Supraphysiological level of estrogen exposure in vivo increases lymphoid cell death in mice

Estrogen can enhance or reduce lymphocyte functions in vitro depending on dose and exposure duration. The purpose of this study was to determine the effect of in vivo 17 beta-estradiol (E2) on apoptosis and necrosis in lymphoid tissue of female C567BL/6 mice. Animals were ovariectomized (OVX), ovariectomized and 17 beta-estradiol supplemented (OVX + E2; 71 micrograms E2 per day for 14 days), sham ovariectomized (SHAM), or unhandled controls (CONTROL). Thymus and spleen were removed aseptically, cells dispersed into single cell suspensions in RPMI-1640, and measures of cell damage performed: an annexin V flow cytometric assay for markers of apoptosis and an enzyme-linked immunoassay for measures of DNA fragmentation and necrosis. OVX + E2 mice had 620 +/- 72 pg/ml 17 beta-estradiol in serum in contrast to OVX mice which had 7.6 +/- 5 pg/ml, the SHAM mice which had 2.8 +/- 1 pg/ml of serum E2, and the CONTROL mice which had 3.9 +/- 0.8 pg/ml of serum E2 (p < 0.001). There was a significantly lower percentage of viable thymocytes in OVX + E2 mice compared to the other treatment conditions (p < 0.001, respectively). There was also a significantly higher percentage of annexin V positive thymocytes in OVX + E2 mice (p < 0.005). Measures of DNA fragmentation by ELISA were higher in splenocytes from OVX + E2 mice than in the OVX, SHAM or CONTROL mice (p < 0.005). These results suggest that supraphysiological levels of estrogen in vivo induce damage in lymphoid cells; however, the impact of estrogen associated lymphoid tissue damage on specific immune functions remains to be determined.     

Regulation of leptin by steroid hormones in rat adipose tissue.

We investigated if steroid hormones regulate the secretion and the expression of leptin in female and male rat adipose tissue fragments in vitro. Dexamethasone time and dose-dependently increased the secretion and mRNA expression of leptin with a half-maximal stimulation of approximately 1 nM. A time-course revealed a maximal stimulatory effect of 17 beta-estradiol after 24 hours. In male adipose tissue 17 beta-estradiol increased leptin secretion (32% by 50 nM 17 beta-estradiol, P = 0.07 and 34% by 500 nM 17 beta-estradiol, P < 1780.05) after 24 hours. An additional effect of estrogen was seen in the dexamethasone (50 nM) stimulated cells (38% with 50 nM 17 beta-estradiol, P < 0.05 and 48% by 500 nM 17 beta-estradiol, P < 0.05). Basal secretion of leptin was equal in female and male adipose tissue, whereas the effects of 17 beta-estradiol (50 nM) and dexamethasone were significantly increased in female as compared with male adipose tissue. Progesterone, testosterone, dihydrotestosterone and dehydroepiandrostendione-sulfate neither affected leptin secretion in male nor female adipose tissue in vitro. Furthermore, to investigate the effect of estrogen female rats were ovariectomized (OVX) and the adipose tissue was incubated in vitro and compared with adipose tissue leptin secretion from sham operated rats (SHAM), and with ovariectomized rats treated with 17 beta-estradiol (EST). A decreased basal and dexamethasone-stimulated leptin secretion from OVX rats compared with SHAM rats was found (P < 0.005) whereas 17 beta-estradiol treatment of ovariectomized rats maintained a normal leptin secretion. However, the dexamethasone stimulation was equally increased above basal levels in SHAM, OVX and EST rats (3.7 +/- 1.2, 2.9 +/- 0.8, 4.2 +/- 1.4, NS, ANOVA) respectively.     

