The testosterone level in the testes of silver foxes during prenatal development]

The level of testosterone in serum and testes of the silver fox fetuses on days 31, 35, 40, 45, and 50 of gestation was determined using radioimmunoassay. In testes, testosterone was first detected at day 31; then its level gradually increased. Serum testosterone was detected only at day 40. Subsequent increase in its concentration was insignificant. Human chorionic gonadotropin stimulated testicular testosterone production in vitro beginning from day 40. We suggest that in the silver fox, testes are capable of responding to gonadotropins already by the day 40 of prenatal development but the formation of pituitary-gonadal axis proceeds until the end of embryogenesis.     

Testosterone production by the fetal testes of silver foxes selected for their domesticated behavior]

Testosterone content was determined by radioimmune assay in the testes of silver foxes between the 31st and 50th days of gestation. Small quantities of testosterone were found in the testes already at the 31st day of prenatal life, gradually increasing up to a maximum value at the 50th day. No significant difference in testosterone content was found between domesticated and undomesticated silver foxes during prenatal life. It is suggested that selection for domesticated behaviour affects rather central control of endocrine functions than steroid biosynthesis in the testes.     

Cortisol production in fetal adrenals of the silver fox

The present study was designed to examine cortisol production by the silver fox fetal adrenals and their response to ACTH at different periods of prenatal life. Serum levels of cortisol were determined on days 35, 40, 45 and 50 of gestation (term on day 52) in embryos of both sexes. Cortisol content in adrenal tissue homogenates and its in vitro adrenal production were also investigated at the same time points. Hormones were measured by RIA. The levels of cortisol changed slightly during embryonic life. The adrenal content and the in vitro production of cortisol increased sharply and progressively (by 4-10 times from days 35 to 50 of gestation). The rises in cortisol content and in vitro production were associated with a rapid growth of the fetal adrenals. There were no sex differences in cortisol level and its adrenal content. ACTH (50 mIU per sample) increased the in vitro cortisol production by the adrenals in the two sexes on all the studied days (by 3-4 times on day 35 and by 1.4-1.7 times on day 50). These results indicate that 1) silver fox fetal adrenals are capable of synthesising cortisol; 2) ACTH is a potent activator of the in vitro cortisol production during embryonic life in this species.     

Sex-related differences in blood and gonad levels of testosterone in silver fox fetuses

The mass of silver fox fetuses of both sexes, their gonads, and adrenals, and the levels of testosterone in blood serum and in gonads and adrenals were determined from day 31 of gestation and every five days thereafter until its termination. Marked sex-related differences were revealed: the blood and gonad levels of testosterone in male fetuses were much higher than those in female fetuses. The fetal adrenals contained significantly less testosterone than the gonads. No sex-related differences in the content of testosterone in the fetal adrenals were found. No differences were found in the body and adrenal mass in female and male fetuses at all the developmental stages studied, while the mass of ovaries exceeded that of testes from day 45 of gestation. The data obtained suggest sex dimorphism in the production of testosterone by gonads in silver foxes appears after day 35 and appears to correspond to the period of morphological differentiation of gonads.     

Sex-Related Differences in Blood and Gonad Levels of Testosterone in Silver Fox Fetuses

The mass of silver fox fetuses of both sexes, their gonads, and adrenals, and the levels of testosterone in the blood serum and in gonads and adrenals were determined from day 31 of gestation and every five days thereafter until its termination. Marked sex-related differences were revealed: the blood and gonad levels of testosterone in male fetuses were much higher than those in female fetuses. The fetal adrenals contained significantly less testosterone than the gonads. No sex-related differences in the content of testosterone in the fetal adrenals were found. No differences were found in the body and adrenal mass in female and male fetuses at all the developmental stages studied, while the mass of ovaries exceeded that of testes from day 45 of gestation. The data obtained suggest sex dimorphism in the production of testosterone by gonads in silver foxes appears after day 35 and appears to correspond to the period of morphological differentiation of gonads.     

