The characteristics of the programming action of androgens on the formation of sexual dimorphism in the level of corticosteroid-binding globulin in rats

The role of sex steroids in the programming of the level of serum corticosteroid binding globulin (CBG) of male and female rats has been studied at different stages of ontogenesis. It was shown that castration of adult males lead to the increase of the level of CBG, but not to the elimination of sex differences. Gonadectomy of males up to 28th day of postnatal life results in complete feminization of the CBG content in these animals at the age of 10-12 weeks. The castration after 35th day of life does not prevent the formation of the male phenotype of CBG content. The results of administration of testosterone-propionate (TP) to castrated males at different periods of ontogenesis suggests that the sensitivity to irreversible negative action of androgens appears after 28th day of life and disappears after the puberty. It was concluded that short period of ontogenesis from 29th to 35th days of life is critical for the realization of the irreversible masculinization of CBG level upon the influence of androgens in the physiological conditions. It was found that injections of both synthetic estrogens diethylstilbestrol or TP in the sensitive period of ontogenesis lead to the expression of male phenotype of CBG level in a similar fashion.     

Sexual differentiation of gonadotrophin secretion, sexual orientation and gender role behavior

The positive estrogen feedback was found to be a relatively sex-specific reaction of the hypothalamo-hypophyseal system in rats as well as in human beings. It is dependent--most of all--on the estrogen convertible androgen level during sexual brain differentiation, but also on an estrogen priming effect in adulthood. The lower the estrogen convertible androgen or primary estrogen level during brain differentiation, the higher is the evocability of a positive estrogen action on LH secretion in later life. In clinical studies, we were able to induce a positive estrogen feedback on LH secretion in most intact homosexual men in clear-cut contrast to intact hetero- or bisexual men. These findings were strongly confirmed by Gladue and associates. In addition, the evocability of a positive estrogen feedback was also demonstrable in most homosexual male-to-female transsexuals in significant contrast to hetero- or bisexual male-to-female transsexuals. These findings suggest that homosexual males possess, at least in part, a predominantly female-differentiated brain, which may be caused by a low estrogen convertible androgen level during brain organization. Recently, the following relations were found between sex hormone levels during brain differentiation and sex-specific responses in adulthood: (1) estrogens--which are mostly converted, however, from androgens--are responsible for the sex-specific organization of gonadotrophin secretion and hence the evocability of a positive estrogen feedback in later life; (2) estrogens and androgens, occurring during brain differentiation, predetermine synergistically sexual orientation and (3) androgens--without conversion to estrogens--are responsible for the sex-specific organization of gender role behaviour in later life. Furthermore, the organization periods for sex-specific gonadotrophin secretion, sexual orientation and gender role behaviour are not identical but overlapping. Thus, combinations as well as dissociations between deviations of the neuroendocrine organization of sex-specific gonadotrophin secretion, sexual orientation and gender role behaviour are conceivable. Most recently, female-type sexual orientation could be converted to male-type sexual orientation in adult rats by administration of the dopamine agonist and serotonin antagonist lisuride.     

Evidence of a sex related difference of transcortin level in adult ducks

Corticosteroid binding globulin (CBG) serum level as evaluated by either equilibrium dialysis or gel filtration was found to be higher in male than in female adult ducks during the reproductive period. Castration did not modify CBG concentrations in females,whereas in males it induced a significant decrease in CBG, to the level observed in intact or castrated females. Testosterone injections administrated to castrated females increased CBG to the level of adult intact males. Finally it was found that testosterone stimulated CBG production in ducks without altering thyroxine levels.     

