Seasonal specificity of hormonal, behavioral, and coloration responses to within- and between-sex encounters in male lizards (Sceloporus undulatus).

This study reports the gender and seasonal specificity of hormonal, behavioral, and coloration responses displayed by "resident" male lizards (Sceloporus undulatus) exposed to male or female "intruders" during staged encounters in outdoor enclosures. Resident males were engaged in staged encounters with males or females for 1 h per day on 9 consecutive days during the breeding and postbreeding seasons. Male-specific responses occurred during the breeding but not the postbreeding season. These included (1) a transient increase in plasma testosterone (T) that was evident on Day 4 and had subsided by Day 10, (2) behavioral displays of aggression (full shows and chases), and (3) a lightening of dorsal integumental color. Female-specific behavioral responses (nod sets) were displayed in both seasons. Season-specific responses consisted only of a transient increase in plasma corticosterone (B) during the breeding season that was evident on Day 4 and had subsided by Day 10. Pushups were displayed in response to both genders during both seasons, although the frequency of pushups was significantly higher in response to females than to males during the postbreeding season. The coloration of residents did not change in response to male intruders during the postbreeding season or to females during either season. These results define the gender and seasonal specificity of hormonal, behavioral, and coloration responses of resident male S. undulatus in social interactions with conspecifics. Thus, our results clarify the biological significance of these responses in terms of potentially aggressive versus courtship interactions and breeding versus postbreeding contexts.     

Prey availability affects territory size,but not territorial display behavior,in green anole lizards

The availability of food resources can affect the size and shape of territories, as well as the behaviors used to defend territories, in a variety of animal taxa. However, individuals within a population may respond differently to variation in food availability if the benefits of territoriality vary among those individuals. For example, benefits to territoriality may differ for animals of differing sizes, because larger individuals may require greater territory size to acquire required resources, or territorial behavior may differ between the sexes if males and females defend different resources in their territories. In this study, we tested whether arthropod abundance and biomass were associated with natural variation in territory size and defense in insectivorous green anole lizards, Anolis carolinensis. Our results showed that both male and female lizards had smaller territories in a habitat with greater prey biomass than lizards in habitats with less available prey, but the rates of aggressive behaviors used to defend territories did not differ among these habitats. Further, we did not find a relationship between body size and territory size, and the sexes did not differ in their relationships between food availability and territory size or behavioral defense. Together, these results suggest that differences in food availability influenced male and female territorial strategies similarly, and that territory size may be more strongly associated with variation in food resources than social display behavior. Thus, anole investment in the behavioral defense of a territory may not vary with territory quality.     

Stress during adolescence enhances locomotor sensitization to nicotine in adulthood in female, but not male, rats

A wide body of research has indicated that perinatal exposure to stressors alters the organism, notably by programming behavioral and neuroendocrine responses and sensitivity to drugs of abuse in adulthood. Recent evidence suggests that adolescence also may represent a sensitive period of brain development, and yet there has been little research on the long-lasting effects of stressors during this period. We investigated the effects of pubertal social stress (PS; daily 1-h isolation followed by pairing with a new cage mate on postnatal days 33-48) on locomotor sensitization to injections of nicotine and corticosterone response to restraint stress when the rats were adults (approximately 3 weeks after PS). There were no differences among the groups in locomotor activity to injections of saline. However, PS females had enhanced locomotor sensitization to repeated doses of nicotine compared to control (non-stressed; NS) females, whereas PS males and NS males did not differ. PS enhanced the corticosterone response to restraint in male rats previously sensitized to nicotine and decreased the corticosterone response in nonsensitized male rats. In contrast, PS females and NS females did not differ in plasma corticosterone levels in response to restraint stress, but NS females showed enhanced corticosterone release to restraint after sensitization to nicotine. Thus, during adolescence, social stressors can have long-lasting effects, and the effects appear to differ for males and females.     