Effect of genistein on the expression of bone metabolism genes in ovariectomized mice using a cDNA microarray

Osteoporosis associated with estrogen deficiency is defined as an abnormal decrease in bone mass leading to an increased fracture risk. Genistein (GEN), as a phytoestrogen, is a type of soybean-derived isoflavone that possesses structural similarity to estrogen. In this study, we assessed the effect of GEN in ovariectomized (OVX) mice. To determine the effect of GEN on bone metabolism, we investigated gene expression profiles using a radioactive cDNA microarray. Eight-week-old female mice were either sham operated (SHAM) or OVX. From 1 week after the operation, OVX mice were injected daily with intraperitoneal GEN (0.1, 0.5, 1.5 and 3.0 mg/day) or 17beta-estradiol (E2, 0.03 microg/day) for 4 weeks. A cDNA microarray was used to evaluate changes in the expression of 1,152 genes. OVX mice showed bone mineral density (BMD) loss versus SHAM mice (5.8+/-0.4 vs. 6.9+/-0.6 mg/cm2). However, femur BMDs were completely restored by GEN and by E2 administration in OVX mice. Serum osteocalcin in OVX mice treated with 0.5 mg/day of GEN was 1.6-fold (44.30+/-5.73 ng/ml) higher than that in untreated mice. GEN treatment up-regulated 38 genes (e.g., mitogen-activated protein kinase 10) and down-regulated 18 (e.g., matrix metalloproteinase 13). Moreover, GEN was found to have a protective effect on bone loss caused by estrogen deficiency in OVX mice. The present study suggests that GEN modulates bone metabolism-related gene expression, including calciotropic receptor, cytokines, growth factors and bone matrix proteins.     

17beta-estradiol downregulates tissue angiotensin-converting enzyme and ANG II type 1 receptor in female rats

Estrogens have been implicated in both worsening and protecting from cardiovascular disease. The effects of 17beta-estradiol (E2) on the cardiovascular system may be mediated, at least in part, by its modulation of local tissue renin-angiotensin systems (RAS). We assessed two critical components, angiotensin-converting enzyme (ACE) and ANG II type 1 receptor (AT(1)R), in the heart, lung, abdominal aorta, adrenal, kidney, and brain in four groups of female Wistar rats (n = 5-6/group): 1) sham ovariectomized, 2) ovariectomized (OVX) treated with subcutaneous vehicle, 3) OVX treated with 25 mug/day (regular) E2 subcutaneously, and 4) OVX treated with 250 mug/day (high) subcutaneous E2 for 2 or 5 wk. After 2 wk, plasma ACE activity was not altered by OVX, but it was 34-38% lower in OVX + regular E2 and OVX + high E2 rats compared with sham OVX rats, and these decreases were no longer present after 5 wk. After 5 wk, OVX alone increased ACE activity and binding densities, and AT(1)R binding densities by 15-100% in right ventricle, left ventricle (LV), kidney, lung, abdominal aorta, adrenal and several cardiovascular regulatory nuclei in the brain. These effects were, for the most part, prevented by regular E2 replacement and were reversed to decreases by high E2 treatment. This regulation of tissue ACE and AT(1)R is significant as the activity of these tissue RAS contributes to the pathogenesis and/or progression of hypertension, atherosclerosis, and LV remodeling after myocardial infarction.     

Production of endothelin-1 from the mesenteric arteries of streptozotocin-induced diabetic rats

Release of endothelin-1 (ET-1) from the mesenteric arteries of Wistar rats with streptozotocin-induced diabetes (STZ-DM) rats and nondiabetic rats was measured by a specific enzyme immunoassay following purification using an immunoaffinity column. The mesenteric arteries from STZ-DM rats released a significantly higher amount of ET-1 as compared to control rats (35.8 +/- 2.8 vs 14.9 +/- 2.0 pg/1hr, p less than 0.05). The plasma level of ET-1 in STZ-DM rats was also elevated to a significant extent as compared to controls (5.1 +/- 0.4 vs 3.0 +/- 0.4 pg/ml, p less than 0.05). The systolic blood pressure of STZ-DM rats was significantly higher than of the controls (p less than 0.05). The increased level of plasma ET-1 as well as its release from the mesenteric artery of STZ-DM rats may suggest its release following damage to the endothelium caused by diabetes and/or by associated changes in blood pressure.     

