The red cell membrane and its cytoskeleton.

Gel-filtration (Sephadex G-75) analysis of hepatic cytosol reveals both qualitative and quantitative sex differences in oestrogen-binding proteins. The elution profile of [³H]oestradiol-labelled cytosol shows four species of oestrogen-binding proteins (peaks I, II, IV and V) common to both sexes. The amount of [³H]oestradiol binding in peak I is equivalent in both males and females and corresponds quantitatively to the specific oestrogen receptor. The amount of binding in the remaining three peaks is greater in males than females. In addition, an oestrogen-binding protein (peak III) is present that is unique to male cytosol. Proteinase-inhibition studies demonstrate that the observed multiplicity of oestrogen-binding proteins is not an artefact of proteolytic breakdown. Sex differences in oestrogen-binding proteins are absent in immature male and female animals; the oestrogen-binding protein profile in immature rats resembles that of an adult female. Gonadectomy of adult animals does not affect the oestrogen-binding-protein profile. In contrast, neonatal (day 1) castration results in partial feminization of the characteristic oestrogen-binding protein profile seen in the adult male; the appearance of Peak III is suppressed and marked decreases in the amount of oestradiol binding occurs in the remaining peaks. Hypophysectomy of adult animals results in near abolishment of the observed sex differences; the male oestrogen-binding protein profile is partially feminized and the female profile is partially masculinized, as characterized by the appearance of [³H]oestradiol binding in the region of peak III and increased amounts of binding in peaks IV and V. The present studies demonstrate a multiplicity of oestrogen-binding proteins in liver cytosol and raise the possibility that the presence of some of these proteins may be imprinted at birth through the hypothalamic–pituitary axis, by a mechanism requiring neonatal androgen exposure.     

Evidence for androgen independence of male mounting behavior in white-crowned sparrows (Zonotrichia leucophrys gambelii)

Previous experiments have shown that expression of mounting behavior in sexually inexperienced, adult male white-crowned sparrows does not require elevated plasma levels of androgen; adult males maintained on nonstimulatory short days mount sexually receptive females. The experiments reported here demonstrate that (1) sexually inexperienced, prepubertal males maintained on nonstimulatory short days show very low mounting rates in response to female sexual displays; (2) these males exhibit high mounting rates when exposed to stimulatory long days but androgen treatment on short days is ineffective in stimulating high mounting rates; and (3) prepubertal castration has no effect on the expression of mounting behavior by photostimulated adult males. Thus, there is no evidence that mounting behavior of adult male white-crowned sparrows depends on androgen.     

Selective actions of growth hormone on rat liver estrogen binding proteins

Levels of hepatic estrogen receptor were 9.0 ± 2.4 fmoles/mg cytosol protein in intact females compared to 3.4 ± 2.2 in hypophysectomized females. Likewise, levels of receptor were 9.8 ± 1.5 fmoles/mg cytosol protein in intact males and 2.7 ± 1.8 in hypophysectomized males. Hypophysectomy abolished the sex differences in a second class of binding sites termed higher capacity lower affinity binding sites by increasing female levels and decreasing male levels. Treatment of hypophysectomized male or female rats with growth hormone (2 units/kg body wt, two times daily) restored normal levels of hepatic estrogen receptor. Administration of growth hormone to hypophysectomized rats did not reverse the effects of hypophysectomy on higher capacity lower affinity binding sites. These studies demonstrate that growth hormone exerts selective actions on different forms of hepatic estrogen binding proteins.     

Androgen receptor in genital tubercle of rabbit fetuses and newborns. Ontogeny and properties

In newborn rabbits of both sexes, an androgen receptor was characterized in the genital tubercle. Homogenates exhibited high affinity (Kd was about 0.4 nM) and saturable binding of [3H]methyltrienolone. The half-life of the [3H]5 alpha-dihydrotestosterone-androgen receptor complex was 72 h at 4 degrees C. The receptor was inactivated by heat and pronase and the binding was specific for potent androgens. Sucrose gradient analysis revealed a 8-9 S [3H]methyltrienolone binding protein in cytosols from both sexes. Androgen binding, in the homogenate, was detected as soon as day 18 of gestation in both sexes and the number of binding sites increased until birth. During sexual organogenesis and at birth there were no major differences between males and females in the amount or affinity of androgen binding. Specific androgen binding was also detected in sexual ducts of male and female newborns.     

