Sex, dietary salt and AT2R impact sodium transporters along the nephron
Authors: Lei Hillmin Andy , Mirabito Collafella Katrina M. , Ralph Donna Lee , Denton Kate M. , McDonough Alicia Ann 
Published: June 2021

BackgroundThe Denton lab has reported that females (F) express 4‐fold more angiotensin type 2 receptor (AT2R) than males (M), that the pressure natriuresis (P‐Nat) is activated at lower blood pressures (BP) in F vs. M wildtype (WT) mice and that this advantage is lost in F AT2R KO (KO) mice. Additionally, 7 day 5% NaCl high salt diet (HS) increased BP in F WT and AT2R‐KO, but not in M mice.AimTest the hypothesis that sexual dimorphism in P‐Nat and response to HS are due to distinct renal Na+ transporter profiles and to AT2R expression.MethodsFVB/N WT mice and AT2R‐KO mice of both sexes (n=6) were fed LS (0.02% NaCl), NS (0.24% NaCl) and HS (5% NaCl) diets. 24 hr urine was collected (day 6); kidneys were quick frozen (day 7), dissected into cortex ‘c’ and medulla ‘m,’ and homogenized for quantitative immunoblotting of: Na+/H+ exchanger 3 (NHE3), Na‐HCO3−cotransporter (NBCe1), claudin 2 (cldn2), Na‐K‐2Cl cotransporter 2 (NKCC2) and phosphorylated form (p), Na,K‐ATPase α1, Na‐Cl cotransporter (NCC, NCCp), cldn7, cotransporter kinases (SPAK and OSR1), epithelial sodium transporter (ENaC) α,β, and γ subunits and cleaved forms (‐cl), and aquaporins (AQP1, AQP2).ResultsComparing KO to WT at baseline, only m‐NKCCp and m‐AQP2 had lower abundance (by 20%) in both KO‐F and KO‐M. During LS, NS, HS diets, both genotypes maintained salt balance and similar body weights within sexes.Proximal tubule (PT)NHE3, cldn2, and NBCe1 were 30% less abundant and AQP1 40% more abundant at baseline in WT‐F and KO‐F vs WT‐M and KO‐M. LS diet paradoxically depressed NHE3 in WT‐M, WT‐F, KO‐M. In HS diet, NHE3 decreased in WT‐M and KO‐M.Loop of HenleNKCC2 and Na,K‐ATPase were 20–30% more abundant at baseline in WT‐F and KO‐F vs WT‐M and WT‐ F. HS decreased m‐NHE3 in WT‐M and KO‐F.Distal tubule (DT)NCC, NCCp, SPAK, SPAKp, OSR1p, and cldn7 were all ~ 50% more abundant in WT‐F and KO‐F vs WT‐M and KO‐M. LS diet increased NCC, NCCp in WT‐M and KO‐F; HS diet depressed NCC, NCCp in WT‐F and KO‐F (25 and 40%), c‐NKCC2 was not altered by LS or HS.Collecting ductα‐ENaC‐cl and γ‐ENaC‐cl were 50% more abundant in WT‐F and KO‐F vs. WT‐M and KO‐M. LS diet increased α‐ENaC‐cl across groups. HS diet decreased α‐ENaC‐cl in WT‐F and KO‐F (40 and 50%), but not in WT‐M or KO‐M. HS diet increased AQP2 abundance across groups and increased m‐Na,K‐ATPase in KO‐M and KO‐F (~25%).Conclusions1) Lower PT and higher DT transporters' abundance in F vs. M is not a function of the higher AT2R in WT‐F vs. M. 2) Overall, the depression of sodium transporters along the nephron (NHE3, NCC, ENaC‐cl) during HS diets is amplified in female KO vs. WT indicating that AT2R expression does not facilitate natriuresis via depression of transporter abundance. 3) The shift in P‐Nat to lower BP in F can be attributed to lower baseline PT transporters' abundance. 4) Since WT‐F have 4‐fold higher AT2R expression than WT‐M, the responses to diet salt observed only in WT‐M and KO‐F (m‐NHE3, NCC and m‐Na,K‐ATPase) may reflect similarly low AT2R levels. 5) Baseline AT2R levels are associated with higher medullary NKCC2 and AQP2 abundance which may impact concentration potential.Support or Funding InformationDK083785This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.