Secretion of endothelin-1 in human endothelial cell line but not in B cell line by transfection of preproendothelin-1 cDNA.

Stable transformants with preproendothelin-1 (preproET-1) cDNA were established for the study of the regulation of endothelin-1 (ET-1) biosynthesis in human cells. ET-1, a potent vasoconstrictor peptide, is produced by endothelial cells and is secreted into the blood at a low level. Human preproET-1 cDNA was introduced into two immortal human cell lines, t-HUE2, an endothelial cell line, and Raji, a B cell line, with Ecogpt selection. Several stable transformants of t-HUE2 expressed extraordinarily high levels of preproET-1 specific mRNA and secreted ET-1 into serum-free culture medium, while the transformants of Raji cells expressed high levels of ET-1 mRNA, but secreted a negligible amount of ET-1. Immunocytochemical studies of intracellular ET-1 content revealed that there were some defects in the translation or processing of preproET-1 in the B cell line transformants. In addition, the ratio of ET-1 to ET-1 precursor (big ET-1) was much higher in the t-HUE2 transformants than in normal endothelial cells, suggesting that t-HUE2 transformants (for example t-HUE2-1) possess high levels of endothelin converting enzyme (ECE). The establishment of stable transformants producing high levels of ET-1 in serum-free medium will be useful for the study of cell-type-specific translation and processing to mature ET-1, and of the regulatory factors of ECE.     

Interleukin-2 enhances the release of endothelin-1 from the rat mesenteric artery

We measured the ET-1 concentration in plasma and in the perfusate of the mesenteric arteries of rats treated with a therapeutic dose of IL-2 for 7 days (100000 U/Kg, iv.). The plasma ET-1 concentration in rats given IL-2 was 14.2 +/- 3.2 pg/ml which was significantly greater than that in the controls (2.5 +/- 0.4 pg/ml, P less than 0.05). The mesenteric arteries also released a significantly greater amount of ET-1 (29.5 +/- 1.6 pg/h) than that in controls (16.8 +/- 2.3 pg/h, P less than 0.01). The arterial blood pressure was significantly lower after IL-2 treatment than the pre-dosing level (P less than 0.05). It is concluded that IL-2 induces ET-1 release from the vascular wall, possibly as a result of reversible endothelial dysfunction caused by IL-2.     

17beta-estradiol deficiency reduces potassium excretion in an angiotensin type 1 receptor-dependent manner

This study examined the effects of ovariectomy (OVX) and 17beta-estradiol (E(2)) replacement (OVX + E(2)) on renal function in Sprague-Dawley rats. OVX caused a 40% decrease in the fractional excretion of potassium (FE(K(+))) that was prevented by E(2) replacement [Sham, 24.2 +/- 2.9%; OVX, 14.5 +/- 2.1% (P < 0.05 vs. OVX + E(2)); and OVX + E(2), 26.2 +/- 2.7%; n = 7-11] and that corresponded to significant increases in plasma potassium [(in mmol/l): Sham, 3.15 +/- 0.087; OVX, 3.42 +/- 0.048 (P < 0.05 vs. OVX + E(2)); and OVX + E(2), 3.19 +/- 0.11; n = 7-11]. No effects of OVX were detected on plasma levels of sodium and aldosterone. Angiotensin II type 1 receptor (AT(1)R) densities in ovariectomized rats were 1.4-fold and 1.3-fold higher in glomerular [maximum binding capacity (B(max); in fmol/mg protein): Sham, 482 +/- 21; OVX, 666 +/- 20 (P < 0.05 vs. OVX + E(2)); and OVX + E(2), 504 +/- 26; n = 7-11] and proximal tubular [B(max) (in fmol/mg protein): Sham, 721 +/- 16; OVX, 741 +/- 24 (P < 0.05 vs. OVX + E(2)); and OVX + E(2), 569 +/- 23; n = 7-11] membranes compared with E(2) replete animals, respectively. Both the angiotensin-converting enzyme inhibitor captopril and the AT(1)R antagonist losartan prevented the OVX-induced decrease in the FE(K(+)) and the increase in renal AT(1)R densities, suggesting that E(2) deficiency reduces potassium excretion in an ANG II/AT(1)R-dependent manner. These findings may have implications for renal function in postmenopausal women as well as contribute to the reasons underlying the age-induced increase in susceptibility to hypertension-associated disease in women.     

