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Bisphenol A (BPA) is an endocrine disrupting, high volume production chemical found in a variety of products. Evidence of prenatal exposure has raised concerns that developmental BPA may disrupt sex-specific brain organization and, consequently, induce lasting changes on neurophysiology and behavior. We and others have shown that exposure to BPA at doses below the no-observed-adverse-effect level can disrupt the sex-specific expression of estrogen-responsive genes in the neonatal rat brain including estrogen receptors (ERs). The present studies, conducted as part of the Consortium Linking Academic and Regulatory Insights of BPA Toxicity program, expanded this work by examining the hippocampal and hypothalamic transcriptome on postnatal day 1 with the hypothesis that genes sensitive to estrogen and/or sexually dimorphic in expression would be altered by prenatal BPA exposure. NCTR Sprague-Dawley dams were gavaged from gestational day 6 until parturition with BPA (0-, 2.5-, 25-, 250-, 2500-, or 25 000-μg/kg body weight [bw]/d). Ethinyl estradiol was used as a reference estrogen (0.05- or 0.5-μg/kg bw/d). Postnatal day 1 brains were microdissected and gene expression was assessed with RNA-sequencing (0-, 2.5-, and 2500-μg/kg bw BPA groups only) and/or quantitative real-time PCR (all exposure groups). BPA-related transcriptional changes were mainly confined to the hypothalamus. Consistent with prior observations, BPA induced sex-specific effects on hypothalamic ERα and ERβ (Esr1 and Esr2) expression and hippocampal and hypothalamic oxytocin (Oxt) expression. These data demonstrate prenatal BPA exposure, even at doses below the current no-observed-adverse-effect level, can alter gene expression in the developing brain.

Concerns have been raised regarding the long-term impacts of early life exposure to the ubiquitous environmental contaminant bisphenol A (BPA) on brain organization. Because BPA has been reported to affect estrogen signaling, and steroid hormones play a critical role in brain sexual differentiation, there is also concern that BPA exposure could alter neural sex differences. Here, we examine the impact of subchronic exposure from gestation to adulthood to oral doses of BPA below the current no-observed-adverse-effect level (NOAEL) of 5 mg/kg body weight (bw)/day on estrogen receptor (ESR) expression in sexually dimorphic brain regions of prepubertal and adult female rats. The dams were gavaged daily with vehicle (0.3% carboxymethylcellulose), 2.5, 25, 260, or 2700 μg BPA/kg bw/day, or 0.5 or 5.0 μg ethinyl estradiol (EE)/kg bw/day from gestational day 6 until labor began. Offspring were then gavaged directly from the day after birth until the day before scheduled sacrifice on postnatal days 21 or 90. Using in situ hybridization, one or more BPA doses produced significant decreases in Esr1 expression in the juvenile female rat anteroventral periventricular nucleus (AVPV) of the hypothalamus and significant decreases in Esr2 expression in the adult female rat AVPV and medial preoptic area (MPOA), relative to vehicle controls. BPA did not simply reproduce EE effects, indicating that BPA is not acting solely as an estrogen mimic. The possible consequences of long-term changes in hypothalamic ESR expression resulting from subchronic low dose BPA exposure on neuroendocrine effects are discussed and being addressed in ongoing, related work., (Published by Oxford University Press on behalf of Toxicological Sciences 2014. This work is written by (a) US Government employee(s) and is in the public domain in the US.)

