Role of dihydrotestosterone in the control of sexual behaviour on castrated male sheep

Wethers (at least 2 1/2 years after castration) were implanted with testosterone propionate (TP), oestradiol dipropionate (ODP), dihydrotestosterone, or a combination of dihydrotestosterone and ODP Silastic capsules. Active immunization against both oestradiol and oestrone or oestradiol only was used to negate effects of oestrogens produced by aromatization of TP. On exposure to oestrous ewes, immunization of wethers implanted with TP significantly (P less than 0.01) reduced all components of mating behaviour (except sniffing and Flehmen) to levels seen in untreated controls. The results support the conclusion that dihydrotestosterone potentiates the action of oestrogens, particularly as regards Flehmen, and has no action on its own within the central nervous system, while oestrogens do not restore mating activity to the same level as that following treatment with testosterone.     

Courtship and copulation in prepubertally castrated male sheep (wethers) treated with 17β-estradiol,aromatizable androgens,or dihydrotestosterone

Groups of sexually inexperienced adult Clun Forest sheep (four animals per group) which had been castrated on the day after birth received one of the following treatments: testosterone propionate (TP, 20 mg/day); estradiol dipropionate (ODP, 2 mg/day); 19-hydroxy-17, 19-dipropionate (19HTP, 20 mg/day); dihydrotestosterone propionate (DHTP, 20 mg/day); or arachis oil vehicle (OIL). Treatments were in the form of sc injections given 5 days/week over a 6-week period during which time individual animals were observed in 18 tests for sexual behavior. The stimulus females used were ovariectomized ewes maintained in a state of continuous receptivity by daily injections of 15 mg of TP. Various measures of sexual and aggressive behavior were recorded during each test. Mounting was induced mainly in animals in the TP group and to a lesser extent in those receiving ODP. The extent to which precopulatory courtship was induced followed the order TP > ODP > 19HTP. Animals treated with DHTP or OIL showed negligible sexual activity.     

The effectiveness of steers and heifers treated with oestrogen or testosterone to detect oestrus in cattle

Two experiments were carried out to determine the effectiveness of steers and heifers, treated with oestrogen or testosterone, in the detection of oestrus in cattle.In the first experiment 17 steers castrated at birth and 16 castrated at 6 months of age were randomly allocated to three groups and received an 800 mg subcutaneous implant of testosterone, subcutaneous injections of 10 mg oestradiol benzoate per week for 16 weeks or no hormone (controls). In addition, six heifers were injected subcutaneously with 10 mg oestradiol benzoate per week for 16 weeks while six untreated heifers served as controls. Animals were observed in a standardised libido test 2, 4, 8, 16, 20 and 24 weeks after treatment commenced. The time to first mount and the number of mounts per animal responding in the presence of oestrous heifers were recorded. Both steers and heifers treated with oestradiol benzoate were superior at detection of oestrus in cattle than animals treated with testosterone or those receiving no hormone. Oestrogen-treated animals generally detected heifers in oestrus in less than 3 min after introduction and mounted these animals between 20 and 30 times in one hour. This response was consistent throughout the duration of the experiment. There was no effect of age at castration of steers on development of male behaviour.The second experiment determined the rate and degree of development of male behaviour in steers in response to weekly subcutaneous injections of 0, 2, 4, 8 or 16 mg oestradiol benzoate per 250 kg body weight, 250 mg testosterone or 150 mg dihydrotestosterone for a period of 15 weeks. Steers treated with oestradiol benzoate again proved to be more successful than untreated or testosterone-treated steers at consistently detecting and mounting oestrous heifers. The best response was obtained from steers treated with 8 mg/250 kg body weight per week. The practical application of this work is discussed.     

Serum gonadotrophin levels in prepubertally castrated male sheep treated for long periods with propionated testosterone, dihydrotestosterone, 19-hydroxytestosterone or oestradiol.

At different times of the year, groups of wethers were treated with 20 mg testosterone, dihydrotestosterone or 19-hydroxytestosterone propionates/day or 2 mg oestradiol dipropionate/day, or the oil vehicle, for 6 weeks after a 2-week control period. LH and FSH values were determined by radioimmunoassay of serum samples collected at regular intervals. Oestradiol and dihydrotestosterone reduced LH and FSH concentrations whereas 19-hydroxytestosterone and testosterone had no effect.     