Enhanced expression of endothelin-1 and endothelin-converting enzyme-1 in acute hypoxic rat aorta

Deeply anesthetized male Wistar rats were perfused by Hanks' balanced salt solution bubbled with either 95%air and 5%CO2 (normoxic group) or 95%N2 and 5%CO2 (hypoxic group) from the thoracic aorta for 30 min, and the isolated abdominal aortae from both groups were used for electron microscopy, immunocytochemistry of endothelin (ET)-1 and ET-converting enzyme (ECE)-1, and in situ hybridization of preproET-1 mRNA. A remarkable increase in the number of Weibel-Palade (WP) bodies, storage sites of ET-1 and ECE-1, occurred in the hypoxic group when compared to the normoxic group. Immunoreactivities for ET-1 and ECE-1, and signals for preproET-1 mRNA were seen along the endothelia of both groups, but the intensities were significantly elevated in the hypoxic group. The increase in the number of ECE-1 immunoreactive gold particles was noticed especially in WP bodies in the hypoxic group. These findings indicate the enhancement of preproET-1 synthesis in the aortic endothelial cells as well as the acceleration of ET-1 processing in increased WP bodies in such cells in an experimentally hypoxic condition of the rat aortae.     

Use of cognitive strategies in rats: the role of estradiol and its interaction with dopamine

Accumulating evidence suggests a role for estrogen in the use of a particular cognitive strategy when solving a maze task. In order to confirm the role of estrogen in this phenomenon, ovariectomized (OVX) female rats receiving either high ( approximately 90 pg/ml) or low ( approximately 32 pg/ml) circulating levels of 17beta-estradiol benzoate (E2) performed a plus maze task for a reward. Consistent with previous research, OVX rats receiving low levels of E2 utilized a striatum-mediated response strategy while OVX rats administered high levels of E2 employed a hippocampus-mediated place strategy. Furthermore, following a systemic injection of a moderate dose of either a dopamine D1 (SKF 83566, 0.1 mg/kg IP) or D2 (raclopride, 0.5 mg/kg IP) receptor antagonist, low E2 rats were seen to use the opposite strategy and exercise a hippocampus-mediated place strategy in order to obtain the reward. At the same doses, high E2 rats did not change from using a place strategy. At a lower dose, these drugs shifted high E2 rats such that they showed an equal propensity for either strategy; this was not observed in low E2 rats. These results corroborate previous findings that E2 plays a significant role in the use of either a response or place strategy when solving a maze for a reward. In addition, the shift in strategy after dopamine receptor blockade implies the importance of central dopamine function in selecting a cognitive strategy to solve such tasks. It is suggested that estrogen alters cognitive strategy not only by improving hippocampal function, but also by altering dopamine-regulated striatal function.     

Endothelin-1 contributes to the sexual differences in renal damage in DOCA-salt rats

We investigated whether gender differences in renal damage in DOCA-salt hypertension are associated with effects of ovarian hormones and/or endothelin-1 (ET-1). Renal injuries and renal pre-pro-ET-1 mRNA expression were enhanced in male and female ovariectomized (OVX) DOCA rats versus female DOCA rats. Treatment with estrogen plus progesterone or progesterone, but not estrogen alone, attenuated renal damage and pre-pro-ET-1 mRNA expression in OVX DOCA rats. The ETA antagonist BMS182874 greatly ameliorated renal damage in male and OVX DOCA rats. In conclusion, the ovarian hormones have a protective role on the renal structural alterations in female DOCA rats by modulating effects of ET-1, via ETA receptors.     

Estrogen status and skeletal muscle recovery from disuse atrophy.

