High affinity estrogen binding by rabbit liver

Macromolecular binding components for [³H]estradiol-17β are present to cytosol prepared from rabbit liver. When cytosol from sexually mature male liver was incubated with [³H]estradiol and analyzed for binding on low ionic strength sucrose gradients, two peaks of binding activity were detected. One peak had a sedimentation coefficient of 4–5 S and the other had a sedimentation coefficient of 8–9 S. The two components differed from each other regarding steroid specicity and various physiocochemical parameters. [³H]-estradiol binding to the 4–5 S component was not inhibited by estrogens, 5α-dihydrotestosterone, progesterone or cortisol. Binding to this component did not appera to be saturable and lavel was rapidly stripped from it by cahrcoal. Estradiol bindng to the 8–9 S component was estrogen specific, saturable and of high affinity. The specific binder dissociates on high ionic strength sucrose gradients and sediments as a 4–5 S moiety. The specific binding protein has a K_d of 3.05 · 10⁻¹⁰ M and a dissociation half-time of 33 h and there are 35.2 fmol of binding sites/mg cytosol protein. Estrogen binders are also present in liver cytosol from sexually mature female and sexually immature male rabbits. During prolonged incbuation of [³H]estradiol with mature male liver cytosol at 0–5°C polar metabolites of estradiol are produced.     

Binding of [H]methyltrienolone to androgen receptor in rat liver

The synthetic androgen methyltrienolone is superior to testosterone and androstenedione for the measurement of androgen receptor in tissues where the native ligands are metabolized into inactive derivatives. [³H]Methyltrienolone binds with a high affinity to androgen receptor in cytosol prepared from male rat livers, as the Scatchard analysis revealed that the K_d value was 3.3 · 10⁻⁸ M and the number of binding sites was 35.5 fmol/mg protein. Since methyltrienolone also binds glucocorticoid receptor which exists in rat liver, the apparent binding of androgen receptor is faulty when measured in the presence of glucocorticoid receptor. The binding of methyltrienolone to glucocorticoid receptor can be blocked by the presence of a 100-fold molar excess of unlabeled synthetic glucocorticoid, triamcinolone acetonide, without interfering in its binding to androgen receptor, because triamcinolone does not bind to androgen receptor. Triamcinolone-blocked cytosol exhibited that the K_d value was 2.5 · 10⁻⁸ M and the number of binding sites was 26.3 fmol/mg protein, indicating a reduction to of that in the untreated cytosol. The profile of glycerol gradient centrifiguration indicated that [³H]methyltriemolone-bound receptor migrated in the 8–9 S region in both untreated and triamcinolone-blocked cytosols, but the 8–9 S peak in triamcinolone-blocked cytosol was reduced to about of that of untreated cytosol.     

Different mechanisms of regulation of nuclear reduced nicotinamide-adenine dinucleotide phosphate-dependent 3-oxo steroid 5alpha-reductase activity in rat liver, kidney and prostate

The regulatory mechanisms involved in the control of the nuclear NADPH-dependent 3-ketosteroid 5α-reductase (5α-reductase) activity were studied in liver, kidney and prostate. The substrate used was [1,2-³H]androst-4-ene-3,17-dione (androstenedione) (for liver and kidney) or [4-¹⁴C]androstenedione (for prostate). The hepatic nuclear 5α-reductase activity was greater in female than in male rats, was greater in adult than in prepubertal female rats, increased after castration of male rats, but was not affected by treatment with testosterone propionate or oestradiol benzoate. These regulatory characteristics are in part different from those previously described for the hepatic microsomal 5α-reductase. The renal nuclear metabolism of androstenedione, i.e. 5α reduction and 17β-hydroxy steroid reduction, was relatively unaffected by sex, age, castration and treatment with testosterone propionate. However, treatment of castrated male rats with oestradiol benzoate led to a significant increase in the 5α-reductase activity and a significant decrease in the 17β-hydroxy steroid reductase activity. Finally, the nuclear 5α-reductase activity in prostate was androgen-dependent, decreasing after castration and increasing after treatment with testosterone propionate. In conclusion, the nuclear 5α-reductase activities in liver, kidney and prostate seem to be under the control of distinctly different regulatory mechanisms. The hypothesis is presented that whereas the prostatic nuclear 5α-reductase participates in the formation of a physiologically active androgen, 5α-dihydrotestosterone, this may not be the true function of the nuclear 5α-reductase in liver and kidney. These enzymes might rather serve to protect the androgen target sites in the chromatin from active androgens (e.g. testosterone) by transforming them into less active androgens (e.g. 5α-androstane-3,17-dione and/or 5α-dihydrotestosterone).     