Measurement of inhibin and an index of inhibin production by rat testes during postnatal development

Our previous studies have shown that inhibin activity in rat testes can be measured using an in vitro inhibin bioassay. In animals that have undergone unilateral efferent duct ligation (EDL), the difference in inhibin content between the ligated and nonligated testis can be used as an index of the rate of inhibin production in vivo. In the present study we examined postnatal changes in inhibin content in the testes, and the production rates of inhibin and seminiferous tubule fluid in groups of neonatal, immature, and adult rats from 1 to 80 days old. We detected inhibin activity in testes of 1-day-old rats; the activity level rose linearly to Day 8, subsequently increasing markedly from Day 30 until it reached a plateau at Day 45. Increments in the content of inhibin and weight of the testis after unilateral EDL, interpreted to represent production of inhibin and seminiferous tubule fluid, commenced at Day 20 and increased rapidly between Days 30 and 50, decreasing thereafter to Day 80. The increase in content and production of inhibin was directly correlated to the rise in serum follicle-stimulating hormone (FSH) and testosterone, suggesting that these two hormones are important in controlling inhibin secretion. In addition, the changes in serum FSH from the high, postnatal levels to those typical of adults were inversely correlated to the content and production rate inhibin in the testes and concentrations of testosterone in the serum. These data support the hypothesis that inhibin is specifically involved in the feedback control of pituitary FSH secretion during postnatal development, although a role for testosterone or a synergistic interaction between the two hormones cannot be excluded.     

Testosterone secretion during gubernacular development and testicular descent in the dog

Serum testosterone concentrations ranged from 0.24 to 1.45 nmol/l between Day 53 post coitum (p.c.) until Day 40 post partum (p.p.) and did not show variations that could be correlated with the process of testicular descent. The intratesticular androgen appeared to be mainly testosterone, its concentration being about 5000-fold higher than that in serum whereas 5 alpha-dihydrotestosterone could not be demonstrated. The intratesticular testosterone concentration at the initiation of gubernacular regression (Day 0) was apparently, but not significantly, higher than at Day 49 p.c. and at Day 40 p.p. The ability of the neonatal canine testis to synthesize testosterone was indicated by increased serum testosterone concentrations after hCG stimulation.     

Effect of oestradiol and tamoxifen on the testosterone response in male rats to a single injection of hCG

A single s.c. injection of hCG (100 i.u.) produced a biphasic serum testosterone response in adult male rats, peaks being noted at 2 h (24 ng/ml) and 3 days (16 ng/ml). The levels fell to control during the intervening interval (8 ng/ml), although there were elevated levels of serum hCG. Maintenance of high oestradiol levels by a s.c. injection of 50 micrograms oestradiol benzoate given on Day 2 after the initial hCG injection failed to prolong the refractory period and the secondary peak of testosterone (16 ng/ml) occurred on Day 3. Administration of the antioestrogen, tamoxifen (2 mg or 3 micrograms), 24 h before or simultaneously with hCG did not prevent testicular refractoriness in vivo because serum testosterone levels still declined after 2 h to reach a nadir at 2 days. The basal in-vitro testosterone production by decapsulated testes from animals injected with hCG was enhanced at 2 h. Stimulation by hCG increased the amount of testosterone produced (X 1.5 that in controls). By 12 h basal production decreased and there was no further increment in testosterone in the presence of hCG. This refractoriness to further hCG stimulation prevailed until Day 3, but the total production of testosterone fell so that at 24 h and 2 days testes were producing basal amounts of testosterone. Testes recovered from refractoriness at 4 and 5 days, when basal and stimulated testosterone production were greater than in controls. Injection of 50 micrograms oestradiol benzoate at 2 days did not prolong the in-vitro refractory period and 2 mg or 3 micrograms tamoxifen had no effect on the in-vitro steroidogenic activity, since testes were still refractory to further hCG stimulation from 12 h to 3 days. The results of the present study do not support the hypothesis that oestradiol is involved in the hCG-induced refractoriness of the Leydig cell. The nadir between the peaks of serum testosterone in vivo corresponds to the period during which the testis is refractory to in-vitro stimulation by hCG.     