Although estrogen loss can alter skeletal muscle recovery from disuse, the specific components of muscle regrowth that are estrogen sensitive have not been described. The primary purpose of this study was to determine the components of skeletal muscle mass recovery that are biological targets of estrogen. Intact, ovariectomized (OVX), and ovariectomized with 17beta-estradiol replacement (OVX+E2) female rats were subjected to hindlimb suspension for 10 days and then returned to normal cage ambulation for the duration of recovery. Soleus muscle mass returned to control levels by day 7 of recovery in the intact animals, whereas OVX soleus mass did not recover until day 14. Intact rats recovered soleus mean myofiber cross-sectional area (CSA) by day 14 of recovery, whereas the OVX soleus remained decreased (42%) at day 14. OVX mean fiber CSA did return to control levels by day 28 of recovery. The OVX+E2 treatment group recovered mean CSA at day 14, as in the intact animals. Myofibers demonstrating central nuclei were increased at day 14 in the OVX group, but not in intact or OVX+E2 animals. The percent noncontractile tissue was also increased 29% in OVX muscle at day 14, but not in either intact or OVX+E2 groups. In addition, collagen 1a mRNA was increased 45% in OVX muscle at day 14 of recovery. These results suggest that myofiber growth, myofiber regeneration, and extracellular matrix remodeling are estrogen-sensitive components of soleus muscle mass recovery from disuse atrophy.     

雌激素受体α和β在不同雌激素干预大鼠骨代谢中的表达

应用雌性大鼠的骨质疏松模型,通过骨密度(BMD)检测、RT-PCR和Westernblot等技术观察去卵巢(Ovariectomy,OVX)、结合性雌激素(ConjugatedEquineEstrogens,CEE)和戊酸雌二醇(EstradiolValerate,EV)对大鼠松质骨骨量以及松质骨中雌激素受体(ER)α和βmRNA和蛋白表达的影响,探讨两受体亚型在介导雌激素参与松质骨代谢的不同作用机制以及不同来源雌激素对ERα和ERβ表达的差异性调节。40只7-8周龄的雌性Sprague-Dawley大鼠,在观察动情周期后随机分成四组:对照组(Control,n=10)、去卵巢组(Ovariectomy,OVX,n=10)、去卵巢后结合性雌激素治疗组(CEE,n=10)和去卵巢后戊酸雌二醇治疗组(EV,n=10)。对照组大鼠行假手术,其余三组行去卵巢手术。术后48天(12个动情周期),对照组和OVX组用生理盐水喂养12天(3个动情周期),CEE组和EV组分别用药物的生理盐水溶液喂养12天。结果显示:在对照组中,大鼠松质骨ERα的蛋白表达水平显著性高于ERβ蛋白表达水平,而ERα的mRNA表达水平显著性低于ERβmRNA水平。与对照组相比,OVX组大鼠松质骨中ERα的蛋白表达水平显著性降低,ERαmRNA表达水平显著性增加,而ERβ蛋白和mRNA的表达水平均显著性增加。与OVX组相比,CEE组大鼠松质骨中ERβ蛋白和mRNA的表达水平均显著性下降,而EV组大鼠松质骨中ERα蛋白表达显著性上升,ERαmRNA表达显著性下降,ERβ蛋白表达显著性下降。此外,OVX大鼠松质骨的骨密度下降均可通过应用CEE和EV得到显著性改善。上述结果提示:⑴ERα可能是大鼠松质骨中优势表达的受体亚型,在介导雌激素参与松质骨代谢中起着主导作用。⑵不同来源雌激素可能侧重不同的ER亚型途径产生骨保护效应。     

ET-1 in the myocardial interstitium: relation to myocyte ECE activity and expression