Effects of reproductive status on behavioral and endocrine responses to acute stress in a biparental rodent, the California mouse (Peromyscus californicus)

In several mammalian species, lactating females show blunted neural, hormonal, and behavioral responses to stressors. It is not known whether new fathers also show stress hyporesponsiveness in species in which males provide infant care. To test this possibility, we determined the effects of male and female reproductive status on stress responsiveness in the biparental, monogamous California mouse (Peromyscus californicus). Breeding (N = 8 females, 8 males), nonbreeding (N = 10 females, 10 males) and virgin mice (N = 12 females, 9 males) were exposed to a 5-min predator-urine stressor at two time points, corresponding to the early postpartum (5-7 days postpartum) and mid/late postpartum (19-21 days postpartum) phases, and blood samples were collected immediately afterwards. Baseline blood samples were obtained 2 days prior to each stress test. Baseline plasma corticosterone (CORT) concentrations did not differ among male or female groups. CORT responses to the stressor did not differ among female reproductive groups, and all three groups showed distinct behavioral responses to predator urine. Virgin males tended to increase their CORT response from the first to the second stress test, while breeding and nonbreeding males did not. Moreover, virgin and nonbreeding males showed significant behavioral changes in response to predator urine, whereas breeding males did not. These results suggest that adrenocortical responses to a repeated stressor in male California mice may be modulated by cohabitation with a female, whereas behavioral responses to stress may be blunted by parental status.     

Asymmetric learning to avoid heterospecific males in Mesocricetus hamsters

If a female mates with a male of a closely related species, her fitness is likely to decline. Consequently, females may develop behavioral mechanisms to avoid mating with heterospecific males. In some species, one such mechanism is for adult females to learn to discriminate against heterospecific males after exposure to such males. We have previously shown that adult, female Syrian hamsters (Mesocricetus auratus) learn to discriminate against male Turkish hamsters (Mesocricetus brandti) after exposure to a single heterospecific male during 8 days across a wire-mesh barrier. Here we repeated that experiment but this time we exposed female Turkish hamsters to a male Syrian hamster for 8 days and then measured sexual and aggressive behaviors towards that heterospecific male and towards a conspecific male. In contrast to female Syrian hamsters, female Turkish hamsters did not differ in their latency to go into lordosis or in any measure of aggression towards either type of male. Female Turkish hamsters spent less time in lordosis with the heterospecific male, but the percentage of trials in which females copulated with conspecific and heterospecific males did not differ. When comparing females from both species that had been exposed to a heterospecific male for 8days, female Syrian hamsters copulated less and were more aggressive towards the heterospecific male compared to the behavior of female Turkish hamsters. We discuss how this asymmetric response between females of the two species may be due to the much larger geographical range of Turkish hamsters compared to Syrian hamsters.     

Wood duck hatch date: relationship to pairing chronology, plasma luteinizing hormone, and steroid hormones during autumn and winter

We examined variation in courtship activity and hormone levels of male and female wood ducks (Aix sponsa) in relation to hatch date. Young wood ducks were assigned in groups of 8 (4 males and 4 females) to 4 experimental pens; 2 pens contained early-hatched ducks (3-12 April) and 2 pens contained late-hatched ducks (7-16 June). Courtship behaviors occurred less frequently in October and November than in December and January-February for both early- and late-hatched groups. Early-hatched wood ducks participated in courtship more frequently and formed pair bonds sooner than late-hatched individuals. Testosterone (T) and androstenedione (AD) levels of males did not differ between treatment groups; however, average levels of luteinizing hormone (LH) were greater for early-hatched males. Differences in LH occurred in 2 of 12 samples (6 October and 3 November); otherwise, LH did not vary by treatment. Levels of T and LH in males varied temporally, but there was no significant temporal variation in AE levels. Temporal variation in hormone levels was similar for early- and late-hatched males. Estradiol (E), progesterone (P), and LH levels of females did not differ between treatment groups. There was temporal variation in levels of E and LH, but not of P; this variation was similar for early- and late-hatched females. These data indicate that behavioral differences occurring temporally and between early- and late-hatched wood ducks were not related to corresponding differences in hormone levels.     