Neonatal hormonal influences on the development of proceptive and receptive feminine sexual behavior in rats

The objective of this study was to examine the influence of androgen and of the inhibiting of aromatization of androgen to estrogen during the early neonatal period on the development of receptive (lordosis and acceptance of stimulus male mounting attempts) and proceptive (affiliation with and solicitation of stimulus males) feminine sexual behavior. Within 8 hr of birth, male rats were castrated or received subcutaneous implants of the aromatase inhibitor androst-1,4,6-triene-3, 17-dione (ATD) while females received injections of testosterone propionate (TP). At 90 days of age all treated animals and controls were tested for receptive and proceptive feminine sexual behavior. It was found that androgen present neonatally blocked proceptive as well as receptive behavior patterns in adult rats. The proceptive and receptive feminine sexual behavior patterns displayed by adult males deprived of the effects of androgen neonatally either by castration or by treatment with ATD were comparable to those of normal females.     

Diazepam: endocrine effects and hypothalamic binding sites in the developing male and female rat

The ontogeny of diazepam's endocrine effects in male and female rats, and of 3H-diazepam binding in the hypothalami of both sexes was studied. Diazepam inhibited basal prolactin levels in 38 day-old male rats and, if prolactin levels were stimulated by Haloperidol the inhibition occurred in 28 day-old males, indicating that the hypoprolactinemic effect of the drug could be evidenced earlier if prolactin titers were high. The prolactin inhibition in females did not reach statistical significance at any studied age. Diazepam significantly released LH only in male rats at 12 days, showing thus, a period of special sensitivity of LH release to the drug. Benzodiazepine-hypothalamic binding sites increased in number from birth to puberty, reaching a plateau at 20 days of age. No sexual differences or changes in affinity were found throughout the studied period. These results suggest that the maturation of diazepam's hypoprolactinemic effect could be partially related to the increase in hypothalamic binding sites, whereas the sexual differences observed in diazepam's endocrine actions could be due to sexual differentiation of endocrine control mechanisms.     

Early androgen treatment and male and female sexual behavior in mice

To investigate the role of neonatal androgen stimulation in the development of the potential for masculine and feminine sexual behavior in the mouse, different groups of mice were hormonally manipulated early in life. One group of female mice was administered testosterone propionate (TP) within 24 hr of birth; a second group of females was given a control injection of oil on the day of birth; a third group of females received an injection of TP on the 10th day after birth. A group of males received a control injection of oil on the day of birth. All mice were gonadectomized at about 30 days of age. At 60 days of age, mice were injected with estrogen and progesterone and tested for sexual receptivity; several weeks later all mice were injected with TP and tested for male sexual behavior. Female behavior: Females given oil at birth and females given TP on the 10th day after birth showed high levels of sexual receptivity as adults following estrogen-progesterone treatment. Females given TP on the day of birth, and male mice, rarely exhibited lordosis following estrogen-progesterone treatment. Male behavior: Most mice, regardless of genetic sex or neonatal treatment, mounted in adulthood following administration of exogenous androgen. There was little difference in mounting frequency between groups, suggesting that exogenous or endogenous androgen stimulation of the neonatal mouse does not facilitate adult mounting behavior. These data for the mouse are in essential agreement with existing data for the rat, and indicate that sexual behavioral differentiation induced by androgen stimulation in infancy is best characterized as an inhibition of the potential to display feminine sexual behavior in adulthood.     

Specific morphogenetic events in mouse external genitalia sex differentiation are responsive/dependent upon androgens and/or estrogens

The objective of this study was to perform a comprehensive morphologic analysis of developing mouse external genitalia (ExG) and to determine specific sexual differentiation features that are responsive to androgens or estrogens. To eliminate sex steroid signaling postnatally, male and female mice were gonadectomized on the day of birth, and then injected intraperitoneally every other day with DES (200ng/g), DHT (1μg/g), or oil. On day-10 postnatal male and female ExG were dissected, fixed, embedded, serially sectioned and analyzed. We identified 10 sexually dimorphic anatomical features indicative of normal penile and clitoral differentiation in intact mice. Several (but not all) penile features were impaired or abolished as a result of neonatal castration. Those penile features remaining after neonatal castration were completely abolished with attendant clitoral development in androgen receptor (AR) mutant male mice (X(Tfm)/Y and X/Y AR-null) in which AR signaling is absent both pre- and postnatally. Administration of DHT to neonatally castrated males restored development of all 10 masculine features to almost normal levels. Neonatal ovariectomy of female mice had little effect on clitoral development, whereas treatment of ovariectomized female mice with DHT induced partial masculinization of the clitoris. Administration of DES to neonatally gonadectomized male and female mice elicited a spectrum of development abnormalities. These studies demonstrate that the presence or absence of androgen prenatally specifies penile versus clitoral identity. Differentiated penile features emerge postnatally and are sensitive to and dependent upon prenatal or pre- and postnatal androgen. Emergence of differentiated clitoral features occurs postnatally in either intact or ovariectomized females. It is likely that each penile and clitoral feature has a unique time-course of hormonal dependency/sensitivity.     

