Early Vaginal Opening Research Articles

Neonatal androgenization-induced early endocrine-metabolic and ovary misprogramming in the female rat.

Androgen excess predisposes the organism to develop metabolic-endocrine and reproductive dysfunctions, among them the development of a phenotype resembling that of human Polycystic Ovary Syndrome (PCOS). We analyzed the impact of a single neonatal (5day-old) testosterone propionate (TP; s.c. 1.25mg/female pup) dose on: a) several metabolic-endocrine activities and b) ovarian steroidogenic and granulosa cell (GC) functions and also follicular population in juvenile and adult TP and control (CT) rats. Compared to CT rats, TP animals were characterized by: a) accelerated growth, hyperadiposity and hyperleptinemia, b) very early (pre-weaning age) vaginal opening, c) hyperinsulinemia in adult life, d) dysfunctional ovarian steroidogenesis, e) conserved GC functionality in both juveniles (in vitro) and adults (in vivo), and f) estrous cycles arrested at estrus. Finally, histological studies of the ovaries indicated that in TP (vs. CT) rats: i) primary and antral follicle frequencies were 3- and 15-fold higher and lower, respectively, in juveniles and ii) secondary and atretic follicle frequencies were 3- and 5-fold lower and higher, respectively, in adults. Large cystic images without corpus luteum were observed in the ovaries from adult TP rats only. Our results strongly suggest that transient neonatal hyperandrogenemia induced early misprogramming of metabolic-endocrine and ovarian (steroidogenesis/folliculogenesis) functions. Conversely, TP rats preserved their ovary GC endocrine function. Our results further support the high risk of developing ovarian hyperstimulation syndrome for infertile women with transient/chronic hyperandrogenemia (PCOS) subjected to assisted reproductive technologies.

Elevated hypothalamic aromatization at the onset of precocious puberty in transgenic female mice hypersecreting human chorionic gonadotropin: Effect of androgens

Transgenic female mice overexpressing the α- and β- subunits of human chorionic gonadotropin (hCGαβ+) exhibited precocious puberty, as evidenced by early vaginal opening. Chronically elevated hCG in 21-day-old hCGαβ+ females stimulated gonadal androgen production, which exerted negative feedback over the endogenous gonadotropin synthesis, and activated the hypothalamic GnRH pulsatility and gene expression. Transgenic females also exhibited elevated hypothalamic aromatization in the preoptic area (POA), which is the sexually-differentiated area that controls the LH surge in adulthood. Ovariectomy at 14days of age was unable to rescue this phenotype. However, the blockade of androgen action by flutamide from postnatal day 6 onwards reduced the aromatase levels in the POA of hCGαβ+ females. Our results suggest that early exposure of females to androgen action during a critical period between postnatal days 6–14 induces sex-specific organizational changes of the brain, which affect the aromatase expression in the POA at the onset of precocious puberty.

Abstract 5452: In utero exposure to bisphenol A promotes mammary tumor development in MMTV-erbB-2 transgenic mice