Estrogen interaction with the anterior pituitary of female rats differential cytosol binding, nuclear translocation and stimulation of RNA synthesis by 17β-estradiol and tamoxifen

The interaction of tamoxifen (trans-1-(p-β-dimethylaminoethoxyphenyl)-1,2-diphenylbut-1-ene) with the cytosol estrogen receptor of the anterior pituitary of female rats was studied. No differences were recorded between incubations of cytosol samples with 17β-[³H]estradiol performed in the presence or absence of unlabeled 17β-estradiol and tamoxifen, respectively, thus suggesting that these interactions were at common receptor sites and excluding possible cooperative interactions. Competition experiments and Scatchard plot analysis of saturation experiments add further evidence for common receptor sites. A dissociation constant for tamoxifen of K_d = 2 nM was recorded. Tamoxifen was found to be bound to a moiety sedimenting in the 4–5 S region, on a 6–24% linear sucrose density gradient at low salt concentrations, whereas 17β-estradiol sedimented in the 8–9 S area. These data suggest possible conformational changes of the receptor in the presence of tamoxifen. Furthermore, nuclear estrogen receptor levels remained elevated for at least 80 h after the application of tamoxifen alone or in a combination with 17β-estradiol, and a concomitant inhibition of cytosol receptor replenishment was noted. Tamoxifen and 17β-estradiol, respectively, were found to stimulate progesterone receptor levels when applied through 5 days. Tamoxifen plus 17β-estradiol administration elevated progesterone receptor contents above those found for each of the two compounds alone. On the other hand, tamoxifen enhanced the 17β-estradiol-induced prolactin serum levels, but did not stimulate prolactin serum levels by itself. These data combine to suggest that tamoxifen interacts with common estrogen receptor sites at the rat anterior pituitary.     

Age-dependent induction and repression of rat liver microsomal hydroxylase systems by estradiol

The activities of the hepatic microsomal 2α-, 2β-, 18- and 7α-hydroxylase systems active on 5α-[4-^14C] androstane-3α,17β-diol were studied in male and female rats which had been castrated and spayed at 14, 24, 35 and 45 days of age, treated for 5 days with 500 μg of estradiol benzoate per kg body weight and killed 6 days after gonadectomy. The hepatic microsomal 15β-hydroxylase enzyme system active on 5α-[1,2-³H] androstane-3α, 17β-diol 3,17-disulphate was also measured in these animals as well as in an additional group of animals that were gonadectomized at 56 days of age, treated with estradiol benzoate and killed at 62 days of age. The hydroxylase systems active on the free steroid substrate were all relatively unresponsive towards enstradiol in the developing rat, in contrast to the strong effects on hepatic hydroxylase activities previously noted following treatment of adult male rats with estradiol. On the other hand, the 15β-hydroxylase system active on disulphurylated 5α-androstane-3α, 17β-diol was inducible in both male and female rats of 41 and 51 days of age and in male rats of 61 days of age.     

Relationships among mammalian gonadotropin-releasing hormone, prostaglandins, and sex steroids in the brain of the crested newt, Triturus carnifex

The present study was carried out to evaluate the in vitro brain release of prostaglandin F_2α (PGF_2α), prostaglandin E₂ (PGE₂), androgens, and 17β-estradiol in male and female crested newt, Triturus carnifex, during three different periods of the annual sexual cycle; in addition, the effects of mammalian gonadotropin-releasing hormone (mGnRH), PGF_2α, and PGE₂ on prostaglandins and steroids release by the brain were evaluated during the same periods. In brain incubations of both sexes, PGF_2α and estradiol were higher during postreproduction, while PGE₂ and androgens were higher during reproduction. In both sexes, mGnRH increased PGF_2α and estradiol during postreproduction, and PGE₂ during reproduction; PGF_2α increased estradiol secretion during postreproduction. Only in the male, did both mGnRH and PGE₂ increase androgens during reproduction. It could be suggested that in Triturus carnifex, the regulation of the reproductive activity in the central nervous system (CNS) depends on the relationships among mGnRH, prostaglandins and steroids. In particular, PGF_2α and PGE₂ seem to play different roles in the CNS of the newt: PGF_2α is involved in the postreproductive processes, through estradiol secretion, while PGE₂ in the reproductive ones (through androgens secretion?).     