Steroid hormone content of the gonads of the tammar wallaby during sexual differentiation.

The gonads of the tammar wallaby, Macropus eugenii, are sexually indifferent at birth (Day 0) despite the fact that phenotypic sexual differentiation has already commenced as evidenced by the presence of a scrotum in males and mammary anlagen in females. The seminiferous cords of the testis first become clearly recognizable on Day 2 of pouch life, and ovarian differentiation is recognizable by Day 10. To monitor the endocrine development of the gonads during sexual differentiation of the urogenital tract, we measured the steroid hormone content in 92 pools of gonads from male and female tammar pouch young from the day of birth to 206 days of pouch life. Progesterone, estradiol, and dihydrotestosterone concentrations were low (less than 0.05 ng/mg protein) in both ovaries and testes at all stages examined, and testosterone concentrations were uniformly low in ovaries. Testosterone concentrations in testes were low on Days 0-4, averaging about 0.2 ng/mg protein; they rose by Days 5-10 to an average of 0.9 ng/mg protein, remained elevated until about Day 40, and thereafter fell to values similar to those in the ovaries. The phallus and urogenital sinus were able to convert testosterone to dihydrotestosterone from the earliest stages examined (Days 10 and 11). Thus in the tammar wallaby, as in eutherian mammals, testosterone is the androgen secreted by the developing testis, and dihydrotestosterone is formed in certain androgen target tissues.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of postnatal treatment with a gonadotropin-releasing hormone antagonist on sexual maturation of male rats

The role of postnatal pituitary-testicular activity in sexual maturation at puberty was studied in male rats. Rats were injected twice daily with a potent gonadotropin-releasing hormone antagonist (N-Ac-4-Cl-D-Phe1, 4-Cl-D-Phe2, D-Trp3, D-Phe6, D-Ala10-NH2-GnRH) (GnRH-Ant.), 2 mg/kg, on Days 1-15 of life, and killed on Day 48, 56 or 90 of life. The treatment delayed the onset of puberty (monitored by balano-preputial separation) by 8 days (from the age of 48 to 56 days). The weights of testes, seminal vesicles and ventral prostates were reduced by 50-60% on days 48 and 56 of life, but only the testis weights remained suppressed by Day 90. Levels of serum luteinizing hormone (LH) and follicle-stimulating hormone (FSH), but not those of prolactin (Prl), were elevated 2-to-4-fold in the treated animals at the three ages studied. Serum and testicular testosterone (T) and the receptors for LH and Prl were suppressed in the peripubertal animals (48 and 56 days), but serum T was elevated and the receptor levels were normal in the 90-day group. The testicular FSH receptors were 50% suppressed at all ages studied. Only minor changes were observed in testicular histology when studied at 48 and 56 days. The 85-day-old animals treated with GnRH-Ant. were infertile when mated with females.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effect of prenatal treatment with busulfan on the hypothalamo-pituitary axis, genital tract and testicular histology of prepubertal male rats

Female Wistar rats were treated with busulfan or with solvent on Day 20 of pregnancy. Thirty male offspring of each group were killed at 38 days of age. In busulfan-treated rats, compared to controls, hypothalamic LH-RH content was decreased by 52%, whereas pituitary LH and FSH concentrations were increased by 60 and 43% respectively. Plasma LH and FSH were increased by 112 and 275% respectively. Prolactin concentrations were not changed, but plasma testosterone concentration was decreased by 48%. The total number of Leydig cells per testis was decreased by 52%, and LH binding sites per testis were decreased by 70%. The total number of Sertoli cells was decreased by 44%, while FSH binding sites per testis were decreased by 62%. Spermatogenesis was practically absent after prenatal exposure to busulfan. These data demonstrate that on Day 20 of pregnancy all the dividing cells in the fetal testes were depleted by an antimitotic treatment. The stimulation of the hypothalamo-pituitary axis could have been partly induced by the decrease in testosterone production, and by the aplasia of germ cells involving modifications of the remaining Sertoli and Leydig cells.     