Abstract Increasing evidence indicates that environmental exposure plays a critical role in breast cancer etiology. Bisphenol A (BPA), the building block of polycarbonate plastics with estrogenic activity, is one of the most common chemicals exposed in daily life. Recent studies suggest a link between BPA exposure and increased breast cancer risk. However, specific conditions and the underlying mechanisms of BPA exposure associated breast cancer risk remain unclear. This study aimed to investigate the effects of in utero exposure to BPA on mammary tumor development in MMTV-erbB-2 transgenic mice, which exemplifies a scenario of gene-environmental interaction. Since erbB-2 is amplified/overexpressed in approximately 30% of breast cancer, which has been associated with poor prognosis and therapeutic resistance, use of this clinically relevant mammary tumor model is of high translational value. In this study, pregnant MMTV-erbB-2 mice were subcutaneously injected with 0, 25 ng, 250 ng and 250 µg/Kg body weight BPA daily between day 13 and day 19 of gestation. We found that in utero exposed mice in the low dose (25 and 250 ng) groups, but not the high dose group (250 µg), displayed earlier vaginal opening and prolonged estrous phase, suggesting a dose dependent pro-estrogenic effect of in utero exposure to BPA. Whole mount analysis showed that mammary glands of mice with in utero exposure to low dose BPA had a longer ductal extension at 6 weeks and increased lateral branching/alveolar structures at 10 weeks, which was more significant in the 250 ng group but less evident in the high dose group. Molecular analysis of mammary tissues at 10 weeks indicated that in utero exposure to BPA increased phosphorylation/activation of erbB-2, EGFR, erbB-3 Erk1/2 and Akt, and expression of erbB-3 and EGFR. Signaling of estrogen receptor (ER) pathway was upregulated concomitantly, as reflected by increased expression and phosphorylation of ERα and upregulation of cyclin D1 and c-myc. More importantly, mice in the 250 ng group but not the high dose group, developed tumor earlier than the mice in the control group, suggesting that dose is a critical factor affecting the outcomes. These data demonstrated that in utero exposure to BPA may promote mammary tumor development in erbB-2 transgenic mice in a dose dependent manner. Concomitant activation of the erbB-2 and ER pathways in the premalignant tissues suggests that induction of ER-erbB-2 crosstalk may play a critical role in mammary tumorigenesis promoted by in utero exposure to BPA, which underscores the significance of gene-environment interaction in early exposure to BPA associated breast cancer risk. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 5452. doi:1538-7445.AM2012-5452

Fibroblast Growth Factor Signaling Deficiencies Impact Female Reproduction and Kisspeptin Neurons in Mice1

Fibroblast growth factor (FGF) signaling is essential for the development of the gonadotropin-releasing hormone (GnRH) system. Mice harboring deficiencies in Fgf8 or Fgf receptor 1 (Fgfr1) suffer a significant loss of GnRH neurons, but their reproductive phenotypes have not been examined. This study examined if female mice hypomorphic for Fgf8, Fgfr1, or both (compound hypomorphs) exhibited altered parameters of pubertal onset, estrous cyclicity, and fertility. Further, we examined the number of kisspeptin (KP)-immunoreactive (ir) neurons in the anteroventral periventricular/periventricular nuclei (AVPV/PeV) of these mice to assess if changes in the KP system, which stimulates the GnRH system, could contribute to the reproductive phenotypes. Single hypomorphs (Fgfr1(+/-) or Fgf8(+/-)) had normal timing for vaginal opening (VO) but delayed first estrus. However, after achieving the first estrus, they underwent normal expression of estrous cycles. In contrast, the compound hypomorphs underwent early VO and normal first estrus, but had disorganized estrous cycles that subsequently reduced their fertility. KP immunohistochemistry on Postnatal Day 15, 30, and 60 transgenic female mice revealed that female compound hypomorphs had significantly more KP-ir neurons in the AVPV/PeV compared to their wild-type littermates, suggesting increased KP-ir neurons may drive early VO but could not maintain the cyclic changes in GnRH neuronal activity required for female fertility. Overall, these data suggest that Fgf signaling deficiencies differentially alter the parameters of female pubertal onset and cyclicity. Further, these deficiencies led to changes in the AVPV/PeV KP-ir neurons that may have contributed to the accelerated VO in the compound hypomorphs.

Effect of the aqueous extract of Justicia insularis T. Anders (Acanthaceae) on ovarian folliculogenesis and fertility of female rats.