Hormonal and developmental regulation of glycosylated alpha 2u-globulin synthesis

We have investigated the regulation of glycosylated α_2u-globulin synthesis by examining the appearance of these molecules in the medium of primary monolayer cultures of hepatocytes. Hepatocytes were isolated from male and female rats of various ages, as well as from castrated or ovariectomized animals. α_2u-Globulin was immunoprecipitated from the culture medium with rabbit antibody specific for α_2u-globulin, and the dissociated precipitates were electrophoresed on sodium dodecyl sulfate-polyacrylamide gels. We found that prepubescent male and female rats synthesized only the high molecular weight glycosylated forms of α_2u-globulin. Hepatocytes from 50-day-old intact and ovariectomized female rats, as well as from ovariectomized rats treated with 17β-estradiol, secreted only glycosylated α_2u-globulin. Hepatocytes from castrated male rats treated with dihydrotestosterone in vivo synthesized the 20,000-dalton nonglycosylated form of α_2u-globulin; the rate of glycosylated α_2u-globulin synthesis was reduced in these cells. The rate of synthesis of glycosylated α_2u-globulin by male rat hepatocytes declined concomitant with increases in the age of the rats, the level of serum testosterone, and the rate of synthesis of nonglycosylated α_2u-globulin. Under our conditions, dexamethasone administration to castrated male rats or ovariectomized female rats in vivo did not alter the species of α_2u-globulin that were synthesized subsequently by hepatocytes in vitro. Our results suggest that the synthesis of glycosylated α_2u-globulin is regulated differently than the synthesis of the 20,000-dalton nonglycosylated form of α_2u-globulin.     

Binding of estrogens by α-fetoprotein in rat amniotic fluid

Amniotic fluid from 15–17-day rat fetuses bound estrone and 17β-estradiol specifically. Related steroids such as estriol, 6-ketoestradiol, 17α-estradiol and testosterone were not bound to any significant extent. The apparent K_a for 17β-estradiol was 2.6·10⁸ M⁻ at 4°C; 6 nmoles of 17β-estradiol were bound per ml of amniotic fluid. The binding component appears to be α-fetoprotein in that it migrates as an α₁-globulin upon polyacrylamide gel electrophoresis and has an isoelectric pH of 4.7 as determined by isoelectric focusing. Furthermore, binding activity was precipitated by antiserum which was shown by immuno-electrophoresis to be specific for α-fetoprotein. Binding activity, partially purified by isoelectric focusing of amniotic fluid, was associated with one of two bands seen by polyacrylamide gel electrophoresis. This band migrated as an α₁-globulin.     

Synthesis and GABAA receptor activity of 2,19-sulfamoyl analogues of allopregnanolone

The synthesis of new analogues of allopregnanolone with a bridged sulfamidate ring over the β-face of ring A has been achieved from easily available precursors, using an intramolecular aziridination strategy. The methodology also allows the synthesis of 3α-substituted analogues such as the 3α-fluoro derivative. GABA_A receptor activity of the synthetic analogues was evaluated by assaying their effect on the binding of [³H]flunitrazepam and [³H]muscimol. The 3α-hydroxy-2,19-sulfamoyl analogue and its N-benzyl derivative were more active than allopregnanolone for stimulating binding of [³H]flunitrazepam. For the binding of [³H]muscimol, both synthetic analogues and allopregnanolone stimulated binding to a similar extent, with the N-benzyl derivative exhibiting a higher EC₅₀. The 3α-fluoro derivative was inactive in both assays.     