Maternal transfer of photoperiodic information in Siberian hamsters. V. Effects of melatonin implants are dependent on photoperiod.

Photoperiodic information is transferred from female Siberian hamsters to their fetuses during gestation. Although maternal melatonin is known to be essential for the transfer of prenatal photoperiodic information, its specific role is not well defined. The duration of the daily melatonin signal, expressed as an elevation of serum melatonin levels in the maternal circulation, has been hypothesized to convey day length information to the fetus. If this hypothesis is valid, it predicts that identical maternal melatonin signals should affect the fetuses identically, regardless of the prenatal photoperiod. To test this hypothesis, adult females received melatonin in beeswax or beeswax alone. They were paired with males and housed in photoperiods of 12L:12D or 16L:8D. On the day of parturition, mother and young were transferred to constant light (LL). Young males were killed on Day 28 of life, and weights of testes were determined. Prenatal treatment with beeswax alone did not affect the nature of the signal transferred from mother to fetus; young gestated in 12L:12D and reared in LL developed small testes, while those gestated in 16L:8D had large testes. On the other hand, the effect of the prenatal melatonin treatment on postnatal testicular development in LL was inversely dependent on the prenatal photoperiod: testicular growth was stimulated in young gestated in 12L:12D, but inhibited in young gestated in 16L:8D. To verify that the melatonin pellets produced equivalent serum melatonin levels in adult females in 12L:12D and 16L:8D, unmated adult females were killed 6-10 wk after receiving melatonin pellets. Serum levels were elevated in both groups throughout the day and night.(ABSTRACT TRUNCATED AT 250 WORDS)     

Retinoid-sensitive steps in steroidogenesis in fetal and neonatal rat testes: in vitro and in vivo studies

Retinoic acid (RA) was recently shown to modify testosterone secretion of the fetal testis in vitro. We characterized this effect by culturing rat testes explanted at various ages, from Fetal Day 14.5 to Postnatal Day 3. In basal medium, RA inhibited, in a dose-dependent manner, both basal and acute LH-stimulated testosterone secretion by testes explanted on Fetal Days 14.5, 15.5, and 16.5. It had no effect on testes from older animals. The negative effect of RA did not result from a diminution in the number of Leydig cells but from a decrease in P450c17 mRNA levels and in LH-stimulated cAMP production. However, the RA-induced decrease in P450C17 mRNA levels was also observed with neonatal testes, suggesting that this enzymatic step is no longer rate limiting at this developmental stage. To study the physiological relevance of RA effects, we used fetuses and neonates issued from mothers fed a vitamin A-deficient (VAD) diet, resulting in a threefold decrease of plasma retinol concentration. On Fetal Day 18.5 and on Posnatal Day 3, testosterone secretion by the testis ex vivo was significantly increased in VAD animals. This shows that the endogenous retinol inhibits differentiation and/or function of fetal Leydig cells before Fetal Day 18.5 and is required for the normal regression of fetal Leydig cell function that occurs after Fetal Day 18.5. In conclusion, our results show that retinoids play a negative role on the steroidogenic activity during the differentiation of rat fetal Leydig cells.     

Virilization of the male pouch young of the tammar wallaby does not appear to be mediated by plasma testosterone or dihydrotestosterone.