Estrogenic support of motoneuron dendritic growth via the neuromuscular periphery in a sexually dimorphic motor system

The lumbar spinal cord of rats contains the sexually dimorphic, steroid-sensitive spinal nucleus of the bulbocavernosus (SNB). In males, the growth of SNB dendrites is steroid-dependent: dendrites fail to grow after castration, but grow in castrates treated with androgens or estrogens. Blocking estradiol synthesis or estrogen receptors in gonadally intact males attenuates SNB dendritic growth, suggesting that estrogens are required and must be able to act at their receptors to support normal masculine dendritic growth. However, SNB motoneurons do not accumulate estrogens, suggesting that estrogens act indirectly to support SNB dendritic growth. In this experiment, we examined whether local estrogen action in the neuromuscular periphery was involved in the postnatal development of SNB motoneurons. Motoneuron morphology was assessed in gonadally intact and castrated males. Gonadally intact males were left untreated or given either blank or tamoxifen implants sutured to the target musculature, or tamoxifen interscapular implants. Castrated males were left untreated or were given estradiol by muscle or interscapular implants or systemic injection during the period of SNB dendritic growth. At postnatal day 28, when SNB dendritic length is normally maximal, SNB motoneurons were retrogradely labeled with cholera toxin-HRP and reconstructed in three dimensions. While interscapular tamoxifen implants were ineffective, blocking estrogen receptors at the target musculature resulted in attenuation of SNB dendritic growth. In contrast, while interscapular implants of estradiol were ineffective, local treatment with estradiol at the target musculature in castrated males resulted in masculinization of dendritic growth. Thus, estrogens may act by an indirect action in the neuromuscular periphery to support SNB dendritic growth.     

Estrogenic influence on the regulation of hepatic estrogen receptor-alpha and serum level of angiotensinogen in female rats

The majority of data regarding biological effects of estrogens is based on studies in male rats or ovariectomized (Ovx) female rats. Therefore, in this study, the effects of estradiol treatment on the regulation of the hepatic estrogen receptor and the level of circulating angiotensinogen were examined in the intact female rat. The data were compared with that of the hypophysectomized (Hx) rat. Animals were treated with either low (physiological) or high (pharmacological) doses of estrogen. In intact rats, the hepatic estrogen receptor (ER) level increased with increasing doses of estrogen. This was in contrast to the Hx rats where growth hormone (GH) and dexametasone (Dex) in combination were the sole modulators of the estrogen receptor. The angiotensinogen level increased in normal rats after estrogen administration in a dose dependent manner, regardless of the mode of administration. The pure antiestrogen ICI 182 780 efficiently blocked the increase in circulating angiotensinogen. The conclusion is that in the normal female, estrogens are important modulators of the serum angiotensinogen level.     

The effect of gonadectomy and aromatase inhibition on the excretion of 19-nordeoxycorticosterone in rats

19-Nordeoxycorticosterone (19-norDOC) is a powerful mineralocorticoid, which has been postulated to be involved in the pathogenesis of some forms of hypertension. The urinary excretion of 19-norDOC by female rats is up to 20 times that of males. To demonstrate the influence of the gonads on the excretion of 19-norDOC, we measured the excretion of 19-norDOC in intact and gonadectomized male and female rats with and without replacement with testosterone (40 mg testosterone enanthate s.c.) or estrogen (4 mg estradiol valerate s.c.) and in intact animals receiving the aromatase inhibitor, 10-propargyl androstenedione (10-pA) (10 mg s.c.). Orchiectomy produced a significant increase in the urinary excretion of 19-norDOC in males. Testosterone treatment decreased 19-norDOC excretion by castrated males to below intact values, while estrogen administration increased its excretion. Oophorectomy had no consistent effect on 19-norDOC excretion. In oophorectomized females, testosterone administration significantly suppressed 19-norDOC excretion and estrogen replacement increased excretion slightly. 10-pA had little effect on the excretion of 19-norDOC in intact rats of either sex. In conclusion, it appears that 19-norDOC production is inhibited by testosterone, but is affected only slightly by estrogens.     

