Androgenic properties and adrenal depressant activity of megestrol acetate observed in castrated male rats.

Megestrol acetate (17alpha-acetoxy-6-dehydro-6-methylprogesterone), a synthetic steroid with high progestational activity, is used in oral contraceptives but also in the treatment of prostatic diseases in man. To investigate whether megestrol acetate has any androgenic properties the growth of the ventral and dorsolateral prostate, the coagulating glands and the seminal vesicles was studied morphologically in castrated rats treated with megestrol acetate and in non-treated castrated rats. The effect of megestrol acetate on the body weight, the levator ani muscle and the adrenals was also studied. Megestrol acetate was administered in daily doses of 0.02 mg, 0.2 mg, 2.0 mg or 20.0 mg for a period of 21 days. Megestrol acetate in the two higher doses retarded growth and gave a low weight for the levator ani muscle at autopsy indicating an anti-anabolic or catabolic action of megestrol acetate in high doses. Megestrol acetate in daily doses of 0.2, 2.0 and 20.0 mg caused an involution of the adrenal glands. After the two higher doses the weight of the adrenals amounted to only about a third of that of the untreated rats. Megestrol acetate in the lower doses had no demonstrable effect on the growth of the accessory reproductive glands. After the two higher doses of megestrol acetate some growth of the dorsal part of the dorsolateral prostate and of the coagulating glands was observed. Only the seminal vesicles exhibited complete morphological criteria of an androgenic stimulation and then only after the largest dose of megestrol acetate. The investigation shows that megesterol acetate has weak androgenic properties which are apparent at a dose per kg body weight approximately 200 times greater than that used in the treatment of prostatic diseases in man.     

The uptake of [3H]testosterone and its metabolites by the brain and pituitary gland of the fetal macaque

Testosterone is secreted by the fetal testis during gestation, and this is thought to influence certain aspects of the brain's subsequent development. To study this action at the neuronal level, nine macaque fetuses were injected with 250 microCi [3H]testosterone via the umbilical vein at about 120 days gestation. After 60 min, samples of brain and peripheral tissue were studied by autoradiography or HPLC. Purified nuclear pellets were prepared, and radioactivity in ether extracts was fractionated by HPLC and identified by coelution with internal standard steroids. Concentrations of radioactivity were significantly higher (P less than 0.05) in the hypothalamus-preoptic area than in amygdala, hippocampus, midbrain, and cerebral and cerebellar cortexes, and most of the radioactivity (75%) in the hypothalamus-preoptic area coeluted with 17 beta-estradiol. Radioactivity coeluting with 17 beta-estradiol was also detected in nuclear fractions from amygdala (44%). In contrast, 80% of the radioactivity extracted from pituitary gland nuclei coeluted with testosterone. Most of the neurons labeled in autoradiograms were located in the hypothalamus and preoptic area, fewer were found in the amygdala, and labeling in the frontal or motor cortex did not exceed chance levels. Results suggested that aromatization and, consequently, estrogen receptors play a role in the effects of testosterone on the hypothalamus and amygdala of the primate fetus at this stage of development.     

Effects of sex steroid hormones on arginine vasopressin in intact and castrated male and female rats

In the present study we have examined the effects of androgens and estrogens on circulating arginine vasopressin (AVP). Adult male Wistar rats had serum AVP levels of 0.4 microU/ml. Two weeks after bilateral castration, AVP rose to 2.6 microU/ml, but daily testosterone administration (100 microgram/100 g BW) to the castrate males prevented the AVP increase (0.8 microU/ml). During a normal estrous cycle, adult female Wistar rats had AVP values of 0.6 microU/ml during diestrus, 4.6 microU/ml on the morning of proestrus, 1.3 microU/ml on the afternoon or proestrus, and 1.5 microU/ml on the day of estrus. These changes in AVP paralleled the presumed changes in serum estradiol. Two weeks after bilateral ovariectomy of the adult female rats, AVP was 1.4 microU/ml but daily estradiol injections (100 microgram/100 g BW) to the castrate females produced a rise of serum AVP to 5.0 microU/ml. The results suggest an androgen inhibition and an estrogen stimulation of serum AVP levels.     

Formation of steroids by the pregnant mare. VI. Metabolism of [14C]farnesyl pyrophosphate and [3H]dehydroepiandrosterone injected into the fetus.

