Potassium Can Regulate Its Own Excretion in Collecting Duct Cells: Role of mTORC2, SGK1 and ENaC

Abstract

BackgroundThe control of potassium (K+) excretion is central to the homeostatic response to variation in environmental K+ in all multicellular organisms. In mammals, the serine‐threonine kinase SGK1 plays a central role in regulating K+ excretion by stimulating electrogenic Na+ transport in the kidney tubules through the epithelial Na+ channel, ENaC. The resulting increase in lumen negative electrical potential enhances the driving force for K+ secretion through apical membrane K+ channels. It is well established that SGK1 phosphorylation state and activity are controlled by the kinase mTORC2, however, the central physiological inputs that control this phosphorylation are poorly characterized. We postulated that K+ itself might act directly in kidney tubule cells to regulate SGK1 through effects on mTORC2 activity.MethodsmpkCCD cortical collecting duct (CCD) cells were grown on Transwell filters, and treated for 4 h with aldosterone to increase SGK1 expression and adapted to either 1 mM or 5 mM [K+]. At t = 0, basolateral [K+] was changed from 1 to 5 mM or from 5 mM to 1 mM, followed by a return to 5 mM in the presence or absence of either an SGK1 (GSK650394) or mTOR inhibitor. Additional experiments were performed using patch clamp to detect ENaC currents in CCD isolated from mice and subjected to acute change in bath [K+] from 5 mM to 1 mM. SGK1 phosphorylation state in response to changes in [K+] was also assessed in wild type and WNK1 deficient HEK‐293 cells.ResultsIn mpkCCD cells, raising basolateral [K+] from 1 to 5 mM increased (approximately 3‐fold), while lowering [K+] from 5 mM to 1 mM decreased (approximately 4.5‐fold) SGK1 phosphorylation (p < 0.01 for both manipulations). Changing apical [K+] had no significant effect. Further, shifting basolateral [K+] from 5 mM to 1 mM markedly reduced ENaC‐dependent Na+ current, and returning [K+] to 5 mM induced a rapid increase in Na+ current. The stimulatory effects on both SGK1 phosphorylation and Na+ current were dependent on mTOR and SGK1 in that they were inhibited by the mTOR inhibitor, AZD 8055 and the SGK1 inhibitor, GSK650394. SGK1 phosphorylation was also strongly stimulated in a graded fashion by raising medium [K+] in HEK‐293 cells. Interestingly, the K+ stimulatory effect on SGK1 was lost in WNK1‐deficient HEK‐293 cells. Finally, in patch clamp performed on cortical collecting duct isolated from mice, ENaC currents were significantly greater in the presence of 5 mM (330 +/−25 pA, N=5) than in the presence of 1 mM (250 +/− 20 pA, N=5) [K+] (p < 0.05).ConclusionThese data strongly support the idea that changes in extracellular [K+] act directly in CCD cells to modulate mTORC2‐dependent SGK1 phosphorylation resulting in altered ENaC‐mediated Na+ transport, and thereby stimulating K+ secretion. This effect appears to be highly dependent on WNK1. In light of recent evidence that Na‐Cl cotransporter (NCC) is directly regulated in distal convoluted tubule cells by extracellular fluid [K+], these data support a new model of coordinated regulation of Na+ transport between distal convoluted tubule and CCD, which is directly modulated by local renal [K+].Support or Funding InformationAm Heart Assoc. 17GRNT33410667;NIH/NIDDK, R01‐DK56695This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.