Role of estrogens in development of prostate cancer

Estrogens have previously been extensively used in prostate cancer treatment. Serious side effects, primarily in cardiovascular system have, however, limited their use. The therapeutic effect of estrogen in preventing prostate cancer growth was mainly obtained indirectly by feedback inhibition of the hypothalamic release of LRH leading to lowered serum androgen levels and castration like effects. Prostate tissue is also most probably a target for direct regulation by estrogens. Prostate contains estrogen receptor alpha (ERalpha) and beta (ERbeta), which are localized characteristically in stroma and epithelium, respectively. The physiological function of these receptors is not known but there is evidence of the role of estrogens in prostatic carcinogenesis. Developing prostate seems particularly sensitive to increased level of endogenous and/or exogenous estrogens. Perinatal or neonatal exposure of rats and mice to estrogens leads to "imprinting" of prostate associated with increased proliferation, inflammation and dysplastic epithelial changes later in life. Prolonged treatment of adult rodents with estrogens along with androgens also leads to epithelial metaplasia, PIN-like lesions and even adenocarcinoma of prostate speaking for the role of estrogen in prostate cancer development. Recent results concerning antiestrogen inhibition of prostate cancer development beyond PIN-type lesions in transgenic mouse models further suggests a role for estrogens in prostate cancer progression. These results also suggest that direct inhibition of estrogen action at the level of prostate tissue may provide an important novel principle of development of prostate cancer therapies.     

Effect of ipriflavone on the response of uterus and thyroid to estrogen

Ipriflavone, 7-isopropoxy-3-phenyl-4H-1-benzopyran-4-one, was uterotropic in intact but not in ovariectomized rats. When it was administered simultaneously with estrone and estradiol-17 beta to ovariectomized rats, ipriflavone increased the uterotropic activities of both estrogens. The uterotropic activity of this compound in intact rats might be attributable to its action on the target organ of estrogen by augmenting the response to the hormone because none of the other possibilities for accelerating uterine growth such as gonadotropin-releasing, gonadotropic, and estrogen metabolism-retarding activities were demonstrated experimentally. In addition, ipriflavone increased the calcitonin releasing activity of estrone when these compounds were administered simultaneously to ovariectomized rats.     

A re-examination of the lordosis response in female rats given high doses of testosterone propionate or estradiol benzoate in the neonatal period

High lordosis quotients (LQ) were observed when female Wistar rats injected with 1.25 mgm of testosterone propionate (TP) on Day 4 of postnatal life were tested as intact adults. The high LQ was not due to testing during the lights-on period, the age at which the females were tested, the use of a strain that was insensitive to the masculinizing action of TP or estradiol benzoate (EB), the age at which the females were injected with TP or EB, or an abnormal response to estrogen. High LQ values were found in similar tests on adult female rats of two other strains injected with 1.25 mgm TP on Day 4 of life. A marked reduction of the facilitatory action of progesterone on receptivity in estrogen-primed animals was demonstrated in the females of all three strains treated with TP or EB during the neonatal period and for males after castration as adults.Analysis of the experimental records of the mating tests showed that females anovulatory following TP or EB administration during the neonatal period and tested either intact and under the influence of endogenous hormones or under the influence of exogenous estrogen showed a rapid and highly significant increase in receptivity during the course of prolonged (20 min) tests with two or three active stimulus males. This effect was very much reduced if the treated females were under the influence of exogenous estrogen plus progesterone. The effect was not seen in males castrated as adults and treated with estrogen, or in females not treated with steroids in the neonatal period and tested intact at proestrus alone or under the influence of exogenous steroids after ovariectomy. A significant increase in LQ during the test period was observed in females of the Wistar strain which were anovulatory as a result of exposure to constant light and were tested intact without any exogenous hormone being administered.It is suggested that although tests involving a limited number of mounts or attempts to mount at low rates over a short period of time may be adequate to determine the degree of receptivity of normal female rats they are not adequate to establish the capacity of female rats treated with steroid hormones during the neonatal period to display the lordosis response.     

Effect of sex hormones on plasma T-kininogen in the rat

The influence of sex hormones on rat plasma T-kininogen concentration was examined. The level of T-kininogen in the post-pubertal female rat is about 3-times that of the male animal. Female rats castrated as adults or 15 days after birth, had low T-kininogen concentrations, near those of male rats. In contrast, castration of mature or immature male animals induced no change in T-kininogen. Treatment of castrated female or male rats with 17 alpha-ethinylestradiol significantly increased the T-kininogen level, whereas administration of testosterone or progesterone had no effect. The influence of estrogen was specific for T-kininogen, since plasma HMW kininogen concentration was the same in male and female rats and was not affected by castration or sex hormone treatment. T-kininogen concentration was not significantly changed in pregnant rat between the 12th and the 20th day of pregnancy, but increased after parturition. It was high in the newborn rat at birth and then decreased similarly over the next 3 weeks in males and females. It continued to decrease in the males, reaching the level of the adult rat, but it increased in the female from 3-4 weeks of age and reached the adult level at about 6-8 weeks. These data indicate that natural estrogens have a physiological influence on the plasma level of T-kininogen in female rats whereas testosterone had no effect on either male or castrated female rats. HMW kininogen is not physiologically dependent on sex hormones.     