Neonatal dihydrotestosterone and estrogen stimulation: Effects on sexual behavior of male rats

Male rats castrated on the second day after birth (Day 2) were, for the next 10 days, given daily injections of one of five steroids or steroid combinations: 200 μg of testosterone propionate (TP); 200 μg of dihydrotestosterone propionate (DHTP); 5 μg of estradiol benzoate (EB); 5 μg of estradiol benzoate plus 200 μg of dihydrotestosterone propionate; oil vehicle (VH). Control male rats castrated on Day 90 received a sham castration and oil vehicle in the neonatal period. All animals were given TP in adulthood and tested for male sexual behavior. There was no difference in mounting activity among the subjects. Day 2 DHTP subjects displayed intromissions but were incapable of ejaculating. The more frequent display of intromissions by Day 2 DHTP animals in comparison to Day 2 VH animals could be solely due to their larger and more highly developed penes. On the other hand, the ejaculatory failure of the Day 2 DHTP subjects was attributed to some deficiency in central neural processes controlling ejaculatory mechanisms rather than inadequate penile development. Equivocal results were obtained with the Day 2 EB and Day 2 EB-DHTP animals in that only a few animals in both groups showed an ejaculatory pattern.     

Androgens and the social behavior of male and female lizards (Anolis carolinensis)

This study investigated the androgen specificity of aggressive and sexual behavior in the lizard Anolis carolinensis and the capacity of females of this species to exhibit male-typical copulation. Gonadectomized males and females were injected with testosterone propionate (TP) or dihydrotestosterone propionate (DHTP) or were implanted with Silastic tubing containing TP or DHTP. Either TP or DHTP activated male-typical sexual behavior in both males and females and activated aggressive behavior in males; DHTP activated aggressive behavior in females. Thus conversion of androgen to estrogen is not essential for these behavior patterns, and endogenous dihydrotestosterone may be important. TP but not DHTP stimulated receptivity in females, suggesting that conversion of testosterone to estrogen may underlie TP-stimulated receptivity. Females treated with TP did not differ from males in their display of male-typical courtship, neck-clasping, and intromission.     

Detection of estrus with cows administered testosterone via injections and/or silastic implants

Over a three year period 8 cows and 2 heifers were administered testosterone via injections and/or silastic implants. During and subsequent to treatments, blood samples were collected for determination of testosterone and cows were observed for male sex behavior. Male sex behavior was induced by administering 200 mg of testosterone propionate every other day for 20 days, and once induced, male sex behavior was maintained by injections of 200 mg of testosterone propionate every 10 days or by two 15 cm silastic implants containing testosterone propionate. Male sex behavior was also induced and maintained by administering an injection of testosterone enanthate (1 gm) and testosterone propionate (200 mg) and two 15 cm silastic implants containing testosterone propionate at the same time. Cows which are administered testosterone in this manner detected 94% of 231 females that were in estrus 342 times.     

Inhibition of adult social behaviour in ducks induced by juvenile administration of gonadal hormones

This experiment was designed to study the long-term behavioural effects of gonadal hormone injections into domestic ducklings. Ten male and four female ducklings were injected daily from their 4th day onwards: the males with testosterone propionate and the females with oestradiol benzoate. Five males were injected for 40 days, the other ducklings (5♀ + 4♂) for 80 days. Behavioural effects of these treatments were assessed when the birds were six months old. Social displays were strongly inhibited in the injected males as compared with control birds but no general significant effect was observed for the aggressive and sexual behaviour. The inhibition of social displays is consistent with the view that these behaviour patterns are regulated by the pituitary-gonadal axis and the different levels at which the functional inhibition could have occured are discussed. Far fewer social displays were performed by the males presented with injected females which, on the other hand, showed less sexual behaviour than the control females. This supports the idea that the female plays an important part in the social display of ducks.     

Induction and Maintenance of Maternal Behaviour in the Domestic Hen: Influence of Testosterone and Oestradiol Treatments

Confining an adult hen with two newly-hatched chicks induced a gradual emergence of maternal behaviour in the hen. The aim of the experiments presented here was to analyse the effects of testosterone or oestradiol treatments on the induction and maintenance of maternal behaviour in hens with no previous maternal experience. The ability of a hen to positively respond towards chicks was not altered by either testosterone or oestradiol injections. However, testosterone therapy prevented both the appearance and the maintenance of the most typical call, clucking. Testosterone injected hens responded to chicks with a contact-type call, but oestradiol treated ones did not. These results indicate that some aspects of maternal behaviour can be modulated by peripheral levels of hormones.     