Perinatal life is a critical window for sexually dimorphic brain organization, and profoundly influenced by steroid hormones. Exposure to endocrine-disrupting compounds may disrupt this process, resulting in compromised reproductive physiology and behavior. To test the hypothesis that neonatal bisphenol A (BPA) exposure can alter sex-specific postnatal Esr2 (Erβ) expression in brain regions fundamental to sociosexual behavior, we mapped Esr2 mRNA levels in the principal nucleus of the bed nucleus of the stria terminalis (BNSTp), paraventricular nucleus (PVN), anterior portion of the medial amygdaloid nucleus (MeA), super optic nucleus, suprachiasmatic nucleus, and lateral habenula across postnatal days (PNDs) 0-19. Next, rat pups of both sexes were subcutaneously injected with 10 μg estradiol benzoate (EB), 50 μg/kg BPA (LBPA), or 50 mg/kg BPA (HBPA) over the first 3 days of life and Esr2 levels were quantified in each region of interest (ROI) on PNDs 4 and 10. EB exposure decreased Esr2 signal in most female ROIs and in the male PVN. In the BNSTp, Esr2 expression decreased in LBPA males and HBPA females on PND 10, thereby reversing the sex difference in expression. In the PVN, Esr2 mRNA levels were elevated in LBPA females, also resulting in a reversal of sexually dimorphic expression. In the MeA, BPA decreased Esr2 expression on PND 4. Collectively, these data demonstrate that region- and sex-specific Esr2 expression is vulnerable to neonatal BPA exposure in regions of the developing brain critical to sociosexual behavior in rat.

Bisphenol A (BPA) is a high volume production chemical used in polycarbonate plastics, epoxy resins, thermal paper receipts, and other household products. The neural effects of early life BPA exposure, particularly to low doses administered orally, remain unclear. Thus, to better characterize the dose range over which BPA alters sex specific neuroanatomy, we examined the impact of perinatal BPA exposure on two sexually dimorphic regions in the anterior hypothalamus, the sexually dimorphic nucleus of the preoptic area (SDN-POA) and the anterioventral periventricular (AVPV) nucleus. Both are sexually differentiated by estradiol and play a role in sex specific reproductive physiology and behavior. Long Evans rats were prenatally exposed to 10, 100, 1000, 10,000μg/kg bw/day BPA through daily, non-invasive oral administration of dosed-cookies to the dams. Offspring were reared to adulthood. Their brains were collected and immunolabeled for tyrosine hydroxylase (TH) in the AVPV and calbindin (CALB) in the SDN-POA. We observed decreased TH-ir cell numbers in the female AVPV across all exposure groups, an effect indicative of masculinization. In males, AVPV TH-ir cell numbers were significantly reduced in only the BPA 10 and BPA 10,000 groups. SDN-POA endpoints were unaltered in females but in males SDN-POA volume was significantly lower in all BPA exposure groups. CALB-ir was significantly lower in all but the BPA 1000 group. These effects are consistent with demasculinization. Collectively these data demonstrate that early life oral exposure to BPA at levels well below the current No Observed Adverse Effect Level (NOAEL) of 50mg/kg/day can alter sex specific hypothalamic morphology in the rat., (Copyright © 2013 Elsevier Inc. All rights reserved.)

Bisphenol A (BPA) exposure is ubiquitous, and in laboratory animals, early-life BPA exposure has been shown to alter sex-specific neural organization, neuroendocrine physiology, and behavior. The specific mechanisms underlying these brain-related outcomes, however, remain largely unknown, constraining the capacity to ascertain the potential human relevance of neural effects observed in animal models. In the perinatal rat brain, estrogen is masculinizing, suggesting that BPA-induced perturbation of estrogen receptor (ESR) expression may underpin later in-life neuroendocrine effects. We hypothesized that prenatal BPA exposure alters sex-specific ESR1 (ERα) and ESR2 (ERβ) expression in postnatal limbic nuclei. Sprague Dawley rats were mated and gavaged on gestational days (GDs) 6-21 with vehicle, 2.5 or 25 μg/kg bw/day BPA, or 5 or 10 μg/kg bw/day ethinyl estradiol. An additional group was restrained but not gavaged (naïve control). Offspring were sacrificed the day after birth to quantify ESR gene expression throughout the hypothalamus and amygdala by in situ hybridization. Relative to the vehicle group, significant effects of BPA were observed on ESR1 and ESR2 expression throughout the mediobasal hypothalamus and amygdala in both sexes. Significant differences in ESR expression were also observed in the mediobasal hypothalamus and amygdala of the naïve control group compared with the vehicle group, highlighting the potential for gavage to influence gene expression in the developing brain. These results indicate that ESR expression in the neonatal brain of both sexes can be altered by low-dose prenatal BPA exposure.