Water-soluble metabolites of the estrogens. Quantitation of C-18 tetrols in rat feces.

Several radioactive estrogens possessing one, two and three hydroxyl groups were injected orally (and in the case of estrone sulfate also intraperitoneally) into adult male rats. The rats were either intact or had ligated or cannulated bile ducts. Two unconjugated estrogen tetrols together represented 21 - 87% of the total metabolites in the intact rat. One of the tetrols was 2-hydroxyestriol (estra-1,3,5(10)-triene-2,3,16alpha,17beta-tetrol); the other may be estra-1,3,5(10)-triene-2,3,6xi,17beta-tetrol but this was not confirmed. It is concluded that poly-hydroxylated estrogens represent a very large proportion of the previously unidentified water-soluble metabolites of the estrogens in the adult male rat.     

Sexual Dimorphism in Development of Kidney Damage in Aging Fischer-344 Rats

BackgroundAging kidneys exhibit slowly developing injury and women are usually protected compared with men, in association with maintained renal nitric oxide.ObjectivesOur purpose was to test 2 hypotheses: (1) that aging intact Fischer-344 (F344) female rats exhibit less glomerular damage than similarly aged males, and (2) that loss of female ovarian hormones would lead to greater structural injury and dysregulation of the nitric oxide synthase (NOS) system in aging F344 rat kidneys.MethodsWe compared renal injury in F344 rats in intact, ovariectomized, and ovariectomized with estrogen replaced young (6 month) and old (24 month) female rats with young and old intact male rats and measured renal protein abundance of NOS isoforms and oxidative stress.ResultsThere was no difference in age-dependent glomerular damage between young or old intact male and female F344 rats, and neither ovariectomy nor estrogen replacement affected renal injury; however, tubulointerstitial injury was greater in old males than in old females. These data suggest that ovarian hormones do not influence these aspects of kidney aging in F344 rats and that the greater tubulointerstitial injury is caused by male sex. Old males had greater kidney cortex NOS3 abundance than females, and NOS1 abundance (alpha and beta isoforms) was increased in old males compared with both young males and old females. NOS abundance was preserved with age in intact females, ovariectomy did not reduce NOS1 or NOS3 protein abundance, and estrogen replacement did not uniformly elevate NOS proteins, suggesting that estrogens are not primary regulators of renal NOS abundance in this strain. Nicotinamide adenine dinucleotide phosphate oxidase-dependent superoxide production and nitrotyrosine immunoreactivity were increased in aging male rat kidneys compared with females, which could compromise renal nitric oxide production and/or bioavailability.ConclusionsThe kidney damage expressed in aging F344 rats is fairly mild and is not related to loss of renal cortex NOS3 or NOS1 alpha.     

Sex differences in the accumulation of estrogen receptors in nuclei of liver cells and increase of angiotensinogen concentration in the blood of rats after administration of low doses of synthetic estrogens

The significance of sex differences in the level of estrogen receptors (ER) in hepatocytes for direct effects of estrogens in male and female rat livers was investigated. 4-5-fold increase in ER level and 20-30%-elevation in plasma angiotensinogen (AG) occurred after a single injection of 0.5 microgram of hexestrol (HE) in female and gonadectomized male rats. In male liver, where the cytosol ER content is two fold lower than that in female rats, nuclear ER level was shown to be very low and unchanged after HE injection, plasma AG also did not change. Injection of 0.5 microgram of ethinylestradiol produced similar effect. Injection of a greater dose of estrogen caused an enhancement in plasma AG level in males. The existence of sex differences in hepatic ER level seems to cause in some conditions different response of metabolic processes in male and female rat liver after estrogenization.     