Differential effects of estrogen/androgen on the prevention of nonalcoholic fatty liver disease in the male rat

It is important to clarify the distinct contributions of estrogen/estrogen receptor (ER) and androgen/androgen receptor (AR) signaling and their reciprocal effects on the regulation of hepatic lipid homeostasis. We studied the molecular mechanisms underlying the preventive effects of estradiol (E2), dihydrotestosterone (DHT), or E2+DHT on high-fat diet-induced nonalcoholic fatty liver disease (NAFLD) in an orchidectomized Sprague-Dawley (SD) rat model. E2 is shown to be associated with decreased fatty acid synthesis in hepatic zone 3-specific manner by increasing the phosphorylation of acetyl coenzyme-A carboxylase via an ERα-mediated pathway. DHT is shown to be associated with decreased lipid accumulation and cholesterol synthesis in a hepatic zone 1-specific manner by increasing expression of carnitine palmitotyltransferase1 and phosphorylation of 3-hydroxy-3-methyl-glutaryl-CoA reductase via an AR-mediated pathway. E2+DHT showed an additive positive effect and normalized all three impaired zones of the liver. Gene expression changes in human severe liver steatosis were similar to those of experimental rat NAFLD. Steroids reversed the histopathological NAFLD changes, likely by decreasing fatty acid and cholesterol synthesis and increasing β-oxidation. The diverse steroid effects (ER/AR) on NAFLD prevention in male rats indicate the potential applicability of ER/AR modulators for NAFLD treatment.     

Lipid rafts constrain basal α1A-adrenergic receptor signaling by maintaining receptor in an inactive conformation

We have reported that the α_1A-adrenergic receptor (α_1AAR) in rat-1 fibroblasts is a lipid raft protein. Here we examined whether disrupting lipid rafts by methyl-β-cyclodextrin (MCD) sequestration of cholesterol affects α_1AAR signaling. Unexpectedly, MCD increased α_1AAR-dependent basal inositol phosphate formation and p38 mitogen-activated protein kinase activation in a cholesterol-dependent manner. It also initiated internalization of surface α_1AAR, which was partially blocked by receptor inhibition. Binding assays revealed MCD-mediated increases in receptor agonist affinity as well as reciprocal decreases in inverse agonist affinity, a behavior that is usually interpreted as a shift toward the active receptor conformation. In untreated cells a fraction of the receptor was found to be present in preassociated receptor/G protein complexes, which rapidly dissociate upon receptor stimulation. Consistent with MCD-induced signaling, raft disruption resulted in an increase in receptor/G protein complexes. These results strongly suggest that lipid rafts constrain basal α_1AAR activity; however, preassembled receptor/G protein complexes could still provide a mechanism for accelerating α_1AAR signaling following stimulation.     

Binding of [3H]methyltrienolone to androgen receptor in rat liver

The synthetic androgen methyltrienolone is superior to testosterone and androstenedione for the measurement of androgen receptor in tissues where the native ligands are metabolized into inactive derivatives. [³H]Methyltrienolone binds with a high affinity to androgen receptor in cytosol prepared from male rat livers, as the Scatchard analysis revealed that the K_d value was 3.3 · 10^?8 M and the number of binding sites was 35.5 fmol/mg protein. Since methyltrienolone also binds glucocorticoid receptor which exists in rat liver, the apparent binding of androgen receptor is faulty when measured in the presence of glucocorticoid receptor. The binding of methyltrienolone to glucocorticoid receptor can be blocked by the presence of a 100-fold molar excess of unlabeled synthetic glucocorticoid, triamcinolone acetonide, without interfering in its binding to androgen receptor, because triamcinolone does not bind to androgen receptor. Triamcinolone-blocked cytosol exhibited that the K_d value was 2.5 · 10^?8 M and the number of binding sites was 26.3 fmol/mg protein, indicating a reduction to $\text{3}\text{4}$ of that in the untreated cytosol. The profile of glycerol gradient centrifiguration indicated that [³H]methyltriemolone-bound receptor migrated in the 8–9 S region in both untreated and triamcinolone-blocked cytosols, but the 8–9 S peak in triamcinolone-blocked cytosol was reduced to about $\text{3}\text{4}$ of that of untreated cytosol.     