Justicia insularis T. Anders (Acanthaceae) is a medicinal plant whose leaves and those of three other plants are mixed for the preparation of a concoction used to improve fertility and to reduce labour pains in women of the Western Region of Cameroon. Previous studies have demonstrated the inducing potential on ovarian folliculogenesis and steroidogenesis of the aqueous extract of the leaf mixture (ADHJ) of four medicinal plants (Aloe buettneri, Dicliptera verticillata, Hibiscus macranthus and Justicia insularis) among which the later represented the highest proportion. This study was aimed at evaluating the ovarian inducing potential of J. insularis in immature female rats. Various doses of the aqueous extract of J. insularis were daily and orally given, for 20 days, to immature female rats distributed into four experimental groups of twenty animals each. At the end of the experimental period some biochemical and physiological parameters of ovarian function were assayed. The administration of the aqueous extract of Justicia insularis significantly induced an early vaginal opening in all treated groups (P < 0.001) as well as an increase (at doses of 50 or 100 mg/kg) in the number of hemorrhagic points, Corpus luteum, implantation sites, ovarian weight, uterine and ovarian proteins. Ovarian cholesterol level (P < 0.05) significantly decreased in animals treated with the lowest dose (12.5 mg/kg). The evaluation of the toxicological effects of the extract on pregnancy showed that it significantly increased pre- and post-implantation losses, resorption index and decreased the rate of nidation as well as litter's weight. These results suggest that the aqueous extract of Justicia insularis induces ovarian folliculogenesis thus justifying its high proportion in the leaf mixture of ADHJ.

Androgen Receptor Antagonism and an Insulin Sensitizer Block the Advancement of Vaginal Opening by High-Fat Diet in Mice1

Reduced hypothalamic sensitivity to steroid negative feedback may contribute to the onset of puberty. In high fat-fed rodents, the timing of vaginal opening (VO) is advanced, suggesting that puberty begins earlier. Because obesity can increase androgens, which interfere with normal steroid feedback in adult females, we hypothesized that androgens reduce hypothalamic sensitivity to negative feedback during puberty and that blocking androgen action would prevent advanced VO in high fat-fed mice. Age at VO was examined in mice fed high-fat or low-fat diets from weaning and treated with the androgen receptor antagonist flutamide or vehicle (controls). VO was advanced in high-fat vs. low-fat controls, and flutamide blocked this advancement. VO was also delayed in low fat-fed flutamide-treated females, suggesting involvement of androgens in the timing of normal puberty. We next investigated if high-fat diet-induced insulin resistance contributes to early VO, as elevated insulin can stimulate androgen production. VO was examined in mice on either diet treated with the insulin sensitizer metformin. Metformin blocked high-fat advancement of VO but did not alter the timing of VO in low fat-fed mice. Insulin was elevated in high fat-fed females that had undergone VO compared with age-matched low fat-fed or metformin-treated animals on either diet that had not undergone VO. Together, these data suggest a model in which metabolic changes induced by high-fat diet, including transient increased circulating insulin, act in part by increasing androgen action to influence the timing of puberty in females.

Effects of in utero through lactational exposure to dicyclohexyl phthalate and p,p′-DDE in Sprague–Dawley rats

Anti-androgenic chemicals alter sexual differentiation by a variety of mechanisms, and the mechanisms between phthalate esters and p,p'-DDE are considered to be different. We performed an in utero through lactational exposure assay using dicyclohexyl phthalate and p,p'-DDE to investigate the sexual differentiation of these chemicals. Pregnant CD (SD) IGS rats were given dicyclohexyl phthalate or p,p'-DDE orally from gestational day (GD) 6 to postnatal day (PND) 20, and the endocrine-mediated effects in dams and their offspring were examined. The reproductive performance of offspring was also examined. The doses of dicyclohexyl phthalate were 0, 20, 100, and 500 mg/kg/day, and those of p,p'-DDE were 5, 15, and 50mg/kg/day. Using the dicyclohexyl phthalate, a dam in the 500 mg/kg group showed dystocia and died. The viability index of offspring on PND 4 decreased in the 500 mg/kg group. Prolonged preputial separation, reduced ano-genital distance, increased areolas/nipple retention, hypospadia, decreased ventral prostate and levator ani/bulbocavernosus muscle weights and decreased testicular germ cells were observed in male offspring in the 500 mg/kg group. In the assay using p,p'-DDE, decreased viability index of offspring on PND 21, prolonged preputial separation in male offspring and early vaginal opening in female offspring were observed in the 50mg/kg group. The copulation and fertility indices decreased in the reproductive performance of offspring in the 50mg/kg group. The endocrine-mediated effects were detected in offspring of dams given 100mg/kg dicyclohexyl phthalate, and in offspring of dams given 20mg/kg p,p'-DDE. Our results suggest that the in utero through lactational exposure assay is a useful method to detect endocrine-mediated effects and that further comparative study between this assay and two-generation reproductive test are necessary when this assay becomes one of the definitive tests.

