Aldosterone (aldo) is a steroid hormone that has a key role in extracellular volume and K+ homeostasis. While aldo levels are expected to be suppressed with high Na+ intake, 5-10% of hypertensives have chronic hyperaldosteronism regardless of Na+ intake. This combination of high sodium and high aldo can worsen end organ damage, especially in the kidney. We examined the effects of elevated aldo in the absence or presence of a high salt diet (HSD) on the expression of Na+ transporters in the distal nephron and on distal nephron remodeling to understand if these changes may play a role in the altered pathology. Male C57Bl/6 mice received either a control (0.4% NaCl) or HSD (4 weeks, 8% NaCl), +/- aldo via minipump for the final 2 weeks of treatment (240 μg/kg/day). Transporter expression levels and localization were analyzed by Western blot, qPCR and immunofluorescent staining. Expression of the α subunit of the epithelial sodium channel (ENaC) was increased with aldosterone regardless of Na+ intake, while subunits increased at both a transcript and protein level in mice receiving aldo and HSD. Both total levels of sodium chloride cotransporter (NCC) and active pNCC significantly increased with aldo alone, in association with a fall in blood [K+]. However, the addition of HSD markedly attenuated this increase in NCC and pNCC, despite more severe hypokalemia. This change was not associated with a difference in the expression of its upstream kinase SPAK. NKCC2 expression was not significantly altered in any of the conditions. We imaged whole tubules in thick kidney slices (300μm) to assess whether remodeling of the distal tubule could account for NCC expression differences. Sections were incubated with antibodies against pNCC and AQP2 and cleared with ethyl cinnamate to allow for imaging and analysis of whole distal tubules. Length of the distal tubule was analyzed as the pNCC positive portion. While aldo has previously been shown to promote hypertrophy of the distal tubule, we found a significant shortening of tubules in animals treated chronically with 2 weeks of aldo (Ctl 434m ± 63μm versus aldo 375m ± 88m, Plt;0.05, n=38 and 30 respectively). However, the addition of HSD with aldo caused a significant increase in distal tubule length (486m ±68μm, Plt;0.05, n=23). HSD by itself did not alter distal tubule length (428m ± 55m, n=17). Our results suggest that aldo-mediated transcriptional regulation of ENaC subunits in the kidney changes according to dietary Na+ status. The attenuation of increased NCC and pNCC expression in kidneys of mice receiving HSD with aldo, despite profound hypokalemia and an increase in DCT length, suggest that factors other than blood [K+] have important roles in regulating NCC expression. Finally, aldo-dependent DCT remodeling is dependent on dietary Na+.
This work was supported by 5R01HL147818-23 and DK007052