Lack of sexual experience does not reduce the responses of LH, estrus or fertility in anestrous goats exposed to sexually active males

We investigated whether LH secretion, estrous behavior and fertility would differ between sexually inexperienced and experienced anestrous goats exposed to the males. Male goats were rendered sexually active during the reproductive rest season by exposure to 2.5 months of artificial long days. Two groups of anovulatory sexually inexperienced and sexually experienced does were exposed to males during 15 days (n = 20 per group). LH pulsatility was determined every 15 min from 4 h before to 8 h after introducing males (Day 0). Estrous behavior was recorded twice daily. Pregnancy rates were determined on Day 50. Fertility was determined at parturition. Male sexual behavior was registered on days 1 and 2 during 1 h. Before introducing the males, the number of LH pulses did not differ between groups. After introduction of the males, all females increased their LH pulsatility, but the number of pulses did not differ between sexually inexperienced and experienced goats. The proportion of females displaying estrous behavior with a high pregnancy rate and fertility did not differ between inexperienced and experienced goats. The sexual behavior of the males did not differ significantly between those interacting with sexually inexperienced or experienced goats. We conclude that goats can show substantial endocrine and reproductive responses to males, even in the absence of previous sexual experience, when sexually active bucks are used.     

Differential effects of prenatal testosterone timing and duration on phenotypic and behavioral masculinization and defeminization of female sheep

The process of sexual differentiation leaves genetically female individuals at risk of being masculinized by exogenous androgens. Previous research with sheep indicates that exposure to excess testosterone from Gestational Day (GD) 30 to GD 90 of the 147-day gestation masculinizes and defeminizes behavior as well as genitalia. Lower doses and shorter durations produce animals with varying degrees of genital virilization and alterations of the hypothalamic-pituitary-gonadal axis, but to our knowledge, the effects on complex behavior and its prediction by the amount of external virilization have not been explored. Previous research in rodents has suggested that sexual differentiation of the central nervous system and the external genitalia can be dissociated. Therefore, we hypothesized that the extent of virilization of external genitalia would not be predictive of the lack of female-typical, or the presence of male-typical, mating behavior. To test this hypothesis, we compared control females, females exposed to exogenous testosterone from GD 30 to GD 90 (T60 females) that have virilized genitalia, and females exposed to testosterone from GD 60 to GD 90 (T30 females) that have female-typical genitalia. Both natural behavioral estrus in the flock and hormonally controlled behavioral tests were used to explore reproductive behavior. The T60 and T30 females exhibited more masculinized reproductive behavior than the controls; however, the T30 females also exhibited feminine behavior. Neither testosterone-treated group was receptive or was mounted at rates comparable to those of controls. These data illustrate that variation in the timing or duration of exposure to prenatal testosterone during a critical period for masculinization can have variable effects on defeminization and that the effects of testosterone on genitalia are not entirely predictive of behavior.     

Prenatal exposure to androgen influences morphology and aggressive behavior of male and female mice

To assess the effects of prenatal exposure to androgen on adult aggressiveness in mice, pregnant mice were given injections of 1.5 mg testosterone propionate (TP) or oil from Days 12 to 16 of pregnancy. All offspring were gonadectomized on the day of birth. Neonatal treatment occurred on the day following birth and consisted of one-half of the animals from each prenatal treatment group being injected with 100 μg TP while the other half were injected with oil, yielding four Prenatal/Neonatal treatment groups for each sex. On postnatal Day 60, all offspring were given subcutaneous implants of encapsulated testosterone (T) and tested for 10 min every other day against a male opponent until aggression was observed. Female offspring of TP-treated mothers were indistinguishable from males on external examination at birth. The duration of exposure to T required to induce aggression provides an index of the sensitivity of the neural substrate to T. When arranged from the most sensitive to the least sensitive to the aggression inducing action of T, the four Prenatal/Neonatal treatment groups of females were significantly different from each other: Group TP/TP > Group OIL/TP > Group TP/OIL > Group OIL/OIL. A similar pattern was observed for the male offspring. There were no differences in the proportion of animals per group that exhibited aggression (virtually all animals fought) or the intensity of aggression once exhibited. The results demonstrate that morphological and behavioral masculinization can occur in response to exposure to androgen during prenatal as well as neonatal life in mice.     