Timing of prenatal androgen exposure: anatomical and endocrine effects on juvenile male and female rhesus monkeys

Prenatal androgen shapes genital differentiation. In humans, genital anatomy determines sex of rearing and subsequent behavioral development. Rhesus monkey genital anatomy and neuroendocrine function are sexually differentiated, and behavioral development occurs in a complex social environment. We investigated prenatal hormonal influences on sexual differentiation by suppressing or increasing androgens in male and female rhesus monkeys. Pregnant multiparous female rhesus monkeys received 35-40 days of testosterone enanthate (TE) treatment, androgen antagonist (flutamide, FL) treatment, or vehicle starting on gestation day (GD) 35 or 40 (early) or GD 110 or 115 (late). Exogenous androgen increased neonatal LH secretion in females when given early and altered female genital differentiation when administered either early or late. TE treatment, early or late in gestation, had no measurable effects on male genital differentiation or neuroendocrine function. Early FL treatment, however, radically altered male genital differentiation, producing in two cases males with a urethral opening separate from the glans. In females, early FL treatment produced detectable alterations in genitalia consistent with a reduced exposure to prenatal androgen, suggesting that female rhesus monkeys are naturally exposed prenatally to meaningful levels of T. Late FL treatment reduced male penis size and increased neonatal T secretion, but had no effect in females. This is the first study to block endogenous prenatal testosterone in rhesus monkeys, thereby altering sexual differentiation. These findings illustrate the complexity of prenatal influences on anatomical and neuroendocrine development. The relationship between the anatomical changes reported here and sex differences in behavior is currently under investigation.     

Characteristics of cytosol androgen receptor in rat thymus

Male and female rat thymic cytosol contained specific androgen receptor. The apparent dissociation constants (Kd) were 2.4 nM in males and 2.5 nM in females, and the number of binding sites (NBS) were 23.7 fmol/mg protein in males and 34.2 fmol/mg protein in females. Transformation of receptor to the DNA binding state was achieved by heat or KCl treatment of [3H]R1881-receptor complex, and the characteristics of transformed and nontransformed receptors were investigated. The nontransformed androgen-receptor complex eluted at 0.20-0.25 M KCl from DEAE-Sephacel and sedimented at 9.1 S and its molecular weight was 255,000 on agarose gel chromatography, while the transformed receptor complex eluted at 0.03-0.15 M KCl with a broad peak and sedimented at 4.5 S and its molecular weight was 80,000-85,000. The minicolumn binding assay revealed that approximately 57% of the total receptor complexes bound to DNA-cellulose following heat treatment (20 degrees C, 1 h). Castration exerted no effect on the physicochemical properties of cytosol androgen receptor, but it increased the number of binding site to the female level.     

Influence of perinatal androgenization on the castration response of adult rats

An unexplained dichotomy exists between the LH (luteinizing hormone) responses to castration of male and female rats, as males show a more prompt increase in serum LH levels. We have tested the hypothesis that neonatal exposure to androgen determines the sexual dimorphism of that response. Control groups of male and female rats were castrated at 60 days of age. Other animals had been castrated at 0 or 25 days of age and then given steroid treatment via testosterone (T) implants from 25 through 60 days of age. At 60 days of age a blood sample was taken from each animal before removal of either the T implant or the gonads. Animals were bled again 24 and 48 h later. Within 24 h after orchidectomy the typical early plateau of plasma LH had occurred, represented by an increment in mean LH concentrations of 316 ng/ml. Orchidectomy at 25 days of age had little or no effect on subsequent response to removal of T. In contrast, neonatal orchidectomy resulted in a markedly diminished response to T removal on Day 60. The response, however, was not reduced to that of normal females. In female rats plasma LH does not increase by 48 h after ovariectomy. Perinatal testosterone propionate (TP) treatment of females partially masculinized (enhanced) the LH response to T implant removal, but only if ovariectomy had been performed prior to puberty (at 0 or 25 days of age).(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of androgen deprivation on branching morphogenesis in the mouse prostate