Effect of genistein on the sexual maturation in immature female rats

Puopose : Exposure to dietary phytoestrogens such as genistein during early childhood is a growing public health concern. We examined the effect of early exposure to genistein on sexual maturation in immature rats. Methods : Weaning (3wk-old) Sprague-Dawley female rats were assigned to three groups (n=6 for each): fed by high dose of genistein (100 mg/kg/d), low dose of genistein (10 mg/kg/d) and control group. First vaginal opening (VO) day was observed. Structural alterations in the ovary and uterus were assessed by histologically. Expression of genes of ER, ER, and progesterone receptor (PR) in the ovary and uterus were investigated by RT-PCR. Results : High genistein group had earlier VO than control and low genistein group. Graafian follicles and corpora lutea were observed from the ovary of genistein-treated groups, while primary, secondary follicles and small atretic follicles were observed in the control group. Hypertrophy of luminal and glandular uterine epithelia were found in the genistein-treated groups while poor development of gland and fewer myometrial cell layers were evident in control group. In ovary, the transcriptional activities of ER and ER were higher in high genistein group than in controls. In uterus, the transcriptional activities of ER, ER and PR were higher in low genistein group than in controls. Conclusion : Acute exposure to genistein during the prepubertal period could activate the reproductive endocrine system resulting in the early onset of puberty in female rats. Further clinical investigation on the effect of genistein on the sexual maturation in children is warranted.

Early Maturation of Gonadotropin-Releasing Hormone Secretion and Sexual Precocity after Exposure of Infant Female Rats to Estradiol or Dichlorodiphenyltrichloroethane1

An increase in the frequency of pulsatile gonadotropin-releasing hormone (GnRH) secretion in vitro and a reduction in LH response to GnRH in vivo characterize hypothalamic-pituitary maturation before puberty in the female rat. In girls migrating for international adoption, sexual precocity is frequent and could implicate former exposure to the insecticide dichlorodiphenyltrichloroethane (DDT), since a long-lasting DDT derivative has been detected in the serum of such children. We aimed at studying the effects of early transient exposure to estradiol (E(2)) or DDT in vitro and in vivo in the infantile female rat. Using a static incubation system of hypothalamic explants from 15-day-old female rats, a concentration- and time-dependent reduction in GnRH interpulse interval (IPI) was seen during incubation with E(2) and DDT isomers. These effects were prevented by antagonists of alpha-amino-3-hydroxy-5-methylisoxazole-4 propionic acid (AMPA)/kainate receptors and estrogen receptor. Also, o,p'-DDT effects were prevented by an antagonist of the aryl hydrocarbon orphan dioxin receptor (AHR). After subcutaneous injections of E(2) or o,p'-DDT between Postnatal Days (PNDs) 6 and 10, a decreased GnRH IPI was observed on PND 15 as an ex vivo effect. After DDT administration, serum LH levels in response to GnRH were not different from controls on PND 15, whereas they tended to be lower on PND 22. Subsequently, early vaginal opening (VO) and first estrus were observed together with a premature age-related decrease in LH response to GnRH. After prolonged exposure to E(2) between PNDs 6 and 40, VO occurred at an earlier age, but first estrus was delayed. We conclude that a transient exposure to E(2) or o,p'-DDT in early postnatal life is followed by early maturation of pulsatile GnRH secretion and, subsequently, early developmental reduction of LH response to GnRH that are possible mechanisms of the subsequent sexual precocity. The early maturation of pulsatile GnRH secretion could involve effects mediated through estrogen receptor and/or AHR as well as AMPA/kainate subtype of glutamate receptors.