Yolk testosterone modulates persistence of neophobic responses in adult zebra finches, Taeniopygia guttata

Individual differences in animal behavior can be attributed to genetic as well as non-genetic influences. One mechanism by which the behavioral phenotype of an individual can be shaped is via transmission of maternal sex steroids. In this study, we examined the role of yolk testosterone (T) in controlling neophobia in 9-month-old, sexually mature zebra finches (Taeniopygia guttata). Offspring hatched from either T-treated or control eggs were subjected to a sequential series of behavioral tests in which we measured the neophobic response and its persistence towards two unfamiliar stimuli. Birds from T-treated and control eggs did not differ in their latencies to approach and eat a novel food source during their first encounter. However, egg treatment affected subsequent habituation. Latencies decreased in both groups over a habituation period of 5 days, but considerably more so in T-offspring. Although males appeared to approach novel food faster than females, there was no overall sex effect during the habituation period. When a novel object was added in combination with the previously learned food stimulus, this caused an behavioral shift in approach latencies. In males, control offspring had significantly shorter latencies than T-offspring, whereas there was no difference among females. The latency to eat in the same test was not significantly affected by sex or egg treatment. Our results demonstrate long-term effects of prenatal T on neophobic responses in adult zebra finches. We hypothesize that prenatal T may be one underlying mechanism for individual differences routine formation.     

Perinatal exposure to 19-Nor-17α-ethynyltestosterone (norethindrone) influences morphology and aggressive behavior of female mice

Pregnant mice were injected with 50 or 100 μg of the synthetic progestin 19-nor-17α-ethynyltestosterone (NET) on Days 14 through 17 of gestation. Others received only the vehicle or were left undisturbed. Exposure to NET during the prenatal period increased anogenital distance of the female offspring as measured on Day 60 but did not influence the duration of adult testosterone (T) exposure required to activate male-like intraspecific fighting behavior. In a second experiment, female mice were exposed to NET either prenatally, postnatally (Day 1), or pre- and postnatally. Exposure during both pre- and postnatal periods increased anogenital distance but only the exposure during the postnatal period enhanced later behavioral sensitivity to the aggression-activating property of T. Thus, NET is similar to testosterone in its ability to virilize morphology and behavior, although, at the dosages administered, behavioral alternation only occurred as a result of treatment during the neonatal period.     

Androgen threshold to activate copulation differs in male rats prenatally exposed to alcohol, stress, or both factors.

Few male rats prenatally exposed to a combination of alcohol and stress copulate spontaneously. This study determined adult sensitivity to testosterone (T) in males prenatally exposed to alcohol, to stress, or to both factors. Sexually naive males were tested with receptive females following castration and implantation of 20-, 30-, or 45-mm Silastic T-filled capsules. Serum T levels provided by these implants were measured. The behavior shown by males exposed only to prenatal alcohol did not differ from untreated control animals at any T dosage. Prenatal stress alone diminished the copulatory potential below control levels only when the intermediate T dosage was provided. Few males exposed to both alcohol and stress copulated under the lowest or the intermediate dose of adult T replacement, but most ejaculated normally when the largest capsule was implanted. The threshold to the sexual behavior-activating-properties of adult T exposure was moderately raised by prenatal stress but was severely affected when prenatal stress was combined with alcohol. We conclude that a diminished sensitivity to androgen in adulthood underlies some copulatory deficits resulting from treatments that alter fetal T levels. Such deficits may be concealed when behavior is evaluated in gonadally intact animals.     

Behavioral masculinization is independent of genital masculinization in prenatally androgenized female rhesus macaques

Genetic female fetuses were exposed transplacentally to testosterone propionate injected into their mothers either early (Days 40 through 64) or late (Days 115 through 139) in gestation. Early and late androgenized females (EAFs and LAFs, respectively) were raised with normal males and females that served as criteria for evaluating degree of behavioral masculinization induced by the prenatal androgen. EAFs were genitally virilized and LAFs were not. Males and untreated females differed reliably on five behavioral measures: males showed more mother-mounting, more peer-mounting, more rough play with peers, a preference for initiating play with male partners, and less grooming of mothers. Neither type of prenatally androgenized female showed masculinization of all five types of behavior. Compared with females, EAFs showed more mother-mounting, more peer-mounting, less mother-grooming, did not differ from females in rough play, and did not manifest a preference for male partners. LAFs, like females, groomed but did not mount their mothers, and did not show a preference for male partners; but unlike females they showed elevated rough play and mounting with peers. EAFs showed a statistically significant delay in puberty onset (menarche), but LAFs did not. Mothers inspected genitalia of their offspring more often if they were males than if they were females. Mothers of EAFs inspected their offspring's genitalia as often as mothers of males, but mothers of LAFs did not. No aspect of maternal behavior was associated with either the amount or kind of masculine behavior shown toward peers. We interpret the results to mean that genital virilization is independent of, and largely irrelevant to, the expression of those behavioral traits that characterize the juvenile male social role. Moreover, the individual behavior traits that are components of the juvenile male role are independently regulated by the organizing actions of androgen and have separable critical periods. Of the two major traits, mounting peers and rough play with peers, the latter has a greater requirement for androgenic stimulation late in prenatal life.     