Androgen-induced prostatic development encompasses many individual processes such as ductal branching morphogenesis, cellular proliferation, and secretory cytodifferentiation. Previous studies of ductal morphogenesis (Y. Sugimura, G.R. Cunha, and A.A. Donjacour, 1986, Biol. Reprod. 34, 961-971) demonstrated that the majority (approximately 70%) of ductal tips and branchpoints in the mouse prostate is generated before 15 days of age. Since circulating androgen levels are low during this neonatal period, it is possible that ductal branching morphogenesis may not require the continuous presence of androgens. To test this hypothesis mice were castrated within 24 hr of birth, and prostates from these mice were microdissected at various ages from 5 to 120 days of age to assess the number of ductal tips and branchpoints; wet weight and DNA content were also determined. In intact males wet weight and DNA content increased rapidly between 15 and 60 days of age, after most of the prostatic ductal architecture had been laid down. Neonatal castration considerably reduced the number of tips and branchpoints in both the ventral and dorsolateral prostate, yet both lobes still underwent significant branching morphogenesis in the absence of testes. The administration of anti-androgens to neonatal castrates did not suppress ductal branching to any greater extent than did neonatal castration alone. Androgen replacement immediately following neonatal castration resulted in precocious attainment of the adult number of tips and branchpoints, but caused only modest increases in wet weight. In contrast, when androgen replacement was delayed until adulthood, prostatic wet weight increased to normal adult levels, but the number of ductal tips and branchpoints did not. These experiments show that neonatal prostatic ductal morphogenesis is sensitive to, but does not require, chronic androgen stimulation.     

Correlation of hepatic cytosolic androgen binding proteins with androgen induction of hepatic microsomal ethylmorphine N-demethylase in the rat

Previous reports have demonstrated the presence of moderate to high affinity binding for androgens in the cytosol of livers from male rats. This binding was significantly lower in female rats or in immature rats of either sex. The hepatic androgen binding protein, which sedimented at approx. 4 S on sucrose density gradients, has been called a receptor which mediates the actions of androgens in the liver. The experiments in the present study were designed to evaluate the hepatic androgen binding protein for characteristics which have been attributed to receptors in other tissues and to correlate the presence of androgen binding with androgen induction of hepatic drug metabolism. In the current studies, we have shown that cytosol from the livers of male rats bound [3H]dihydrotestosterone [( 3H]DHT) and translocated this steroid ligand to the nucleus in a time and temperature dependent manner. Cytosol prelabeled with [3H]DHT, when passed over a column of denatured DNA cellulose, eluted in three radioactive peaks. Two of these peaks were absent when cytosol from livers of female or hypophysectomized males was used. In addition, the presence of high concentrations of hepatic androgen binding correlated well with the ability of androgen to induce ethylmorphine N-demethylase, a marker of microsomal cytochrome P-450-dependent drug metabolism. Values for both parameters were higher in males than in either females or hypophysectomized males. Testosterone treatment induced both parameters in ovariectomized females and 17 beta-estradiol repressed both in males. However, testosterone treatment failed to induce hepatic androgen binding in hypophysectomized males and immature males, both of which are also unresponsive to androgen induction of drug metabolism. The results suggest that one or more hepatic cytosolic androgen binding proteins possess several characteristics associated with steroid receptors in reproductive tract tissue. Furthermore, this binding may be implicated as a mediator for the androgen induction of at least one component of hepatic drug metabolism.     

Sexual orientation, proceptivity, and receptivity in the male rat as a function of neonatal hormonal manipulation

The influence of neonatal androgen on the potential to exhibit feminine sexual behavior was investigated. Male rats castrated on Day 0 but not those castrated on Day 4 or later showed hop/darting, ear wiggling, and lordotic behavior in response to treatment with estrogen and progesterone in adulthood at a frequency equal to that of females. Neonatal treatment with testosterone propionate (1 mg/rat for 4 days) abolished the capacity to show these behaviors. In subsequent experiments, involving castration of male rats at 0 or 4 hr after cesarean delivery, the effect of the postnatal surge of testicular secretions on the expression of female sexual behavior was investigated. No differences were seen in the frequency of hop/darting, ear wiggling, and receptivity between males castrated immediately or 4 hr after delivery. In a preference test where the experimental male could choose between an estrous female and a sexually active male, the neonatally castrated males preferred the company of a male when treated with estrogen and progesterone. The implantation of testosterone resulted in a preference for an estrous female. It was concluded that testicular secretions in the newborn male influence adult sexual orientation and suppress the ability to show proceptive and receptive behaviors.     