Maraviroc Attenuates Trauma-Hemorrhage-Induced Hepatic Injury through PPAR Gamma-Dependent Pathway in Rats

Maraviroc is a CC-chemokine receptor 5 (CCR5) antagonist with potent antiviral and cancer preventive effects. Recent evidence suggests that the co-existence of CCR5 in various cell types is involved in inflammation. However, the effects that CCR5 antagonists produce in trauma-hemorrhage remain unknown. The peroxisome proliferator-activated receptor gamma (PPAR_γ) pathway exerts anti-inflammatory effects in injury. In this study, we hypothesized that maraviroc administration in male rats, after trauma-hemorrhage, decreases cytokine production and protects against hepatic injury through a PPAR_γ-dependent pathway. Male Sprague-Dawley rats underwent trauma-hemorrhage (mean blood pressure maintained at approximately 35-40 mmHg for 90 minutes), followed by fluid resuscitation. During resuscitation, a single dose of maraviroc (3 mg/kg, intravenously) with and without a PPAR_γ antagonist GW9662 (1 mg/kg, intravenously), GW9662 or vehicle was administered. Plasma alanine aminotransferase (ALT) with aspartate aminotransferase (AST) concentrations and various hepatic parameters were measured (n=8 rats/group) at 24 hours after resuscitation. The results showed that trauma-hemorrhage increased hepatic myeloperoxidase activity, intercellular adhesion molecule-1 and interleukin-6 levels, and plasma ALT and AST concentrations. These parameters were significantly improved in the maraviroc-treated rats subjected to trauma-hemorrhage. Maraviroc treatment also increased hepatic PPAR_γ expression compared with vehicle-treated trauma-hemorrhaged rats. Co-administration of GW9662 with maraviroc abolished the maraviroc-induced beneficial effects on the above parameters and hepatic injury. These results suggest that the protective effect of maraviroc administration on alleviation of hepatic injury after trauma-hemorrhage, which is, at least in part, through PPAR_γ-dependent pathway.     

Antagonism of the estradiol-mediated repression of microsomal 3β-hydroxysteroid dehydrogenase activity in rat liver by antiestrogenic substances

5α-Dihydrotestosterone, 17-hydroxyprogesterone caproate, 2-methoxyestrone and a number of nonsteroidal antiestrogens (clomiphene citrate, nafoxidine hydrochloride, tamoxifen, MER-25) were tested for their ability to block estradiol-mediated repression of the androgen-dependent 3β-hydroxy-steroid dehydrogenase activity of male rat liver. With the exception of 5α-dihydrotestosterone, which induced activity in females, none of these substances affected 3β-hydroxy-steroid dehydrogenase activity when administered alone to otherwise untreated male and female rats. Tamoxifen (100 or 500 μg/day) was the only substance which prevented a decrease in enzyme activity when given simultaneously with estradiol (5 μg/day). The estradiol-mediated decrease in activity was not antagonized by a 100-fold higher dose of androgen (5α-dihydrotestosterone, 0.5 mg/day), demonstrating the potent antiandrogenic effect of estradiol on this hepatic androgen-dependent enzyme activity.     

4-Methyl-3-oxo-4-aza-5α-androst-1-ene-17β-N-aryl-carboxamides: an approach to combined androgen blockade [5α-reductase inhibition with androgen receptor binding in vitro]

4-Aza-5α-androstan-3-one 17β-(N-substituted carboxamides) are potent human type 2 5α-reductase (5aR) inhibitors with generally poor binding to the human androgen receptor (hAR). When the 17-amide N-substituent included an aromatic residue, potent dual inhibitors of both type 1 and 2 5aR are produced, but hAR binding remained poor. Tertiary-substituted-17-amides have reduced inhibition of both 5aR isozymes. The addition of an N⁴-methyl substitutent to the A-ring profoundly increased hAR affinity and the addition of unsaturation to the A-ring (Δ¹) modestly augmented hAR binding. The unsubstituted carbanilides in the Δ¹-N⁴-methyl series show some selectivity for type 1 5aR over the type 2 isozyme, whereas addition of aryl substituents, particularly at the 2-position, increased type 2 5aR binding to provide dual inhibitors with excellent hAR binding, e.g. N-(2-chlorophenyl)-3-oxo-4-methyl-4-aza-5α-androst-1-ene-17β-carboxamide (9c). Compounds of this type exhibit low nanomolar IC₅₀s for both human 5aR isozymes as well as the human androgen receptor. Kinetic analysis confirms that the prototype 9c displays reversible, competitive inhibition of both human isozymes of 5aR with K_i values of less than 10 nM. Furthermore, this compound binds to the androgen receptor with an IC₅₀ equal to 8 nM. Compounds in this series are projected to be powerful antagonists of testosterone and dihydrotestosterone action in vivo, with potential utility in the treatment of prostatic carcinoma (PC).     