Otoacoustic emissions,auditory evoked potentials and self-reported gender in people affected by disorders of sex development (DSD)

Both otoacoustic emissions (OAEs) and auditory evoked potentials (AEPs) are sexually dimorphic, and both are believed to be influenced by prenatal androgen exposure. OAEs and AEPs were collected from people affected by 1 of 3 categories of disorders of sex development (DSD) — (1) women with complete androgen insensitivity syndrome (CAIS); (2) women with congenital adrenal hyperplasia (CAH); and (3) individuals with 46,XY DSD including prenatal androgen exposure who developed a male gender despite initial rearing as females (men with DSD). Gender identity (GI) and role (GR) were measured both retrospectively and at the time of study participation, using standardized questionnaires. The main objective of this study was to determine if patterns of OAEs and AEPs correlate with gender in people affected by DSD and in controls. A second objective was to assess if OAE and AEP patterns differed according to degrees of prenatal androgen exposure across groups. Control males, men with DSD, and women with CAH produced fewer spontaneous OAEs (SOAEs) – the male-typical pattern – than control females and women with CAIS. Additionally, the number of SOAEs produced correlated with gender development across all groups tested. Although some sex differences in AEPs were observed between control males and females, AEP measures did not correlate with gender development, nor did they vary according to degrees of prenatal androgen exposure, among people with DSD. Thus, OAEs, but not AEPs, may prove useful as bioassays for assessing early brain exposure to androgens and predicting gender development in people with DSD.     

The production of and sensitivity to cues that delay puberty and prolong subsequent oestrous cycles in female mice are influenced by prior intrauterine position

Four experiments were conducted with CF-1 house mice (Mus domesticus) to examine the relationship between a female's prior intrauterine position (2M = between 2 male fetuses, 1M = next to one male fetus, 0M = not next to a male fetus) and the timing of puberty and length of subsequent oestrous cycles under a variety of housing conditions. The results of Exp. 1 confirmed that the presence of males was required for females to enter puberty and exhibit regular oestrous cycles, regardless of prior intrauterine position. When housed individually after weaning either with a male in the cage or separated by a wire mesh partition, 0M-females ovulated and mated at a younger age and had shorter post-pubertal oestrous cycles than did 2M-females (1M-females were intermediate between 0M- and 2M-females). Other females were also housed after weaning with a male or separated by a wire mesh partition from a male in a variety of social environments (4 0M-females and 1 2M-female, 4 2M-females and 1 0M-female, 5 1M-females, 3 0M-females and 3 2M-females). The objective of these different housing conditions was to determine whether prior intrauterine position influenced the transmission of and/or sensitivity to cues that delay puberty and prolong subsequent oestrous cycles. Relative to 2M-females, 0M-females both produced more potent cues and were more sensitive to the cues (again, 1M-females were intermediate between 0M- and 2M-females). Specifically, in the presence of other females, puberty was delayed and post-pubertal oestrous cycles were prolonged to the greatest extent in 0M-females. Prior intrauterine position is therefore a source of individual variation in the production of and sensitivity to cues that modulate the timing of puberty and the length of subsequent oestrous cycles in female mice. The findings suggest that prenatally-androgenized 2M-females may have a reproductive advantage over other females at high population densities.     