Gonadal defects and hormonal alterations in transgenic mice expressing a single chain human chorionic gonadotropin–lutropin receptor complex

To study the effects of premature and chronic ligand-mediated luteinizing hormone receptor (LHR) activation on reproductive development, we have generated transgenic mice expressing a genetically engineered, constitutively active yoked hormone-receptor complex (YHR), in which a fusion protein of human chorionic gonadotropin (hCG) is covalently linked to the N-terminus of rat LHR. YHR-expressing mice (YHR(+)) were analyzed at pre- and post-pubertal ages. Relative to wild type (WT) controls, male mice exhibited prepubertal increases in testosterone levels and seminal vesicle weights, and decreases in serum FSH, serum LH, testes weight, and the size of the seminiferous tubules. In adult male YHR(+) mice, testosterone and LH levels are not significantly different from WT controls. However, FSH levels and testes weights remain decreased. Female YHR(+) mice undergo precocious puberty with early vaginal opening, accelerated uterine development, enhanced follicular development, including the presence of corpora lutea, and an increase in serum progesterone. At 12 weeks of age, the ovary exhibits a relative increase in the amount of interstitial tissue, comprised of cells that are hypertrophic and luteinized, as well as follicles that are degenerating. Additionally, hemorrhagic cysts develop in approximately 25% of the transgenic mice. These degenerative changes are consistent with an aging ovary suggesting that CG-induced LHR activation in female mice leads to precocious sexual development and ovarian lesions. Taken together, these data indicate that the single chain YHR is functional in vivo and demonstrate that YHR(+) mice provide a novel system to further understand the reproductive consequences of aberrant LHR activation.

Prepubertal Zearalenone Exposure Suppresses N-Methyl-N-nitrosourea-Induced Mammary Tumorigenesis but Causes Severe Endocrine Disruption in Female Sprague-Dawley Rats

The effect of prepubertal exposure to zearalenone, an estrogenic mycotoxin, on N-methyl-N-nitrosourea (MNU)-induced mammary tumorigenesis and its influence on reproductive organs were examined in female Sprague-Dawley rats. Prepubertal rats were treated daily with either 0.1 or 10 mg/kg body weight of zearalenone between 15 and 19 days of age and compared with zearalenone-untreated animals (30 rats in each group). Six rats in each group were autopsied at 28 days of age, and their growth was evaluated. All remaining rats were given 50 mg/kg body weight MNU at 28 days of age and followed by monitoring for occurrence of mammary tumors ≥1 cm in diameter. Zearalenone did not affect body weight increase, and mammary glands showed similar development at 28 days of age (time at carcinogen administration). Both low- and high-dose zearalenone treatment significantly reduced incidence of mammary tumors ≥1 cm in diameter but did not influence latency (time between MNU administration and harvest of mammary tumor ≥1 cm in diameter) compared with untreated controls. Zearalenone dose dependently suppressed the number of histologically detected tumors (carcinomas) and multiplicity; the suppression was significant with high-dose treatment. However, high-dose treatment caused significantly earlier vaginal opening, both low- and high-dose treatment significantly caused irregularity of estrous cycle (persistent estrus or prolonged diestrus) at 8 to 10 wk of age, and zearalenone dose dependently increased the number of anovulatory rats (ovaries without newly formed corpora lutea) at 37 wk of age. Thus, short-duration zearalenone treatment in the prepubertal period suppressed subsequent mammary cancer occurrence but also severely damaged ovarian functions. This suggests that ingestion of foods containing zearalenone in the infantile period can have dramatic effects in later life.

