the DCT regulates Kir4.1/Kir5.1 channels and thiazide-sensitive-Na-Cl cotransporter, which play a role in regulating renal K + excretion. Method: We used patch-clamp-technique to examine basolateral Kir4.1/Kir5.1 in early-DCT, performed immunoblotting to examine NCC expression and in vivo measurement of urinary K + -excretion (E K ) to determine baseline renal-E K. The experiments were conducted in Fkbp1a flox/flox mice (control), kidney-specific FKBP12-KO (Ks-FKBP12-KO) mice treated with rapamycin and DCT-specific Rapamycin-sensitive-companion- of-mTOR knockout (DCT-RAPTOR-KO) mice. Results: Application of rapamycin decreased Kir4.1/Kir5.1-mediated K + currents and depolarized DCT-membrane-potential in Fkbp1a flox/flox mice. However, the effect of rapamycin on Kir4.1/Kir5.1 was absent in Ks-FKBP12-KO. Rapamycin decreased basolateral 40-pS K + -channel activity (Kir4.1/Kir5.1 heterotetramer) of the DCT. This effect was absent in the DCT treated with H 2 O 2 which stimulated the 40-pS K + -channel activity, suggesting the role of reactive-oxygen-species (ROS) in mediating the effect of mTORc1 on Kir4.1/Kir5.1. Rapamycin treatment significantly decreased the expression of both phosphorylated-NCC and total-NCC in Fkbp1a flox/flox mice but not in Ks-FKBP12-KO mice. Moreover, in vivo measurement of urinary-Na + and urinary-K + demonstrated that rapamycin treatment decreased hydrochlorothiazide-induced natriuresis but increased renal-K + -excretion in Fkbp1a flox/flox mice. Moreover, Kir4.1/Kir5.1-mediated K + currents of the DCT were lower and DCT membrane potential was less negative in DCT-specific RAPTOR-conditional knockout mice (DCT-RAPTOR-KO) than those of Ncc-Cre-Raptor flox/flox mice (control). Also, the expression of phosphorylated-NCC was lower in DCT-RAPTOR-KO mice than Ncc-Cre-Raptor flox/flox mice. In contrast, the expression of phosphorylated-NKCC2 was the same between two genotypes while the expression of aENaC and bENaC was higher in DCT-RAPTOR-KO mice than Ncc-Cre-Raptor flox/flox mice. Consequently, DCT-RAPTOR-KO mice had a higher urinary K + excretion and lower plasma K + concentrations than Ncc-Cre-Raptor flox/flox . Summary: The inhibition of mTORc1 or deletion of RAPTOR in the DCT inhibits the basolateral Kir4.1/Kir5.1 activity, decreases NCC expression/activity and increases renal K + excretion. Conclusion: mTORc1 in the DCT plays a significant role in maintaining K + homeostasis by controlling the basolateral Kir4.1/Kir5.1 of the DCT and NCC. NIH grants DK136491 and DK133220 This abstract was presented at the American Physiology Summit 2025 and is only available in HTML format. There is no downloadable file or PDF version. The Physiology editorial board was not involved in the peer review process.