Genomic organization of the rat α2u-globulin gene cluster

The α_2u-globulins are a group of similar proteins, belonging to the lipocalin superfamily of proteins, that are synthesized in a subset of secretory tissues in rats. The many α_2u-globulin isoforms are encoded by a multigene family that exhibits extensive homology. Despite a high degree of sequence identity, individual family members show diverse expression patterns involving complex hormonal, tissue-specific, and developmental regulation. Analysis suggests that there are approximately 20 α_2u-globulin genes in the rat genome. We have used fluorescence in situ hybridization (FISH) to show that the α_2u-globulin genes are clustered at a single site on rat Chromosome (Chr) 5 (5q22-24). Southern blots of rat genomic DNA separated by pulsed field gel electrophoresis indicated that the α_2u-globulin genes are contained on two NruI fragments with a total size of 880 kbp. Analysis of three P1 clones containing α_2u-globulin genes indicated that the α_2u-globulin genes are tandemly arranged in a head-to-tail fashion. The organization of the α_2u-globulin genes in the rat as a tandem array of single genes differs from the homologous major urinary protein genes in the mouse, which are organized as tandem arrays of divergently oriented gene pairs. The structure of these gene clusters may have consequences for the proposed function, as a pheromone transporter, for the protein products encoded by these genes. Received: 12 August 1998 / Accepted: 13 January 1999     

Sexual experience changes sex hormones but not hypothalamic steroid hormone receptor expression in young and middle-aged male rats

Testosterone is well known to regulate sexual behavior in males, but this is dependent upon prior sexual experience. Aging is associated with decreased libido and changes in testosterone, but the role of experience in these age-related processes has not been systematically studied. We examined effects of age and sexual experience on serum hormones (total testosterone, free testosterone, estradiol, LH) and on numbers of androgen receptor (AR) and estrogen receptor α (ERα) immunoreactive cells in the hypothalamus. Extensive sexual experience was given to male rats at 4 months of age. Rats were euthanized at either 4 months (young) or 12 months (middle-aged (MA)). Comparable sexually naïve male rats were handled and placed into the testing arena but did not receive any sexual experience. Thus, we had four groups: young-naïve, young-experienced, MA-naïve and MA-experienced. Serum hormone levels were assayed, and numbers of AR and ERα cells were quantified stereologically in the medial preoptic nucleus (MPN) and the anteroventral periventricular nucleus (AVPV). Sexually experienced males had significantly elevated serum testosterone and free testosterone in both age groups. Both total and free testosterone were higher, and estradiol lower, in middle-aged than young rats. Experience did not alter either AR or ERα expression in the preoptic brain regions studied. Aging was associated with increased expression of AR, but no change in ERα. These results show that sexual experience can induce short-term and long-term alterations in serum hormones but these effects are not manifested upon their receptors in the hypothalamus.     

Effect of maternal fatness on fetal steroids and semi-quantitative real-time PCR expression of receptor genes in sheep