Changes in reproductive organs of female rats treated with bisphenol A during the neonatal period

The effects of neonatal administration of estrogenic agents on rat reproductive organs were examined. Either bisphenol A (BPA; 0.25, 1, or 4 mg/pup) or 10 μg 17β-estradiol (E 2) was given subcutaneously to Sprague–Dawley female rats during the neonatal period from post-natal day (PND) 0 to 9. Animals ovariectomized at 80 days were given subcutaneous injections of 1 μg/kg E 2 for 3 days from PND 94 to 96. Clefts in the clitoris, early vaginal opening, irregular estrous cycles, a decrease in the area occupied by the corpora lutea (CL) in the ovary, and multiple cystic follicles in the ovary were found in the animals treated neonatally with 1 mg BPA. Uterine fluid weight measured after E 2 treatment on PND 94–96 was less than controls. In addition to these abnormalities, unusual body weight gains, persistent vaginal cornification, and lack of CL were observed in females treated neonatally with 4 mg BPA. The ovary weight on PND 80 and uterine fluid weight measured after E 2 treatment on PND 94–96 were less than controls for the 4 mg BPA group. Neonatal treatment with 10 μg E 2 induced similar abnormalities as found in the 4 mg BPA group. These results show that BPA when given during the neonatal period caused changes in female reproductive organs.

Low dose effect of in utero exposure to bisphenol A and diethylstilbestrol on female mouse reproduction

In utero exposure to bisphenol-A (BPA) at doses relevant to human consumption has been reported to accelerate weight gain and puberty in female mice, but the effect of low dose BPA on female reproduction has not been described. In this study, we investigated low dose effects of BPA on sexual maturation and reproduction in female ICR/Jcl mice. Pregnant ICR mice (F0) were injected (s.c.) with BPA (2 and 20 μg/kg), diethylstilbestrol (DES; 0.02, 0.2, and 2 μg/kg) or oil vehicle once per day from gestational days 11–17. For both female and male offspring (F1), body weights were measured on postnatal day (PND) 0 (the day of birth), 11, 22, and 60, and anogenital distance (AGD) was measured on PNDs 22 and 60. Pups were weaned at PND 22 and males were caged separately from females. Vaginal smears were taken daily beginning the day of vaginal opening for 30 days. The age at vaginal opening was significantly earlier in all exposed females except for 2 μg/kg BPA females compared to oil controls. Body weight at vaginal opening was lower than controls in all exposed females. The first vaginal estrus was earlier in all exposed females except for the 2 μg/kg BPA group females compared to controls. From PND 90 to 120, gestationally exposed F1 female mice were mated with unexposed males. Total numbers of pups and sex ratio in F1 mice exposed to BPA or DES, and those of their offspring (F2) were not different from controls in any treatment group. The present results indicate that prenatal exposure to low doses of BPA and DES induces early vaginal opening, but does not affect reproductive functioning at the first breeding.

Characterization of diethylstilbestrol-induced hypospadias in female mice.