A mixture of [4,8,12-14C]farnesyl pyrophosphate and [3H]dehydroepiandrosterone was injected into a horse fetus im during laparotomy, after which maternal urine was collected for 6 days. Steroid conjugates in the urine were extracted with Amberlite XAD-2 resin, hydrolyzed, and separated into phenolic and neutral fractions. Estrone, 17 alpha-estradiol, equilin [3-hydroxy-1,3,5(10),7-estratetraen-17-one], and 17 alpha-dihydroequilin [1,3,4(10),7-estratetraene-3,17 alpha-diol] were isolated from the phenolic fraction and their radiochemical purities were established. Only estrone and 17 alpha-estradiol contained both 3H and 14C, while the B ring unsaturated estrogens, equilin and 17 alpha-dihydroequilin, contained only 14C. From the neutral fraction, 14C-labeled 3 beta-hydroxy-5 alpha-pregnan-20-one, 5 alpha-pregnane-3 beta-20 beta-diol, and 5 alpha-pregnane-3 beta, 20 alpha-diol were isolated. These results together with our previous findings demonstrate that the route of biosynthesis of both the ring B saturated and unsaturated estrogens is the same up to the stage of farnesyl pyrophosphate. Thus, the bifurcation in the classical pathway of steroid biosynthesis is occurring at a point after the formation of farnesyl pyrophosphate and before the formation of squalene and cholesterol.     

Influence of steroid hormone treatments or pregnancy on [3H]prazosin and [3H]rauwolscine binding to myometrial alpha-adrenoceptors in the ewe.

The adrenergic antagonists [3H]prazosin and [3H] rauwolscine were used to identify alpha 1- and alpha 2-adrenoceptors respectively in the ovine myometrium. Ewes were allocated to four groups according to steroid hormone treatments or physiological status, namely ovariectomized ewes either as untreated controls, treated with oestradiol-17 beta or progestagen plus oestradiol-17 beta, and pregnant ewes at mid-gestation. Binding of both [3H]prazosin and [3H]rauwolscine to membrane preparations from the ovine myometrium was saturable, of high affinity and rapidly reversed by phentolamine (10 mumol/l). Based on the relative order of potency of selected adrenergic agonists and antagonists, the myometrial binding sites labelled by [3H]prazosin and [3H]rauwolscine were characterized as alpha 1- and alpha 2-adrenoceptors respectively. Saturation binding studies with [3H]prazosin showed that the number of alpha 1-adrenoceptors was low (maximal binding capacity, Bmax, between 19 and 24 fmol/mg protein) and there were no noticeable differences between the animal groups. Moreover, the equilibrium dissociation constant (Kd) did not vary significantly between groups (Kd between 0.10 and 0.17 nmol/l). In contrast, saturation binding studies with [3H]rauwolscine revealed the presence of a high number of alpha 2-adrenoceptors. Values of Bmax were far higher in the pregnant ewes (1096 +/- 241 fmol/mg protein; means +/- S.D.) than in any of the non-pregnant ovariectomized ewes. For these latter groups, the highest Bmax values were found in the group treated with both progestagen and oestrogen (382 +/- 77 fmol/mg protein) compared with treatment with oestrogen alone (101 +/- 8 fmol/mg protein) or with controls (82 +/- 12 fmol/mg protein.(ABSTRACT TRUNCATED AT 250 WORDS)     

Comparisons of the nuclear uptake of [3H]-testosterone and its metabolites by the brains of male and female macaque fetuses at 122 days of gestation

Testosterone secreted by the testis of the macaque fetus is thought to influence certain aspects of the brain's subsequent development which may be responsible for the ontogeny of sexually dimorphic patterns of behavior. To compare the interactions between testosterone and the receptors for androgens and estrogens in brain cell nuclei in the two sexes, 7 intact female fetuses and 5 intact male fetuses were injected in utero at about 120 days of gestation with [3H]-testosterone (250 microCi i.v. or 500 microCi s.c.). One hour later, fetuses were delivered by cesarean section, and samples of brain and peripheral tissues were homogenized and separated into purified nuclear and supernatant fractions. Fractions were analyzed by high performance liquid chromatography to measure levels of [3H]-testosterone and its metabolites. Concentrations of radioactivity extracted from cell nuclei were significantly higher in the hypothalamus-preoptic area than in other brain areas (p less than 0.001); [3H]-estradiol represented 65.0 +/- 5.7% of this radioactivity and nuclear concentrations of this metabolite were 73% lower in males than in females (p less than 0.001). Nuclear concentrations of [3H]-testosterone in the pituitary gland (68.9 +/- 8.8% of extracted radioactivity) were 48% lower in males than in females (p less than 0.001). There was no evidence of a sex difference in the tissue uptake of radioactive steroids from blood, but in males, levels of endogenous plasma testosterone (599.8 +/- 208.2 ng/100 ml) were significantly higher than in females (37.7 +/- 28.5 ng/100 ml; p less than 0.01), and the specific activity of [3H]-testosterone in blood was consequently lower in males than in females.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of dopamine agonists and antagonists on the binding of [3H]estradiol to its receptors in the anterior pituitary gland of male rats