Sexual differentiation of the brain occurs between d 30 and 70 in the fetal lamb. The objective of this experiment was to determine if maternal fatness affects fetal steroid production and expression of their receptors which may ultimately alter endocrine systems postnatally. Fetuses were collected from ewes fed at either 100% (Control; n = 5) or 150% (Fat; n = 6) of NRC recommendations from 60 d prior to breeding until collection at 75 d of gestation. Hypothalamic and amygdala neural tissues were collected from twin male/female fetuses. Serum concentrations of testosterone were greater (P < 0.001) in male fetuses compared to female fetuses. Further, male fetuses from Fat ewes had greater (P < 0.05) serum concentrations of testosterone than male fetuses from Control ewes, but differences in testicular steroidogenic enzyme mRNA were not detected (P = 0.18). Quantity of hypothalamic mRNA for estrogen receptor (ER) β tended (P = 0.1) to be influenced by a sex by treatment interaction. Messenger RNA for ER-β was greater in female fetuses than male fetuses from Control ewes (P = 0.05). Although amount of ER-β mRNA did not differ among male fetuses (P = 0.7), amounts tended to be less (P = 0.07) in female fetuses from Fat ewes compared to those from Control ewes, and did not differ (P ≥ 0.8) from male fetuses. Hypothalamic ER-α mRNA tended (P = 0.1) to be less in fetuses from Fat ewes compared to Control fetuses but was not influenced (P = 0.3) by fetal sex or their interaction. Amount of mRNA for hypothalamic progesterone receptor tended (P = 0.06) to be greater in male fetuses than female fetuses and tended to be less (P = 0.06) in fetuses from Fat ewes than in Control fetuses, but did not differ by any sex by treatment interaction (P = 0.6). Hypothalamic RNA for the androgen receptor did not differ by sex, dam nutritional treatment, or the interaction. Likewise, amygdala RNA for the estrogen or androgen receptor did not differ (P ≥ 0.3) by sex, treatment, or their interaction. Dam fatness appears to decrease the expression of progesterone receptor, ER-α, and decrease amount of ER-β in the female fetuses while increasing circulating concentrations of testosterone in male fetuses. Altered expression of hypothalamic receptor genes by the uterine environment may affect adult responses to stress, sexual behavior and/or the pattern of gonadotropin release in response to gonadal steroids.     

Age and sex dependence of the effects of an aqueous extract of Physalis alkekengi fruits on rat hepatic glucose 6-P dehydrogenase activity

Intraperitoneal injections of an aqueous extract of winter cherry fruits (Physalis alkekengi) to new-born, weanling and adult female rats and to weanling and adult male rats had no effect on body weight, liver weight and liver cytosol protein content. The specific activities of hepatic glucose 6-P dehydrogenase (an estrogen induced protein) in rats of different age and sex groups in terms of mU/mg protein were: treated new-born females, 15.9 ± 0.5; control, 29.1 ± 0.6; treated weanling females, 14.9 ± 0.3; control, 24.8 ± 0.7; treated adult females, 25.7 ± 0.5; control, 26.1 ± 0.5; treated weanling males, 7.9 ± 0.2; control, 7.9 ± 0.1; treated adult males, 9.6 ± 0.4; and control, 9.7 ± 0.3. Treatment of new-born and weanling female rats with the extract resulted in 40–45% reduction in hepatic G6PD activity. However, treatment of adult females, and weanling and adult males produced no significant change in the activity of this enzyme. The data are discussed both in terms of the increase in the capacity of rodent liver to metabolize steroidal compounds with age and the presence of low levels of circulating estradiol necessary for enzyme induction in male rats.     

Tissue- and temporal-specific regulation of 11β-hydroxysteroid dehydrogenase type 1 by glucocorticoids in vivo

11β-hydroxysteroid dehydrogenase type 1 (11β-HSD-1) catalyses the interconversion of active corticosterone and inert 11-dehydrocorticosterone. Short-term glucocorticoid excess upregulates 11β-HSD-1 in liver and hippocampus leading to suggestions that 11β-HSD-1 ameliorates the deleterious effects of glucocorticoid excess by its 11β-dehydrogenase activity. However the predominant activity of 11β-HSD-1 in vivo is 11β-reduction, thus generating active glucocorticoid. We have re-examined the time-course of glucocorticoid regulation of 11β-HSD-1 in the liver, hippocampus and kidney of adult male rats in vivo.Sham operation markedly reduced 11β-HSD-1 mRNA expression in all tissues, and reduced 11β-HSD bioactivity in liver and hippocampus when compared to untouched controls. Adrenalectomy reduced 11β-HSD-1 expression in all tissues in the short-term (7 days), followed by subsequent recovery of enzyme activity by 21 days in liver and hippocampus. Dexamethasone replacement of adrenalectomised rats attenuated the initial decrease in hepatic 11β-HSD-1 activity, but by 21 days dexamethasone reduced activity compared to control levels.Thus glucocorticoids regulate 11β-HSD-1 in a complex tissue- and temporal-specific manner. This pattern of regulation suggests glucocorticoids repress 11β-HSD-1 at least in the liver, a pattern of regulation more consistent with the evidence that 11β-HSD-1 is an 11β-reductase in vivo. Operational stress per se down-regulates 11β-HSD-1 which has implications for interpretation and design of in vivo studies of 11β-HSD-1.