Hypothalamic neurons, which produce the kisspeptin family of peptide hormones (Kp), are critical for initiating puberty and maintaining estrous cyclicity by stimulating gonadotropin-releasing hormone (GnRH) release. Conversely, RFamide-related peptide-3 (RFRP3) neurons inhibit GnRH activity. It has previously been shown that neonatal exposure to bisphenol A (BPA) can alter the timing of female pubertal onset and induce irregular estrous cycles or premature anestrus. Here we tested the hypothesis that disrupted ontogeny of RFamide signaling pathways may be a mechanism underlying advanced puberty. To test this, we used a transgenic strain of Wistar rats whose GnRH neurons express enhanced green fluorescent protein. Pups were exposed by daily subcutaneous injection to vehicle, 17beta-estradiol (E2), 50 μg/kg BPA, or 50 mg/kg BPA, from Postnatal Day (PND) 0 through PND 3, and then cohorts were euthanized on PNDs 17, 21, 24, 28, and 33 (5-8 animals per age per exposure; males were collected on PNDs 21 and 33). Vaginal opening was advanced by E2 and 50 μg/kg BPA. On PND 28, females exposed to E2 and 50 μg/kg BPA had decreased RFRP-3 fiber density and contacts on GnRH neurons. RFRP3 perikarya were also decreased in females exposed to 50 μg/kg BPA. Data suggest that BPA-induced premature puberty results from decreased inhibition of GnRH neurons.

Developmental exposure to Bisphenol A (BPA), a component of polycarbonate and epoxy resins, has been purported to adversely impact reproductive function in female rodents. Because neonatal life is a critical window for the sexual dimorphic organization of the hypothalamic-pituitary-gonadal (HPG) axis, interference with this process could underlie compromised adult reproductive physiology. The goal of the present study was to determine if neonatal BPA exposure interferes with sex specific gene expression of estrogen receptor alpha (ERα), ER beta (ERβ) and kisspeptin (Kiss1) in the anterior and mediobasal hypothalamus. Long Evans (LE) neonatal rats were exposed to vehicle, 10μg estradiol benzoate (EB), 50mg/kg BPA or 50μg/kg BPA by subcutaneous injection daily from postnatal day 0 (PND 0) to PND 2. Gene expression was assessed by in situ hybridization on PNDs 4 and 10. Within the anterior hypothalamus ERα expression was augmented by BPA in PND 4 females, then fell to male-typical levels by PND 10. ERβ expression was not altered by BPA on PND 4, but significantly decreased or eliminated in both sexes by PND 10. Kiss1 expression was diminished by BPA in the anterior hypothalamus, especially in females. There were no significant impacts of BPA in the mediobasal hypothalamus. Collectively, BPA effects did not mirror those of EB. The results show that neonatal hypothalamic ER and Kiss1 expression is sensitive to BPA exposure. This disruption may alter sexually dimorphic hypothalamic organization and underlie adult reproductive deficiencies. Additionally, the discordant effects of EB and BPA indicate that BPA likely disrupts hypothalamic organization by a mechanism other than simply acting as an estrogen mimic., (Copyright © 2011 Elsevier Inc. All rights reserved.)