Testosterone promotes paternal behaviour in a monogamous mammal via conversion to oestrogen

Although high testosterone (T) levels inhibit paternal behaviour in birds breeding in temperate zones many paternal mammals have a very different breeding biology, characterized by a post-partum oestrus. In species with post-partum oestrus, males may engage in T-dependent behaviours such as aggression and copulation simultaneously with paternal behaviour. We previously found that T promotes paternal behaviour in the California mouse, Peromyscus californicus. We examine whether this effect is mediated by the conversion of T to oestradiol (E(2)) by aromatase. In the first experiment, gonadectomized males treated with T or E(2) implants showed higher levels of huddling and pup grooming behaviour than gonadectomized males treated with dihydrotestosterone or empty implants. In the second experiment, we used an aromatase inhibitor (fadrozole) (FAD) to confirm these results. Gonadectomized males treated with T + vehicle or E(2) + FAD showed higher levels of huddling and pup grooming behaviour than gonadectomized males treated with T + FAD or empty implants. Although E(2) is known to promote the onset of maternal behaviour to our knowledge our results are the first to demonstrate that E(2) can promote paternal behaviour in a paternal mammal. These results may explain how mammals express paternal behaviour while T levels are elevated.     

Organizational effects of testosterone,estradiol, and dihydrotestosterone on vasopressin mRNA expression in the bed nucleus of the stria terminalis

In adulthood, male rats express higher levels of arginine vasopressin (AVP) mRNA in the bed nucleus of the stria terminalis (BST) than do female rats. We tested whether this sex difference is primarily due to differences in neonatal levels of testosterone. Male and female rats were gonadectomized on the day of birth and treated with testosterone propionate (TP) or vehicle on postnatal days 1, 3, and 5 (P1, P3, and P5). Three months later, all rats were implanted with testosterone-filled capsules. Two weeks later, brains were processed for in situ hybridization to detect AVP mRNA. We found that neonatal TP treatment significantly increased the number of vasopressinergic cells in the BST over control injections. We then sought to determine the effects of testosterone metabolites, estradiol and dihydrotestosterone, given alone or in combination, on AVP expression in the BST. Rat pups were treated as described above, except that instead of testosterone, estradiol benzoate (EB), dihydrotestosterone propionate (DHTP), a combination of EB and DHTP (EB+DHTP), or vehicle was injected neonatally. Neonatal treatment with either EB or EB+DHTP increased the number of vasopressinergic cells in the BST over that of DHTP or oil treatment. However, treatment with DHTP also significantly increased the number of vasopressinergic cells over that of oil treatment. Hence, in addition to bolstering evidence that estradiol is the more potent metabolite of testosterone in causing sexual differentiation of the brain, these data provide the first example of a masculinizing effect of a nonaromatizable androgen on a sexually dimorphic neuropeptide system.     

The effects of the administration of testosterone propionate alone or with phenobarbitone and of testosterone metabolites to neonatal female rats

The effects of graded doses of testosterone propionate administered to female rats on Day 4 of postnatal life have been determined. The incidence of failure of ovulation as adults was related to the dose. With increasing dosage over the range 1–100 μg no significant evidence of a progressive decrease in immunoassayable luteinizing hormone concentration in the plasma or anterior pituitary was obtained. The reported protective action of sodium phenobarbitone when administered with testosterone propionate could not be confirmed. Single injections of 100 μg of testosterone, dihydrotestosterone, 5 α-androstanediol, or a combination of the two latter compounds had no masculinizing effect. When the 17-β propionate derivatives of these compounds were administered at the same dose level only testosterone propionate had a masculinizing effect.     