Virilization of the male urogenital tract of all mammals, including marsupials, is mediated by androgenic hormones secreted by the testes. We have previously demonstrated profound sexual dimorphism in the concentrations of gonadal androgens in pouch young of the tammar wallaby Macropus eugenii during the interval when the urogenital sinus virilizes. To provide insight into the mechanisms by which androgens are transported from the testes to the target tissues, we measured testosterone and dihydrotestosterone in plasma pools from tammar pouch young from the day of birth to Day 150. Plasma testosterone levels were measurable (0.5-2 ng/ml) at all times studied, but there were no differences between males and females. These low concentrations of plasma testosterone appear to be derived from the adrenal glands and not the testes. Plasma dihydrotestosterone levels in plasma pools from these animals were also low and not sexually dimorphic. We conclude that virilization of the male urogenital tract cannot be explained by the usual transport of testosterone or dihydrotestosterone in plasma but may be mediated by the direct delivery of androgens to the urogenital tract via the Wolffian ducts. Alternatively, circulating prohormones may be converted to androgens in target tissues.     

Effect of experimental infection with Trypanosoma congolense and scrotal insulation on Leydig cell steroidogenesis in the ram

Testicular steroid content and Leydig cell steroidogenesis in vitro were investigated in rams on Days 28 and 58 after Trypanosoma congolense infection and were compared with those of rams in which testicular temperature had been raised artificially by insulation of the scrotum for 58 d. Testicular testosterone content increased significantly on Day 28 after infection but was lower than that of controls on Day 58 while it increased in scrotal-insulated rams compared with that of controls by Day 58. Testicular progesterone was undetectable in the control and trypanosome-infected groups throughout the experiment, but it increased in the insulated rams by day 58. Basal (unstimulated) Leydig cell testosterone production in the infected rams was similar to that of control rams on Day 28 but was significantly lower on Day 58. Stimulation of Leydig cell testosterone production with hCG or 22R-hydroxycholesterol (22ROHC) significantly reduced in infected rams at both 28 and 58 d after infection as well as in scrotal-insulated rams on Day 58. It is concluded that the increase in testicular testosterone content in the infected and scrotal-insulated rams on Days 28 and 58, respectively, was induced by elevation of testicular temperature by trypanosome infection, perhaps through an effect on testicular blood flow. Reduced testosterone production by Leydig cells from infected and scrotal-insulated rams in response to hCG and 22ROHC suggests that trypanosome-induced pyrexia might be involved in reducing Leydig cell steroidogenesis and subsequent plasma testosterone levels, possibly by affecting enzymes involved in steroid biosynthesis.     

Testosterone response of cryptorchid and hypophysectomized rats to human chorionic gonadotrophin (hCG) stimulation

The testosterone responses to a single injection of hCG (100 i.u.) in hypophysectomized (hypox.), cryptorchid or sham-operated rats were followed over a 5-day period. In sham-operated rats, hCG induced a biphasic rise in serum testosterone, peaks being observed at 2 and 72 h. Reduced testis weights, elevated FSH and LH levels and reduced serum testosterone levels were found after 4 weeks of cryptorchidism, but hCG stimulation resulted in a normal 2 h peak in serum testosterone. However, the secondary rise at 72 h in cryptorchid rats was significantly lower than sham-operated rats. Reduced testis weight and undetectable serum FSH and LH levels together with decreased testosterone levels were found 4 weeks after hypophysectomy. Serum testosterone levels rose 2 h after hCG in comparison to hypox. controls but this peak was significantly reduced compared with sham-operated rats. The second rise in serum testosterone began on day 2, peaking on day 4 at levels comparable to that seen in sham-operated rats after hCG. The in vitro basal and hCG stimulated secretion of testosterone by cryptorchid testes was greater than that secreted by normal rat testes (518.0 +/- 45.9 and 3337.6 +/- 304.1 pmol per testis per 4 h compared with 223.6 +/- 24.9 and 1312.9 +/- 141.4 pmol per testis per 4 h for normal rat testes). In cryptorchid animals a single injection of 100 i.u. hCG resulted in a pattern of in vitro refractoriness similar to normal rats, lasting from 12 h to 2 days, during which testosterone secretion was reduced to near basal levels. The in vitro basal and hCG-stimulated secretion of testosterone by hypox. rat testes was severely diminished compared with normal rat testes. The temporal pattern of in vitro secretion of testosterone from hypox. rat testes mimicked the in vivo serum testosterone pattern seen in these animals. This study demonstrates important differences in the in vivo and in vitro testosterone response to hCG after testicular damage.     