Increased plasma levels of endothelin-1 (ET-1) have been identified in congestive heart failure (CHF), but local myocardial interstitial ET-1 levels and the relation to determinants of ET-1 synthesis remain to be defined. Accordingly, myocardial interstitial ET-1 levels and myocyte endothelin-converting enzyme (ECE)-1 activity and expression with the development of CHF were examined. Pigs were instrumented with a microdialysis system to measure myocardial interstitial ET-1 levels with pacing CHF (240 beats/min, 3 wk; n = 9) and in controls (n = 14). Plasma ET-1 was increased with CHF (15 +/- 1 vs. 9 +/- 1 fmol/ml, P < 0.05) as was total myocardial ET-1 content (90 +/- 15 vs. 35 +/- 5 fmol/g, P < 0.05). Paradoxically, myocardial interstitial ET-1 was decreased in CHF (32 +/- 4 vs. 21 +/- 2 fmol/ml, P < 0.05), which indicated increased ET-1 uptake by the left ventricular (LV) myocardium with CHF. In isolated LV myocyte preparations, ECE-1 activity was increased by twofold with CHF (P < 0.05). In LV myocytes, both ECE-1a and ECE-1c mRNAs were detected, and ECE-1a expression was upregulated fivefold in CHF myocytes (P < 0.05). In conclusion, this study demonstrated compartmentalization of ET-1 in the myocardial interstitium and enhanced ET-1 uptake with CHF. Thus a local ET-1 system exists at the level of the myocyte, and determinants of ET-1 biosynthesis are selectively regulated within this myocardial compartment in CHF.     

Estrogen receptor-alpha mediates estrogen protection from angiotensin II-induced hypertension in conscious female mice

It has been shown that the female sex hormones have a protective role in the development of angiotensin II (ANG II)-induced hypertension. The present study tested the hypotheses that 1) the estrogen receptor-alpha (ERalpha) is involved in the protective effects of estrogen against ANG II-induced hypertension and 2) central ERs are involved. Blood pressure (BP) was measured in female mice with the use of telemetry implants. ANG II (800 ng.kg(-1).min(-1)) was administered subcutaneously via an osmotic pump. Baseline BP in the intact, ovariectomized (OVX) wild-type (WT) and ERalpha knockout (ERalphaKO) mice was similar; however, the increase in BP induced by ANG II was greater in OVX WT (23.0 +/- 1.0 mmHg) and ERalphaKO mice (23.8 +/- 2.5 mmHg) than in intact WT mice (10.1 +/- 4.5 mmHg). In OVX WT mice, central infusion of 17beta-estradiol (E(2); 30 microg.kg(-1).day(-1)) attenuated the pressor effect of ANG II (7.0 +/- 0.4 mmHg), and this protective effect of E(2) was prevented by coadministration of ICI-182,780 (ICI; 1.5 microg.kg(-1).day(-1), 18.8 +/- 1.5 mmHg), a nonselective ER antagonist. Furthermore, central, but not peripheral, infusions of ICI augmented the pressor effects of ANG II in intact WT mice (17.8 +/- 4.2 mmHg). In contrast, the pressor effect of ANG II was unchanged in either central E(2)-treated OVX ERalphaKO mice (19.0 +/- 1.1 mmHg) or central ICI-treated intact ERalphaKO mice (19.6 +/- 1.6 mmHg). Lastly, ganglionic blockade on day 7 after ANG II infusions resulted in a greater reduction in BP in OVX WT, central ER antagonist-treated intact WT, central E(2) + ICI-treated OVX WT, ERalphaKO, and central E(2)- or ICI-treated ERalphaKO mice compared with that in intact WT mice given just ANG II. Together, these data indicate that ERalpha, especially central expression of the ER, mediates the protective effects of estrogen against ANG II-induced hypertension.     