Attenuating the defeminization of the neonatal rat brain: Mechanisms of action of cyproterone acetate, 1,4,6-androstatriene-3,17,-dione and a synthetic progestin, R5020

Although defeminization of the rat brain appears to depend significantly on the conversion of testosterone (T) to estradiol (E₂), the antiandrogenic steroid cyproterone acetate (CA) is able to attenuate defeminization. In order to study the mechanism of action of CA on brain sexual differentiation, newborn male rats were given subcutaneous injections of this steroid on postnatal Days 2–6. When castrated on Day 70 and given estrogen and progesterone, these CA-treated males displayed elevated lordosis quotients (LQ) compared to controls. CA-treated neonatal males were also examined at the end of the drug treatment to ascertain the mechanism of drug action: (1) Serum T levels were normal; (2) Brain cell nuclear estrogen receptor occupation, estimated by an exchange assay, was reduced by ≈ 30% in the brains of the CA-treated males, although the ability of exogenous E₂ to occupy these brain estrogen receptors was not reduced. Other work has demonstrated a weak competitive effect of cyproterone on aromatization, and thus cyproterone acetate may have interfered with the conversion of T to E₂ CA also has progestogenic activity, and 5-mm capsules of a potent synthetic progestin, R5020, given to newborn male rats on Days 2–6, are shown to elevate the LQ after postnatal Day 70 to the same extent as CA. However, R5020 did not reduce estrogen receptor occupation in the neonatal male rat brain and was without effect on serum T levels in the neonatal male. Because of the implied role of T-derived estrogens in defeminization, an experiment was conducted showing that the defeminizing action of estradiol benzoate given to 3-day-old female rat pups is attenuated by the antiestrogen, CI628, and not by the potent inhibitor of aromatization, 1,4,6-androstatriene-3,17-dione (ATD). This result complements previous experiments showing that both ATD and CI628 attenuate the defeminization produced by T. Taken together, the results lend further support to a pivotal role for aromatization and for estrogen-receptor interactions in the defeminizing effects of T. The actions of progestins such as CA and R5020 in attenuating defeminization are discussed in relation to the recent demonstration of progestin receptors in the neonatal rat brain. It is concluded that CA may act by a combination of actions, both by inhibiting aromatization and by acting as a progestin.     

Oestrogènes et reproduction masculine

The role of estrogen on male reproductive function has become clearer in the last decade. During these years the study of the effect of testosterone, estrogen or an aromatase inhibitor in hypogonadal men provided a first evidence of the effects of estrogens in the regulation of gonadotropin secretion. At the same time, the development of a line of transgenic male mice lacking estrogen receptor α, estrogen receptor β or aromatase gene provided further evidence about the role of estrogens not only in the regulation of gonadotropin secretion, but also on the effects of estrogens on testicular function and development. A confirmation of these actions of estrogens came from the observation of naturally occurring mutations of the estrogen receptor and of the aromatase gene in human males. Based on these data it has been demonstrated that estrogens are major regulators of gonadotropin secretion acting both at pituitary and hypotalamic level. The presence in the human reproductive structures of estrogen receptor α, estrogen receptor β and the aromatase enzyme indicates the existence of receptor α, estrogen receptor β or aromatase estrogen actions at this level. Anyway, the precise role of estrogens in testicular development and function and on the regulation of human spermatogenesis has not yet been precisely clarified.     

Site of negative feedback action of estrogen on gonadotropin secretion in normal women

We have reported that iv administration of conjugated estrogens results in no significant change in the plasma LH-RH level during the negative feedback phase of LH, suggesting that estrogen does not suppress LH by decreasing hypothalamic LH-RH. To determine the site of estrogen action during the negative feedback phase, we studied the pituitary response to a small amount of LH-RH after estrogen administration in normal cyclic women in the mid-follicular phase. The pituitary responses to an iv bolus of 2.5 micrograms of synthetic LH-RH were evaluated by measuring serum LH and FSH 2 h before and 8 h after administration of 20 mg of conjugated estrogens (Premarin). The mean levels of serum LH and FSH were significantly (p less than 0.05) decreased 8 h after the injection. The peak responses of LH and FSH to LH-RH were also significantly (p less than 0.05) reduced after Premarin administration. These findings suggest that the negative feedback effect of estrogen on gonadotropin secretion is caused by its direct suppression on the pituitary response to LH-RH.     

