Critical role of the mineralocorticoid receptor in aldosterone-dependent and aldosterone-independent regulation of ENaC in the distal nephron

Abstract

The epithelial sodium channel (ENaC) constitutes the rate-limiting step for sodium absorption in the aldosterone-sensitive distal nephron (ASDN) comprising the late distal convoluted tubule (DCT2), the connecting tubule (CNT) and the collecting duct. Previously, we demonstrated that ENaC activity in the DCT2/CNT transition zone is constitutively high and independent of aldosterone, in contrast to its aldosterone dependence in the late CNT and initial cortical collecting duct (CNT/CCD). The mineralocorticoid receptor (MR) is expressed in the entire ASDN. Its activation by glucocorticoids is prevented through 11β-hydroxysteroid dehydrogenase 2 (11β-HSD2) abundantly expressed in the late but probably not the early part of ASDN. We hypothesized that ENaC function in the early part of the ASDN is aldosterone-independent but may depend on MR activated by glucocorticoids due to low 11β-HSD2 abundance. To test this hypothesis, we used doxycycline-inducible nephron-specific MR-deficient mice (MR KO). Whole-cell ENaC currents were investigated in isolated nephron fragments from DCT2/CNT or CNT/CCD transition zones using the patch-clamp technique. ENaC activity was detectable in CNT/CCD of control mice but absent or barely detectable in the majority of CNT/CCD preparations from MR KO mice. Importantly, ENaC currents in DCT2/CNT were greatly reduced in MR KO mice compared to ENaC currents in DCT2/CNT of control mice. Immunofluorescence for 11β-HSD2 was abundant in CCD, less prominent in CNT and very low in DCT2. We conclude that MR is critically important for maintaining aldosterone-independent ENaC activity in DCT2/CNT. Aldosterone-independent MR activation is probably mediated by glucocorticoids due to low expression of 11β-HSD2.

Mineralocorticoid Receptor Antagonists Cause Natriuresis in the Absence of Aldosterone

“…17,21 The data also show that both ENaC and NCC are more severely reduced in the KS-MR −/− mice than in AS −/− mice; this likely reflects direct (ENaC) and indirect (NCC) effects, as we and others have shown that both ENaC and NCC must be reduced for the full salt-wasting phenotype. 17,21,32 Together with results showing that amiloride-sensitive sodium currents along the DCT2/CNT are similar in control and AS −/− mice (see Figure 4 and the study by Nesterov et al 10 ), the results suggest an essential homeostatic role for non- aldosterone MR activation in the kidney. If this is true, then mice lacking aldosterone should still respond to MRA with natriuresis.…”

“…In support of this, we found differences between nuclear MR intensity along the CCD of control versus AS −/− mice. Nevertheless, the current data indicate that factors other than nuclear trafficking of MRs are required for aldosterone to stimulate ENaC, as ENaC activity along the CCD is highly dependent on aldosterone concentration (see Figure 4 and the studies by Nesterov et al 10,11 and Wu et al 12 ). In contrast, no difference in nuclear trafficking between control and AS −/− mice was noted along the DCT2/CNT, which exhibits aldosterone-independent ENaC activity.…”

“…The canonical renal target for aldosterone is the ASDN, where the effects derive predominantly from activating ENaC. Yet, ENaC activity along the DCT2 and early CNT is largely independent of circulating aldosterone, 11 even though it is reduced in mice lacking MR. 10,12 One possible mechanism for aldosterone-independent MR activation is through Rac1 (encoded by Rac1 ), which causes paradoxical activation in salt-sensitive models. 26 Yet, the C57/BL6 mice we tested here are relatively salt resistant, making this an unlikely explanation.…”

“…RNA sequencing data from Chen et al showed that Nr3c2 (encoding MR) is expressed along distal nephron, and its expression along CNT/CCD is much higher than along DCT (Figure S2), 16 but MR expression levels along DCT2, in which ENaC current is higher than in CCD, [10][11][12] have not been as clear. To address this, immunofluorescence was performed on kidneys from control and kidney-specific MR knockout (KS-MR −/− ) mice.…”

“…ENaC is expressed throughout this segment, but Nesterov et al showed that activity along the DCT2/CNT is independent on aldosterone, whereas it is aldosterone dependent along the CD. 10–12 This differential activation profile correlates inversely with 11βHSD2, which is minimally expressed along the DCT2 but increases along the CNT and CD, 10,13–15 suggesting that aldosterone-independent MR activity may be mediated by glucocorticoid binding to MR. Here, we tested whether MR activity in the absence of aldosterone maintains ENaC activity in the kidney and whether this is responsible for phenotypic differences between mice lacking aldosterone and mice lacking MR.…”