Growth and sexual maturation of the male house musk shrew (Suncus murinus)

The sexual maturation in the male house musk shrew, Suncus murinus, was investigated by weighing gonads, accessory sexual organs and various other organs, and measuring testosterone concentrations in the plasma from birth to 50 days of age. The histological and histochemical studies of the testis were also carried out. Testicular weight increased markedly from 10 days of age. On 15 days of age, the activity of delta 5-3 beta-Hydroxysteroid dehydrogenase (HSD) was detected in Leydig cells. In accordance with an increase of HSD activity, the concentrations of plasma testosterone raised rapidly from 15 days of age and reached a peak on Day 30. Growth of prostate glands and seminal vesicles showed a sigmoidal pattern and their significant growth began from Day 25. Sperms were detected first in the seminiferous tubules on Day 28, and most tubules were filled with many sperms on Day 33. From these results, the beginning of sexual maturation, i.e. puberty, of the male shrew is considered to be around Day 25. The weight gain of testes, accessory sexual organs and the body became slower on Day 40. The male animal of Day 40 could mate and impregnate the female. Therefore, the male shrew seems to attain to the sexual maturity between days 35 and 40.     

The effect of prenatal stress on activity of the neurosteroids synthesis enzyme in the "critical period" of brain sexual differentiation in male rats

The effect of daily immobillisation stress in female rats on the 15th to 18th days of pregnancy upon synthesis enzyme for neurosteroids of alpha-reductase in their male offspring brain, was studied. A decrease in the enzyme activity in the cortex and hypothalamus of male foetuses occurred within 24 hr following the latest stress, whereas it was increased in the cortex of newborn offspring. An enhancement of the 5 alpha-reductase activity in the cortex, hippocampus and hypothalamus was also found in prenatally stressed males on the 5th day of life. A decrease in the testosterone and progesterone contents in the blood plasma of the animals under study was revealed on the 19th day of their embryonic life as well as in newborn rats, the blood level of progesterone, at that, remained decreased even at the age of 5 days. A possible part ofneurosteroids in action of prenatal stress upon sexual differentiation of the brain is discussed.     

Brain aromatase activity and behavioral consequences in the male rat treated in utero with 4-hydroxy-androstenedione

Pregnant female rats were given daily injections of a potent aromatase inhibitor, 4-hydroxy-delta 4-androstenedione (4OHA), throughout the latter half of the pregnancy (days 11 to 22; the day of insemination was designated as Day O) and male fetuses and pups were obtained. Control animals were male offspring of mothers treated with oil vehicle. When measured by the tritium-water method, significant reductions in the aromatase activity were detected in the hypothalamic and preoptic continuum (HPOA) of the male fetuses and pups, over a period from day 16 of the pregnancy until a day after birth. All parturitions in the experimental as well as control animals occurred on day 22 of the pregnancy. Behavioral and anatomical consequences of the prenatal treatment were examined in adulthood. When mounted by stud male rats, the male litters of the 4OHA-treated mother showed lordosis at a significantly higher frequency both in terms of the number of positive test sessions (each consisted of a 20-min period in which the subject showed lordosis at least once) and the lordosis quotient (percent lordosis occurrence per 10 mounts). When placed with receptive female rats, the experimental animals were no less active in mounting or ejaculating than the control. No significant difference existed between the experimental and control animals in the weight of the testes or the accessorial genitalia, or the serum testosterone levels. Partial but significant intervention of behavioral defeminization of the brain was associated with decreased HPOA aromatase activity during the prenatal period.