Estrogens and male reproduction

The role of estrogen on male reproductive function has become clearer in the last decade. During these years the study of the effect of testosterone, estrogen or an aromatase inhibitor in hypogonadal men provided a first evidence of the effects of estrogens in the regulation of gonadotropin secretion. At the same time, the development of a line of transgenic male mice lacking estrogen receptor α, estrogen receptor β or aromatase gene provided further evidence about the role of estrogens not only in the regulation of gonadotropin secretion, but also on the effects of estrogens on testicular function and development. A confirmation of these actions of estrogens came from the observation of naturally occurring mutations of the estrogen receptor and of the aromatase gene in human males. Based on these data it has been demonstrated that estrogens are major regulators of gonadotropin secretion acting both at pituitary and hypotalamic level. The presence in the human reproductive structures of estrogen receptor α, estrogen receptor β and the aromatase enzyme indicates the existence of receptor α, estrogen receptor β or aromatase estrogen actions at this level. Anyway, the precise role of estrogens in testicular development and function and on the regulation of human spermatogenesis has not yet been precisely clarified.     

Antagonism of estrogen-induced prolactin release by dihydrotestosterone

Previous work from our laboratory has demonstrated that progesterone can inhibit estrogen-induced prolactin release in female rats. Since androgens have been reported to mimic progesterone actions in certain systems, and to antagonize estrogen action in rat uteri, the purpose of this study was to determine whether androgens, like progestins, can inhibit estrogen-induced prolactin release. The ovariectomized (26 days of age) immature rat was used as the model for analysis of this question. Dihydrotestosterone (DHT) was chosen to be used throughout the study since it does not undergo aromatization to estrogens. In response to estradiol exposure (2 micrograms/rat), prolactin release reached peak values at 12 h and returned to control levels by 24 h. A second injection of estradiol 13 h after its initial injection stimulated a second increase in serum prolactin at 25 h. A single injection of DHT (0.8 mg/kg BW) 1 h before the second estradiol injection blocked the increase in serum prolactin. DHT had no effect on basal serum prolactin levels. The DHT inhibition of estrogen-induced prolactin release required estrogen priming. A dose dependency for the DHT effect was demonstrated, with low doses effective and high doses ineffective, in inhibiting estrogen action. This effect of DHT seemed to be androgen receptor-mediated, since flutamide blocked the effect. However, the possibility of progestin receptor mediation could not be ruled out since RU486 also blocked DHT's effect. Flutamide was also effective in blocking progesterone's inhibition of estrogen-induced action. This is perhaps consistent with an overlap of activities in androgens and progestins reported by several investigators.(ABSTRACT TRUNCATED AT 250 WORDS)     

Sex difference in glucocorticoid binding in rat pituitary is estrogen dependent

Sex-dependent differences in corticosteroid binding were assessed in individual pituitaries from adult male and female rats that had been adrenalectomized 12 h before sacrifice. Soluble binding was assayed in duplicate on LH-20 columns. Gonadally intact females showed significantly less 3H-dexamethasone binding than did intact males (p less than 0.01). This difference was confirmed in a second study (p less than .001). However, when ovariectomized females were compared with gondadectomized males, there was no difference in receptor concentration. Estrogen was able to reverse the effect of ovariectomy: ovariectomized females receiving estrogen (10 micrograms/rat/day) had significantly fewer receptors than intact males; p less than 0.01). Progesterone (500 micrograms/rat/day) did not antagonize the effect of estrogen in the pituitary. A sex difference was also found in the Type I (mineralocorticoid) receptor subpopulation which comprised approximately 10% of the total receptors, with females having fewer receptors than males. These results demonstrate that in the pituitary, the level of functional corticosteroid receptors is subject to a 20% down-regulation by circulating levels of estrogen. This raises the possibility that the lower number of receptors in females may act to reduce their sensitivity to the negative feedback effects of glucocorticoids at the level of the pituitary.