Ontogeny of androgen receptors in fetal guinea pig brain

Sexual differentiation of the guinea pig brain is androgen dependent. To understand the cellular mechanisms of androgen action, we studied the ontogeny of cytosolic (ARc) and nuclear (ARn) androgen receptors in the brains and anterior pituitaries of fetal, neonatal, and adult guinea pigs. Using cytosol from the hypothalamus-preoptic area-amygdala-septum of 60- to 65-day fetuses and nuclear preparations from 6-day-old neonates treated with testosterone propionate, validation studies revealed an AR with an apparent Kd of 1.9 +/- 1.1 (mean +/- SEM, n = 3) x 10(-10) M (ARc) and 3.4 +/- 3.2 (n = 3) x 10(-10) M (ARn). The cytosolic receptors were highly specific for androgens. After assay validation, AR content was determined from specific brain regions of fetuses obtained on Days 30, 40, 50, and 59 of gestation and on Days 6 and 120 postpartum. ARc differed significantly (p less than 0.05) between brain regions and times of gestation, but no sex differences were apparent. In contrast, ARn showed little difference between tissues or with gestational age, but there were significant differences between males and females, especially in late gestation and early postnatal life, with males having greater ARn binding (p less than 0.05). These data demonstrate the presence of ARc and ARn in the fetal brain and pituitary gland during the critical period of sexual differentiation (Days 30-37 of gestation), thus establishing the identity of cellular structures involved in androgen action.     

Sex Steroid Receptors in the Perinatal Rat Brain

Many sex differences in the regulation of reproductive functionin rats are the result of a differentiation of the brain whichoccurs neonatally. Although injections of either androgens orestrogens are capable during the neonatal period of alteringhypothalamic systems involved in reproductive behavior and gonadotropinregulation, the physiological role of each type of hormone hasnot been clearly established. In both sexes, circulating estrogensare normally kept from interacting with estrogen receptors inthe limbic brain by the high levels of alpha-fetoprotein inthe blood. The local aromatization of androgens in the braincould circumvent alpha-fetoprotein, since androgens do not bindto this serum protein. The "paromatization hypothesis" statesthat testosterone, which is abundant in neonatal male circulationbut absent in females, is locally converted to estradiol inthe limbic brain. There it binds to estrogen receptor proteinsto produce tissue differentiation. The ontogeny of estradiolbinding proteins in limbic areas is consistent with the aromatizationhypothesis, with a rapid increase in receptor levels occurringshortly after birth. Also, the presence of endogenous estradiol-receptorcomplexes has been demonstrated in the cell nuclei of male neonatesbut not female neonales. Furthermore, the presence of estradiolbinding proteins in other regions of the neonatal male and femalebrain suggests an additional role of estradiol. unknown as ofyet. Several studies with agents which block the aromatizationof androgens to estrogens or the binding of estrogens to theirreceptors are consistent with the aromatization hypothesis,since these agents prevent the differentiation of intact neonatalmales. However, specific androgen binding proteins are alsopresent in neonatal brains, and androgen-receptor complexescan be found in cell nuclei of neonates after an injection oftritiated androgen. The possible involvement of these receptorsin sexual differentiation of the brain is suggested by the findingthat an antiandrogen inhibits both the binding of androgens(but not estrogens) and the differentiation of males.     

Aromatase activity in adult guinea pig brain is androgen dependent

Androgen metabolism in target tissues constitutes an important step for understanding hormone action. The in situ aromatization of androgen represents one of these metabolic events. We characterized aromatase activity (AA) in a microsomal preparation of brain tissue from adult guinea pigs since earlier reports questioned its presence in neural tissues of this species. Analyses revealed an apparent substrate affinity (approximately 17 nM) that was equivalent in adult males and females. However, adult male brains contained greater quantities of AA than female brains. Specifically, AA in the preoptic area (POA: p less than 0.05) and the medial basal hypothalamus (MBH; p less than 0.01) was greater in males than in females. AA was concentrated in the limbic system and hypothalamus (amygdala greater than POA greater than septum greater than MBH), whereas low levels were consistently measured in cortical tissue. In vitro estrogen formation was significantly lower in POA (p less than 0.05) and MBH (p less than 0.01) after castration. After dihydrotestosterone treatment, AA returned to levels equal to or greater than those observed in intact males. These data indicate that AA does exist in the guinea pig brain and is modulated by androgens through the androgen receptor. The presence of high levels of aromatase activity may suggest a role for locally formed estrogens in brain function in this species.     