Gonadal steroid preservation of central synaptic input to hamster facial motoneurons following peripheral axotomy

In recent work, we have demonstrated that testosterone propionate accelerates recovery from facial nerve injury in the adult male hamster. Central synaptic stripping following peripheral motor neuron damage is a well-established component of the injury response. Gonadal steroids regulate synaptogenesis in the normal nervous system. In this study, we tested the hypothesis that testosterone propionate administration at the time of facial nerve transection alters the synaptic connectivity of injured facial motoneurons. Adult hamsters were subjected to right facial nerve transection at the level of the stylomastoid foramen. Half the animals received subcutaneous implants of testosterone propionate; the other half were sham implanted. At 5 days postoperative, the animals were killed by intracardiac perfusion-fixation, and the control and axotomized facial nuclear groups from the brainstems of nonhormone- and testosterone propionate-treated animals processed for routine transmission electron microscopy. Quantiative analysis of the synaptic ratio (percent somal membrane covered by synaptic profiles) and the average length of axosomatic synapses was accomplished. The results indicate that axotomy alone resulted in an 81% reduction in the synaptic ratio and a 26% decrease in the average synaptic length of axosomatic synapses. Exposure to testosterone propionate from the time of facial nerve transection resulted in only a 48% reduction in the synaptic ratio and a 16% decrease in the average synaptic length of axosomatic synapses following injury. Thus, testosterone propionate significantly attenuated the amount of synaptic stripping that occurred at 5 days postoperative and the decrease in average length of the remaining synapses as well. It is concluded that gonadal steroids modulate central synaptic plasticity following peripheral nerve injury. The results are discussed in light of our recent findings of steroidal effects on the central astrocyctic response to facial nerve injury as well.     

Organizational effects of testosterone,estradiol, and dihydrotestosterone on vasopressin mRNA expression in the bed nucleus of the stria terminalis

In adulthood, male rats express higher levels of arginine vasopressin (AVP) mRNA in the bed nucleus of the stria terminalis (BST) than do female rats. We tested whether this sex difference is primarily due to differences in neonatal levels of testosterone. Male and female rats were gonadectomized on the day of birth and treated with testosterone propionate (TP) or vehicle on postnatal days 1, 3, and 5 (P1, P3, and P5). Three months later, all rats were implanted with testosterone‐filled capsules. Two weeks later, brains were processed for in situ hybridization to detect AVP mRNA. We found that neonatal TP treatment significantly increased the number of vasopressinergic cells in the BST over control injections. We then sought to determine the effects of testosterone metabolites, estradiol and dihydrotestosterone, given alone or in combination, on AVP expression in the BST. Rat pups were treated as described above, except that instead of testosterone, estradiol benzoate (EB), dihydrotestosterone propionate (DHTP), a combination of EB and DHTP (EB+DHTP), or vehicle was injected neonatally. Neonatal treatment with either EB or EB+DHTP increased the number of vasopressinergic cells in the BST over that of DHTP or oil treatment. However, treatment with DHTP also significantly increased the number of vasopressinergic cells over that of oil treatment. Hence, in addition to bolstering evidence that estradiol is the more potent metabolite of testosterone in causing sexual differentiation of the brain, these data provide the first example of a masculinizing effect of a nonaromatizable androgen on a sexually dimorphic neuropeptide system. © 2003 Wiley Periodicals, Inc. J Neurobiol 54: 502–510, 2003     

Neonatal hormone manipulations and the maintenance of perineal muscles and their motor neurones in Albino Swiss rats

Newborn female Albino Swiss rats received testosterone propionate, dihydrotestosterone benzoate or oestradiol benzoate for 4 days after birth. The neonatal administration of all three hormones maintained neurones of the spinal nucleus of bulbocavernosus (SNB) complex in adulthood at levels intermediate between those found in normal females (approximately 40 neurones) and those found in normal males (approximately 220 neurones). Dihydrotestosterone benzoate was the most effective treatment. Oestradiol benzoate, while as potent as testosterone propionate in maintaining SNB neurone numbers, could not maintain the perineal muscles which are their normal target. Dihydrotestosterone benzoate and testosterone propionate maintained both neurones and muscles. Newborn male Albino Swiss rats received either the aromatase inhibitor 4-OH-androstenedione, or the 5 alpha-reductase inhibitor aza-steroid 17 beta-N,N-diethylcarbamoyl-4-methyl-4-aza-5 alpha-androstan-3-one(4-MA). Only neonatal treatment with 4-MA led to reduced SNB neurone numbers in adulthood, but the reduction was modest (-16%). The results of the two experiments suggest that several hormones can maintain SNB neurone numbers in Albino Swiss rats, but that 5 alpha-reduced metabolites of testosterone may be particularly effective.     

Changed levels of endogenous sex steroids in women on oral contraceptives.