The urethral duct and vagina are formed from the urogenital sinus (UGS) during the early neonatal period in mice. Neonatal estrogen exposure results in hypospadias, or the malpositioning of vaginal and urethral openings, with wide cleft clitoris. We sought to characterize diethylstilbestrol (DES) influence on UGS morphogenesis and hypospadias formation. Newborn (day 0) and 1-4-day-old female mice (ICR/Jcl) were given (s.c.) oil or 3.0 microg DES. Animals were killed 24 hr later; then hypospadias formation and epithelial apoptosis and proliferation within the developing UGS were assessed. DES did not alter normal UGS morphogenesis by day 1, in comparison with controls. However, hypospadias formation was observed in DES-treated mice by day 3. In these mice, the distal dorsal urethral duct appeared to fuse with and open into the lower vaginal solid cord region. Further, DES treatment produced a gradual significant increase in dorsal urethral epithelial apoptosis (P < 0.05) just prior to and during fusion and hypospadias formation. DES-induced urethral epithelial and sinus cord proliferation appeared significantly increased (P < 0.05) and unchanged, respectively, just prior to fusion. By day 5, DES-treated mice exhibited wide cleft clitoris. In addition, if DES was given on day 3 or 5, a gradual, distinct caudal shift in the vaginal-urethral junction was observed compared to mice treated on days 0-2. Although hypospadias was not induced when neonates were given DES on day 7, these mice continued to display early vaginal opening. Dose-response analysis indicated that 0.03 microg DES for 5 days is the lowest known critical dose for hypospadias induction. We have shown for the first time that DES-induced hypospadias onset may primarily be the result of changes in developing dorsal urethral epithelial cell apoptotic and proliferative activity, and that the location of DES-induced hypospadias formation is dependent on age at time of exposure.

Alterations of events related to ovarian function in tumor necrosis factor receptor type I knockout mice.

C57BL6 mice with targeted disruption of tumor necrosis factor (TNF) type 1 receptor (TNFRI) exhibited early vaginal opening when compared with wild-type mice (Day 24 +/- 0.6, n = 10, vs. 28 +/- 0.2, n = 11, P < 0.001). Equine CG- and hCG-treated TNFRI null mice ovulated more ova than did controls at two distinct times during the prepubertal period (Day 21: 13.4 +/- 1.7 vs. 7.3 +/- 1.4, P < 0.05; Day 25: 20.7 +/- 2.7 vs. 13.0 +/- 1.3, P < 0.05). Enhanced responsiveness to gonadotropins was not observed in adult mice. At 6 mo of age only 40% of TNFRI null mice exhibited estrous cycles. Those TNFRI null mice with estrous cycles spent significantly more time in diestrus and less time in estrus than controls. TNFRI null mice delivered significantly fewer litters (P < 0.001) than did C57BL6 and TNFRII null mice (TNFRI null 2.59 +/- 0.39; C57BL6 4.91 +/- 0.57; TNFRII null 5.40 +/- 0.60 litters/mo/10 pairs over a 12-mo period). Ovarian dispersates prepared on Day 25 of age from control and TNFRI knockout mice were cultured with and without 10 ng TNF/ml. TNF inhibited LH-stimulated progesterone and estradiol secretion by control dispersates but had no effect on cAMP. In contrast, TNF did not affect LH-stimulated accumulation of progesterone, estradiol, or cAMP by ovarian dispersates from TNFRI knockout mice. The results indicate that lack of TNFRI enhances ovarian responsiveness to gonadotropins during the prepubertal period and may be related to early vaginal opening. The lack of TNFRI is associated with early senescence and poor fertility. These studies demonstrate that the mechanism of TNF-mediated inhibition of steroidogenesis is most likely via TNFRI.

Sexual maturation and seasonal changes in plasma levels of sex steroids and fecundity of wild Norway rats selected for reduced aggressiveness toward humans

Parameters of sexual maturation and reproductive function were compared in two groups of Norway rats, one selected for reduced aggressiveness towards man and the other for aggressiveness. It was found that selection for reduced aggressiveness produced an acceleration of sexual maturation as judged by earlier vaginal opening in the female and presence of mature spermatozoa in the testes of the male as well by earlier prepubertal increase in plasma testosterone in the male. In domesticated adults, seasonal variations in the parameters of the reproductive function narrowed; they showed neither the winter decrease in sexual activity nor the marked fall in plasma testosterone and estradiol exhibited by their aggressive counterparts.