The synthesis of PRL and DNA in PRL cells is regulated by estrogens and dopamine. To investigate a possible relationship between these two components, we studied the influence of dopamine agonists and antagonists on the binding of [3H]estradiol ([3H]E2) to its receptors in the anterior pituitary gland of estrogenized male rats. The administration of sulpiride (dopamine antagonist) or bromocriptine (dopamine agonist) decreased the binding of [3H]E2 to cytosolic receptors when the concentration of [3H]E2 in the assay mixture was 1 nM. Both drugs also diminished the binding of [3H]E2 when they were added in vitro to the incubation media, apparently in a competitive way. Dopamine and alpha-methyltyrosine also inhibited competitively the binding of [3H]E2 to cytosolic receptors. The inhibition constants were determined by the Lineweaver-Burk plot. To overcome the competitive inhibition of dopamine agonists and antagonists, the concentration of the titrated steroid in the incubation mixture was increased to 16 nM. This concentration was established by saturation analysis. The administration of alpha-methyltyrosine increased the binding of [3H]E2 to nuclear receptors without modifying the binding to cytosolic receptors. This increase paralleled an increment in the levels of plasma PRL. Bromocriptine prevented the increase in [3H]E2 binding produced by alpha-methyltyrosine and had no effect on the binding when administered to nontreated rats. These results suggest that dopamine can regulate the biological effects of estradiol in the anterior pituitary gland by decreasing the binding of this hormone to its receptors.     

Studies on the feedback regulation of gonadotropin concentrations in male rats (I). The effects of hypothalamic implantation of testosterone and its metabolites on gonadotropin release in castrated male rats (author's transl)

Testosterone (T), 5 alpha-dihydrotestosterone (5 alpha-DHT), 5 alpha-androstane-3 alpha, 17 beta-diol (3 alpha-diol) and estradiol-17 beta (E) or cholesterol (control) were implanted into the hypothalamic median eminence--arcuate nucleus (ME-ARC) region of long-term castrated male rats. Serum LH and FSH levels were measured before and after the implantation. T or E implantation resulted in a 65.5% (P less than 0.01) or 80.2% (P less than 0.01) decrease in serum LH levels respectively, within 7 days after the intrahypothalamic application. 5 alpha-DHT or 3 alpha-diol implantation in the ME-ARC region also resulted in a significant lowering of serum LH levels, although their inhibitory action was less effective than that of T or E. There were no significant differences in serum LH levels 20 min after the intraperitoneal injection of 2 micrograms/kg of LH-RH between the control group and the T, 5 alpha-DHT, 3 alpha-diol or E group. E implantation also resulted in a 34.7% (P less than 0.01) decrease in serum FSH levels at the 7th day, but other steroids failed to reduce serum FSH levels after the intrahypothalamic application. These studies suggest that aromatization of T to E is not indispensable to the negative feedback effect of androgen on the hypothalamic LH-RH release. It might also be supposed that E is one of the factors regulating the serum FSH level.     

Role of the adrenal cortex in chronic stress-induced inhibition of prolactin secretion in male rats

The response of prolactin to chronic stress in intact, adrenalectomized and adrenomedullectomized male rats was studied. Immobilization stress in intact animals induced a significant increase in plasma concentrations of prolactin after 20 and 45 min and a significant decrease when the rats were submitted to chronic restraint (6 h daily for 4 days). Five weeks after adrenomedullectomy, plasma prolactin and corticosterone responses to chronic stress were not modified. In contrast, the inhibitory effect of chronic stress on prolactin secretion was totally suppressed by adrenalectomy. When treated with dexamethasone during the 4 days of restraint, adrenalectomized stressed rats showed similar plasma concentrations of prolactin to the intact stressed rats. These data indicate that the adrenal cortex is able to play an inhibitory role on prolactin secretion during stress only through a prolonged release of glucocorticoids.