Serum and urinary levels of unconjugated testosterone, dihydrotestosterone, and oestradiol were measured by specific radioimmunoassays in 10 healthy women in the early follicular phase of their menstrual cycle and in nine healthy women taking oral contraceptives. The contraceptive group had testosterone levels 1-3 times higher and dihydrotestosterone levels two times higher than those in the controls. Serum oestradiol levels in the contraceptive group were much lower than those in the controls and similar to levels in postmenopausal women. The contraceptive group had about twice the urinary excretion of unconjugated (free) testosterone and dihydrotestosterone of the controls, but their excretion of unconjugated oestradiol was 2-7 times lower. The great increase in serum and urinary androgen concentrations, as well as the suppression of oestradiol, may be related to the antiovulatory effect of oral contraceptives.     

Plasma testosterone and sexual behavior following intracerebral implantation of testosterone propionate in the castrated male rat

Interpretation of behavioral and other effects of intracranial steroid implants depends on knowledge of the rate of release of the implanted hormones into the general circulation. Testosterone propionate implants (200 μg, pellets) in the median eminence and pituitary were found to result in circulating levels of testosterone (T) twice as high as those in the anterior hypothalamus-preoptic area (AHPOA), posterior hypothalamus (PH), and cortex (Ctx). Implants in all cranial areas examined resulted in plasma T levels in the lower range of circulating T found in normal rats for the first 24 hr postoperatively, decreasing thereafter and reaching very low levels by the end of 2 weeks. There were no significant differences in the plasma T levels in rats with implants in the AHPOA, PH, and Ctx, but AHPOA implants were slightly more effective in restoration of sexual behavior than PH implants, and both of these implants were considerably more effective than those in the cortex. There was no apparent correlation between behavioral responses and peripheral levels of T. The major conclusion of this study was that the effects of hypothalamic implants of T on male sexual behavior cannot be explained by the presence of T in the peripheral circulation.     

Effects of estradiol benzoate in combination with dihydrotestosterone on postejaculatory vocalization and refractory period in castrated male rats

The objective of this experiment was to compare the effects of estradiol ben-zoate (EB) treatment with those of testosterone propionate (TP) on the postejaculatory vocalization and refractory period in castrated dihydrotestosterone (DHT)-treated male rats. Twelve reliable maters were tested, castrated, and then treated with subcutaneous implants of DHT and daily injections of either 200 μg of TP (N = 6) or 200 μg of EB (N = 6). Testing continued weekly for 9 weeks with treatments interchanged after the fourth week. EB treatment resulted in: (1) a reduction in the number of males that vocalized, (2) a reduction in the duration of vocalization, and (3) the exhibition of extraordinarily abbreviated postejaculatory refractory periods by a few males. It was suggested that high doses of estradiol act to counter inhibitory processes during the refractory period.     

Testosterone metabolism in the medial basal hypothalamus of the starved male rat

The conversion of testosterone to estradiol by aromatase and to dihydrotestosterone by 5 alpha-reductase was measured in the medial basal hypothalamus of starved and control male rats. Activities of both enzymes were significantly reduced in starved animals. Aromatase activity was 18.2 +/- 2.3 versus 29.8 +/- 5.7 fmol E2/mg protein/90 min (mean +/- SEM, P less than 0.02) and 5 alpha-reductase was 4.95 +/- 0.35 versus 5.96 +/- 0.30 pmol DHT/mg protein/90 min (P less than 0.02) for starved and control animals respectively. The results indicate that hypothalamic metabolism of testosterone is decreased during starvation. Therefore the increased sensitivity of the T-LH feedback described earlier in starved rats [4] cannot be explained by changes in central testosterone metabolism.     

Effects of exogenous gonadotropic and steroid hormones on the social behaviour and gonadal maturation of male domestic ducklings Anas platyrhynchos L.

Male domestic ducklings were injected during their first month of life with mammalian gonadotrophins (ovine LH or FSH, HMG) or gonadal steroids (testosterone or oestradiol). LH and testosterone stimulated sexual behaviour while oestradiol inhibited the increase of aggression observed in control birds during the experiment. The mammalian gonadotrophins did not increase plasma testosterone but nevertheless they all stimulated the testis growth. Several hypotheses which could explain this finding (stimulation of spermatogenesis without any apparent effect on testosterone) are discussed and the possibility of a direct action of LH on the sexual behaviour is analysed. Social displays were only moderately stimulated by testosterone and not at all by gonadotrophins. The hormonal controls of these behaviour patterns remains thus largely unknown.