Role of Estrogen Receptors in a Model of Dilutional Hyponatremia

Hyponatremia is the most frequently occurring electrolyte disorder encountered and is an independent risk factor for increased mortality in liver failure patients. Dilutional hyponatremia seen in liver failure is due to inappropriate arginine vasopressin (AVP) release and can be studied using animal model of bile duct ligation (BDL). Our previous studies to address the knowledge gap of sex differences in this model showed that unlike male BDL rats, female and ovariectomized (OVX) BDL rats did not develop hyponatremia, supraoptic AVP neuron activation, or increased copeptin (CPP), a surrogate to measure AVP, secretion compared to sham ligated females. Interestingly, we also found an increased adrenal estradiol (E2) concentration that could explain the increased plasma E2 observed in OVX BDL rats. An intracerebroventricular infusion of estrogen receptor (ER) antagonist, ICI 182,780 (ICI) in female BDL rats increased CPP concentration compared to BDL‐vehicle and sham groups. These data suggest ER involvement in the prevention of BDL‐induced hyponatremia in female rats. However, ICI is also a G protein‐coupled estrogen receptor 1 (GPER) agonist at high concentrations. We tested GPER expression within the hypothalamo‐neurohypophyseal system of female rats to further elucidate the role of ER in this model. Adult female Sprague‐Dawley rats (n=6) were anesthetized with inactin (100 mg/kg ip) and transcardially perfused with 4% paraformaldehyde and brains harvested. Double‐immunostaining was performed on three separate sets of coronal sections (40μm thick) of forebrain containing PVN and SON. All sets were processed for GPER using polyclonal rabbit anti‐GPER (1:1000, Abcam) and Cy3 donkey anti‐rabbit (1:500, Jackson ImmunoResearch). First set: GPER and AVP, using polyclonal guinea pig anti‐AVP (1:10000, Peninsula Laboratories) and Cy2 donkey anti‐guinea pig (1:500, Jackson ImmunoResearch). Second set: GPER and oxytocin (OXY), using monoclonal mouse anti‐oxytocin (1:5000, Millipore) and Cy2 donkey anti‐mouse (1:500, Jackson ImmunoResearch). Third set: GPER and glial fibrillary acidic protein (GFAP), using monoclonal mouse anti‐GFAP (1:1000, Millipore) and Cy2 donkey anti‐mouse (1:500, Jackson ImmunoResearch). Images of the sections were captured using an Olympus BX41 fluorescence microscope. Co‐localization of GPER+AVP and GPER+OXY was observed in a subpopulation of neurons in the PVN and SON of all rats. Unilateral SON counts for co‐localization: GPER+AVP, 64.8% and GPER+OXY, 64.0%. GPER+GFAP co‐localization was not observed in either region of interest. To conclude, GPER is expressed on a subset of AVP and OXY neurons and not astrocytes in the hypothalamo‐neurohypophyseal system of female rats. Future studies combining GPER antagonist and ICI in this model of hyponatremia will provide further insight toward mechanisms underlying sex differences in hyponatremia related to liver failure. Support or Funding Information NHLBI R01 HL142341, NIH 2 T32 AG020494‐16A1