Effects of uterine position on rate of sexual development in female Mongolian gerbils

Examination of the rate of sexual maturation of 79 female gerbils from 32 Caesarean-delivered, foster-reared litters revealed that those females that, as fetuses, occupied uterine positions adjacent to one or two males (1M and 2M females) were less likely than those females occupying uterine positions not adjacent to males (0M females) to exhibit early vaginal opening. Our data further indicated that the uterine environment provided by late-maturing female gerbils biased their fetal daughters to themselves be late maturing to a greater extent than the daughters' uterine positions could explain. Daughters of late-maturing females in 1M and 2M uterine locations were more likely to be late-maturing than were daughters of early-maturing females in similar uterine locations. Because in female Mongolian gerbils age at vaginal opening is a powerful predictor of future reproductive strategy, the present results indicate that in female gerbils both prenatal maternal influence and uterine location are important determinants of future reproductive behaviors.

Effects of social environment on sexual maturation in female cotton rats (Sigmodon hispidus).

Age at sexual maturation among female cotton rats was measured in a variety of intraspecific social environments. Two experiments were conducted. In Experiment I, female cotton rats attained vaginal perforation and first estrus at younger ages and lighter body masses when paired from weaning with a conspecific juvenile male than when caged alone. In Experiment II, these findings were replicated and extended. Females housed with juvenile males matured at the youngest ages, while those housed alone matured at the oldest ages. Females housed with adult males matured at intermediate ages. Presence of a second juvenile female during maturation was significantly associated with early vaginal opening but not with early first estrus. The results of this study are discussed in context of similar social environmental effects on female sexual maturation that have been identified in other rodent species.

Nucleolus: Changes at puberty in neurons of the suprachiasmatic nucleus and the preoptic area

The nucleolus is directly involved in RNA and protein metabolism and may serve both as an indicator of growth and of increased metabolic activity. Neurons in the medial preoptic area (POA), the suprachiasmatic nuclei (SCN), and the caudate nucleus of the rat were examined at ages ranging from 1 day to 70 days. Mean cross-sectional areas of nucleoli were determined at each age, including samples taken from animals killed at the time of vaginal opening. The pattern of changes for the mean nucleolar area of the caudate nucleus, POA, and SCN indicated that the primary growth phase was completed between 15 and 25 days of age. In each of the hypothalamic nuclei, but not the caudate nucleus, the nucleoli were found to be significantly increased at the age of vaginal opening. An estrogen-induced early vaginal opening also resulted in an increase in the area of the nucleolus in the hypothalamic nuclei. The position of the nucleolus in relation to the nuclear membrane does not appear to be as labile as the size of the nucleolus. The data show that significant changes in the cross-sectional areas of these neuronal nucleoli may be associated with the onset of puberty.

Plasma levels of LH and FSH in female rats with precocious puberty induced by hypothalamic lesions.

To determine whether precocious puberty induced by anterior hypothalamic lesions was associated with alterations in plasma gonadotropin levels during the prepubertal period, lesions were made by radiofrequency in the anterior hypothalamus of 15 day old female rats and plasma levels of gonadotropins were determined by radioimmunoassay (RIA) at different time intervals following the lesion. Nonoperated and sham-operated littermates were used as control. Lesioned animals showed precocious puberty as evidenced by early vaginal opening and first ovulation. They also became pregnant at an earlier age than controls and delivered their pups normally. Lesioned animals showed a more pronounced rate of increase in ovarian weight during the first 15 days following the lesion than that of control rats. Nevertheless, plasma FSH levels were not different in both groups. Plasma LH levels during the morning of the days preceding vaginal opening were somewhat higher in control rats than in lesioned animals. However, during the afternoons LH levels tended to be more elevated in lesioned rats than in controls, the difference being significant at Day 23. Estradiol injected subcutaneously inhibited compensatory ovarian hypertrophy in lesioned rats demonstrating the integrity of the estrogen negative feedback mechanism. The results were interpreted as not supporting the view that hypothalamic lesions induce precocious puberty by eliminating negative feedback to estrogen. The possibility arises that precocious puberty induced by hypothalamic lesions is determined by other mechanisms and an alternative explanation is suggested.