Effect of in-utero exposure to diethylstilboestrol on ontogeny of uterine glands in neonatal mice

Pregnant CD-1 mice were injected with diethylstilboestrol (10 micrograms/kg body weight) in 0.1 ml maize oil, or maize oil alone, on Day 16 of gestation. Six experimental and 6 control female progeny were killed daily from birth until Day 7 and uterine tissues were examined by light microscopy. In-utero exposure to diethylstilboestrol resulted in hypertrophy of luminal epithelial cells and premature formation of uterine glands. The initial sign of uterine gland formation was invagination of the uterine surface epithelial cell layer into the underlying connective tissue stroma. A temporal difference occurred between control animals and those exposed to diethylstilboestrol: uterine gland formation first occurred in experimental progeny on Day 4, but not until Day 5 in control progeny. Uterine glands which extended deep into the connective tissue stroma to the myometrium were present in diethylstilboestrol-treated progeny by Day 7, but remained in the superficial endometrial connective tissue stroma in control animals. The results indicate that prenatal exposure of mice to diethylstilboestrol causes uterine epithelial cell hypertrophy at birth and the premature formation of uterine glands during the first week of neonatal uterine development.     

Prenatal stress reduces fertility of male offspring in mice, without affecting their adult testosterone levels

The male offspring of mice stressed by crowding during the final third of pregnancy showed reductions in sexual behavior and fertility. When paired with receptive females, their latencies to mount and to achieve intromission and ejaculation were greater than controls, and 30% of them failed to ejaculate in the 100-min test. When housed continuously for 4 days with females, 31% of them failed to impregnate their partners, compared with 4% of controls. The sexual receptivity of the untreated females paired with prenatally stressed males was not affected. Resting testosterone levels of prenatally stressed males did not differ from those of controls, and the pattern of rise and fall of testosterone during a 60-min interaction with a female showed only minor differences. The results suggest a central, rather than peripheral, mediation of the behavioral effects of prenatal stress.     

Serum prolactin concentrations and the initiation of maternal behavior in the rat

Maternal behavior and serum prolactin were measured in pregnant and virgin female rats. Pregnant rats were either ovariectomized or shamovariectomized on Day 17 of pregnancy, while virgin females were ovariectomized at the same age. Two days after surgery nests were rated and the three treatment groups were tested for responsiveness to rat pups. Both pregnant treatment groups built superior nests compared to the virgin group and also responded more frequently to rat pups within a 1 hr test period than the virgin controls. In addition, significantly more ovariectomized pregnant subjects responded to pups than did intact pregnant females. Serum prolactin levels did not differ among the three treatments nor did exposure to pups affect serum prolactin levels. In each treatment group serum prolactin was less than 15 ng/ml, well below the 139.7 ng/ml mean found on Day 23 of pregnancy. These data suggest that high levels of serum prolactin during late pregnancy are not essential for the initiation of maternal behavior in the rat.     

Influences of pre- and postnatal testosterone treatment on defeminization of sexual receptivity in pigs

Sexual receptivity was evaluated in female and male pigs that had experienced varying periods of exposure to testosterone pre- and postnatally. For prenatal exposure, pregnant sows were treated with testosterone propionate (TP) from Day 29-35 or Day 39-45 of gestation at a dosage that caused virilization of the external genitalia of their female offspring. Eighty-three percent of the females that received TP prenatally had regular estrous cycles, but reached puberty later than control females. Only 26% of the females that received TP both pre- and postnatally (4-6 mo of age) were observed in estrus by 10 mo of age. After ovariectomy and acute treatment with estradiol benzoate (EB), the proportion of females that showed the immobilization response (receptivity) was similar for all groups of females independent of pre- or postnatal TP treatment. Females treated prenatally from Day 39-45 showed the immobilization response for fewer days after treatment with a high dosage of EB than did controls. On the basis of these observations, we conclude that receptivity in female pigs is not affected greatly by testosterone treatment at the stages of development that were investigated. Males castrated at birth and treated with a single injection of EB after 9.5 mo showed the immobilization response. In contrast, few males castrated at 8 mo or castrated at birth and treated with TP from 3 to 6 mo showed the immobilization response after EB treatment. These observations provide direct evidence for a postnatal component of testosterone-dependent defeminization of receptivity in male pigs.     

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.