Plasma endothelin-1 levels and albumin excretion rate in normotensive,microalbuminuric type 2 diabetic patients

BACKGROUND: Endothelin-1 (ET-1) is able to determine functional and structural renal alterations and plasma levels of this vasoconstrictor peptide are increased in diabetic patients. In a selected group of type 2 normotensive diabetic patients with microalbuminuria, we investigated circulating ET-1 levels compared to a control group and verified whether there is a relationship between ET-1 levels and albumin excretion rate in diabetics. SUBJECTS AND METHODS: Thirty-two microalbuminuric type 2 diabetic patients (12 males and 20 females; mean age 57 +/- 8 years) without hypertension, renal failure, hypercholesterolemia or atherosclerotic damage were selected. The control group was made up of 28 healthy subjects matched for sex and age. Blood pressure, creatinine clearance, serum cholesterol and plasma ET-1 values were determined in diabetic and control group. In diabetic patients, glycosilated hemoglobin and urinary albumin excretion rate were also assayed. Mean ET-1 values in diabetics and controls were compared using Student's t-test. Linear regression test was done to relate two variables. Statistical significance was set at p<0.05. RESULTS: Mean ET-1 values were significantly higher in the diabetic group than in controls (11.77 +/- 1.16 pg/ml vs 8.9 +/- 2.1 pg/ml; p<0.05). No relationship (p>0.05) was found between circulating ET-1 and blood pressure, creatinine clearance, serum cholesterol and metabolic control in diabetics. There was a significant positive correlation (r=0.403; p=0.03) between plasma ET-1 levels and albumin excretion rate in diabetic patients. CONCLUSIONS: Our results showed that circulating ET-1 values were increased in microalbuminuric, normotensive, type 2 diabetic patients and correlated with albumin excretion rate. These findings confirm that endothelial dysfunction, as expressed by ET-1 levels, occurs early in these patients and support the hypothesis of a potential role for this peptide in development of microalbuminuria in diabetic nephropathy.     

Estrogen effects on osmotic regulation of AVP and fluid balance

To determine estrogen effects on osmotic regulation of arginine vasopressin (AVP) and body fluids, we suppressed endogenous estrogen and progesterone using the gonadotropin-releasing hormone (GnRH) analog leuprolide acetate (GnRHa). Subjects were assigned to one of two groups: 1) GnRHa alone, then GnRHa + estrogen (E, n = 9, 25 +/- 1 yr); 2) GnRHa alone, then GnRHa + estrogen with progesterone (E/P, n = 6, 26 +/- 3). During GnRHa alone and with hormone treatment, we compared AVP and body fluid regulatory responses to 3% NaCl infusion (HSI, 120 min, 0.1 ml. min(-1). kg body wt(-1)), drinking (30 min, 15 ml/kg body wt), and recovery (60 min of seated rest). Plasma [E(2)] increased from 23.9 to 275.3 pg/ml with hormone treatments. Plasma [P(4)] increased from 0.6 to 5.7 ng/ml during E/P and was unchanged (0.4 to 0.6 ng/ml) during E. Compared with GnRHa alone, E reduced osmotic AVP release threshold (275 +/- 4 to 271 +/- 4 mosmol/kg, P < 0.05), and E/P reduced the AVP increase in response during HSI (6.0 +/- 1.3 to 4.2 +/- 0.6 pg/ml at the end of HSI), but free water clearance was unaffected in either group. Relative to GnRHa, pre-HSI plasma renin activity (PRA) was greater during E (0.8 +/- 0.1 vs. 1.2 +/- 0.2 ng ANG I. ml(-1). h(-1)) but not after HSI or recovery. PRA was greater than GnRHa during E/P at baseline (1.1 +/- 0.2 vs. 2.5 +/- 0.6) and after HSI (0.6 +/- 0.1 vs. 1.1 +/- 1.1) and recovery (0.5 +/- 0.1 vs. 1.3 +/- 0.2 ng ANG I. ml(-1). h(-1)). Baseline fractional excretion of sodium was unaffected by E or E/P but was attenuated by the end of recovery for both E (3.3 +/- 0.6 vs. 2.4 +/- 0.4%) and E/P (2.8 +/- 0.4 vs 1.7 +/- 0.4%, GnRHa alone and with hormone treatment, respectively). Fluid retention increased with both hormone treatments. Renal sensitivity to AVP may be lower during E due to intrarenal effects on water and sodium excretion. E/P increased sodium retention and renin-angiotensin-aldosterone stimulation.