Androgens and male mating behavior in rough-skinned newts, Taricha granulosa

Cyproterone acetate was administered either orally or intraperitoneally to intact, adult male newts, Taricha granulosa. The number of males that exhibited the courtship behavior of clasping when tested with nuptial females was not altered by the antiandrogen treatments. In males which were unresponsive to nuptial females, the occurrence of clasping was not evoked by injections for 4 days of testosterone, dihydrotestosterone, or 11-ketotestosterone. Further, the incidence of clasping was not significantly elevated by injections of prolactin and/or testosterone for 30 days. The effect of sexual activity on testosterone and dihydrotestosterone levels in male newts was determined by radioimmunoassay of plasma collected from males which were: (1) isolated from females; (2) allowed to clasp a female for 2 min; or (3) allowed to clasp a female for 1 hr. The testosterone and dihydrotestosterone levels were unchanged during this period of clasping. In February and again in June, plasma androgen concentrations were measured in males which differed in their propensity to initiate courtship when paired with females. Androgen levels were similar for males that clasped a female and males that never attempted to clasp a female. Plasma androgen levels in the male newt are apparently not correlated with sexual responsiveness.     

Adrenergic receptor properties of hepatocytes from male and female rats

alpha 1- and beta-adrenergic receptor properties of intact hepatocytes from adult male and female rats were evaluated in ligand binding studies using [3H]prazosin and [3H]CGP-12177 (4-(t-butylamino-2-hydroxypropoxy)-[5,7-3H]benzimidazole-2-one-HCl), a hydrophilic beta antagonist. Prior work had suggested that the response of hepatocytes from males to alpha 1-adrenergic stimulation was greater than that of cells from females. However, little sexual difference in prazosin affinity, number of binding sites or kinetics of association/dissociation with the cells was found. Epinephrine, [3H]prazosin competition for binding sites on intact cells was performed at 2 degrees C and 80-90% of agonist sites remained in a high affinity state with an epinephrine Kd comparable to that previously found in glucose release and phosphorylase alpha activation studies. Agonist Kd inferred from these competition experiments also showed no sexual dimorphism. These data suggest that the greater rise in the concentration of cytosolic free calcium and release of 45Ca from cells of males in response to epinephrine stimulation is not due to male/female alpha 1-receptor differences but, rather, may be a function of the previously observed sexual difference in cell calcium metabolism. [3H]CGP binding to hepatocytes from females was stereospecific, saturable and identified a single, high affinity site. Comparable sites were not found on cells from males, however, [3H]CGP binding to crude membrane preparations from both sexes was identical. This suggests that the loss of hepatic beta-receptor function in the adult male is due to an inaccessibility of beta-receptors at the external surface of the plasma membrane of the intact cell. Further studies with other beta-receptor ligands are being carried out to confirm these initial findings.     

Sex hormone-dependent brain maturation and sexual behaviour in rats.

In male and female rats the endogenous steroid and gonadotrophin secretion was inhibited by injecting high doses of chlormadinone acetate (CmAc) from day 14 to 24 of life, i. e. during the period of brain maturation. In adulthood the males treated prepubertally with CmAc exhibited reduced sexual activity and fertility, whereas the females did not differ from the controls. More complete sex hormone deficiency during brain maturation was achieved by castration on day 14 of life. Controls were castrated at normal puberty time (40--60 days). Both groups were then substituted with androgens or oestrogens. In the females castrated on day 14 no impairment of sexual behaviour was observed as compared to the later castrated controls. In contrast, the early castrated males showed delayed onset of mounting behaviour. At autopsy, the weights of their sex organs were found to be lower than in the controls despite equal testosterone replacement for several months. These findings speak in favour of a permanently diminished responsiveness to androgens in males having been exposed to more or less severe androgen deficiency during sex specific brain maturation. Hence, the maturation of a male hypothalamus as well as the differentiation appears to depend at least in part on the presence of androgens, whereas in females it runs without hormonal influence.