Modulation of NHE3 Activity in Rat Renal Proximal Tubules during the Estrous Cycle Phases and Direct Impact of Estradiol and Progesterone

Abstract

Estradiol (E2) and progesterone (P4) are two of the major steroid hormones involved in the control of reproductive functions in females. Among other effects, E2 and P4 can modulate sodium reabsorption along the nephron, altering the hydroelectrolytic balance, leading to hydrosaline retention. Na/H exchanger, isoform 3, is the main route for sodium bicarbonate reabsorption in renal proximal tubules (PT). The aims of the present study are to evaluate the activity of NHE3 in the different estrous cycle phases and to investigate the direct effects of E2 and P4 on NHE3 activity in kidneys of control (sham) or ovariectomized rats (OVX). All the protocols were approved by the Committee on Animal Research and Ethics from the Ceara State University (protocol #2722214/2016). The estrous cycle of female Wistar rats were evaluated during 15 days and animals randomly assigned to OVX or sham group. Thereafter, two weeks after surgery, the activity of NHE3 was evaluated in vivo by using the stationary microperfusion (SM) technique and after this procedure, the kidneys were stored for evaluation of NHE3 protein abundance by Western Blot (WB). In a first set of experiments, PT were punctured to perfuse control solution (PCS) in order to compare the different phases of the estrous cycle against OVX‐group. In a second set, PTs were perfused with E2 or P4 to compare with OVX animals. SM allows the measurement of H+ secretion in proximal tubules, representing an indirect measure of NHE3 function. It was observed that OVX presented lower H+ secretory activity than all cyclic groups and the estrous group presented higher secretion rates: [JHCO3‐ nmol.cm‐2.s‐1 OVX 1.74 ± 0.07 (10); Proestrus 1.90 ± 0.12 (12)a; Estrus 2.35 ± 0.08 (13)ab; Metaestrus 1.97 ± 0.12 (15)ac and Diestrus 1.89 ± 0.08 (14)ac, where a p <0.05 vs OVX, b p <0.05 vs Proestrus and c p <0.05 vs Estrus]. In addition, an increase in proximal H+ secretion/bicarbonate reabsorption was observed in all E2 and P4 concentrations when administered intratubularly and compared to the OVX group: [JHCO3‐, nmol.cm‐2s‐ OVX+Vehicle 1.84 ± 0.06 (11); E2 (0.1 nM) 2.18 ± 0.07 (15)a; E2 (1 nM) 2.24 ± 0.08 (11)a; E2 (10 nM) 2.20 ± 0.09 (12)a; P4 (0.1 nM), 2.18 ± 0.11 (13)a, P4 (10 nM), 2.10 ± 0.07 (10)a, where a p <0.05 vs OVX + Vehicle]. NHE3 total/villin expression in brush border membranes of PT is lower in the Diestrus group when compared to all other groups: OVX 0.79 ± 0.26 (n=5); Proestrus 0.64 ± 0.08 (n=6); Estrus 0.76 ± 0.30 (n=6); Metaestrus 0.69 ± 0.12 (n=6); Diestrus 0.22 ± 0.12 (n=6)*, p <0.05 vs OVX, Proestrus, Estrus and Metaestrus. Significative changes in NHE3 activity occurs in the different phases of estrous cycles with higher activity in the estrus and lower in diestrus. In addition, it was observed a positive regulation of NHE3 by E2 and P4 in PT which may contribute to sodium retention if this stimulus is maintained for a long‐term.Support or Funding InformationCNPq/FUNCAPThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.