Release of gonadotropins in adult rodents is sex specific and dependent upon kisspeptin (Kiss1) neurons. This crucial pathway within the hypothalamic-pituitary-gonadal (HPG) axis is profoundly influenced by neonatal estrogens, which induce a male-like phenotype. Classically, estrogen activity is mediated via the estrogen receptors α and β (ERα and ERβ), but the relative roles each plays in organizing the sex-specific ontogeny of kisspeptin signaling pathways remain unresolved. Thus, the present study used in situ hybridization histochemistry (ISHH) to map the temporal and sexually dimorphic neonatal mRNA expression profiles of ERα, ERβ, and Kiss1 in the anterioventral periventricular nucleus (AVPV), medial preoptic area (MPOA), ventromedial nucleus (VMN), and arcuate nucleus (ARC), all regions critical for kisspeptin regulation of gonadotropin secretion. In general, females had higher levels of ERα, in all regions examined, a sex difference that persisted until postnatal day (PND) 19 except in the ARC. Males had significantly more ERβ expression in the AVPV at birth, but this sex difference was lost and then re-emerged on PND 19, with females having more than males. VMN ERβ levels were higher in females until PND 19. Kiss1 was not detectable until PND 11 in the anterior hypothalamus, but expression levels were equivalent at birth in the ARC. By PND 2, ARC ERα and Kiss1 levels were abundant, sexually dimorphic (higher in females), and, respectively, showed a U- and a bell-shaped pattern with age. Sex differences in ARC Kiss1 expression provide evidence that Kiss1 may play a role in the sexual dimorphic organization of the neonatal brain. These detailed profiles of neonatal Kiss1 and ERs mRNA levels will help elucidate the relative roles each plays in the sex-specific, estrogen-dependent organization of gonadotropin signaling pathways., (Copyright © 2011 Wiley-Liss, Inc.)

Neonatal exposure to estrogenic endocrine disrupting compounds (EDCs) can advance pubertal onset and induce premature anestrous in female rats. It was recently discovered that hypothalamic kisspeptin (KISS) signaling pathways are sexually dimorphic and regulate both the timing of pubertal onset and estrous cyclicity. Thus we hypothesized that disrupted sex specific ontogeny of KISS signaling pathways might be a mechanism underlying these EDC effects. We first established the sex specific development of KISS gene expression, cell number and neural fiber density across peripuberty in the anteroventral periventricular nucleus (AVPV) and arcuate nucleus (ARC), hypothesizing that the sexually dimorphic aspects of KISS signaling would be most vulnerable to EDCs. We next exposed female rats to the phytoestrogen genistein (GEN, 1 or 10 mg/kg bw), estradiol benzoate (EB, 10 μg), or vehicle from post natal day (P) 0-3 via subcutaneous (sc) injection. Animals were sacrificed on either P21, 24, 28, or 33 (n=5-14 per group at each age). Vaginal opening was significantly advanced by EB and the higher dose of GEN compared to control animals and was accompanied by lower numbers of KISS immunoreactive fibers in the AVPV and ARC. Ovarian morphology was also assessed in all age groups for the presence of multiple oocyte follicles (MOFs). The number of MOFs decreased over time in each group, and none were observed in control animals by P24. MOFs were still present, however, in the EB and 10 mg/kg GEN groups beyond P24 indicating a disruption in the timing of ovarian development., (Copyright © 2010 Elsevier Inc. All rights reserved.)

Developmental exposure to endocrine-disrupting compounds is hypothesized to adversely affect female reproductive physiology by interfering with the organization of the hypothalamic-pituitary-gonadal axis. Here, we compared the effects of neonatal exposure to two environmentally relevant doses of the plastics component bisphenol-A (BPA; 50 microg/kg and 50 mg/kg) with the ESR1 (formerly known as ERalpha)-selective agonist 4,4',4''-(4-propyl-[(1)H]pyrazole-1,3,5-triyl)trisphenol (PPT; 1 mg/kg) on the development of the female rat hypothalamus and ovary. An oil vehicle and estradiol benzoate (EB; 25 microg) were used as negative and positive controls. Exposure to EB, PPT, or the low dose of BPA advanced pubertal onset. A total of 67% of females exposed to the high BPA dose were acyclic by 15 wk after vaginal opening compared with 14% of those exposed to the low BPA dose, all of the EB- and PPT-treated females, and none of the control animals. Ovaries from the EB-treated females were undersized and showed no evidence of folliculogenesis, whereas ovaries from the PPT-treated females were characterized by large antral-like follicles, which did not appear to support ovulation. Severity of deficits within the BPA-treated groups increased with dose and included large antral-like follicles and lower numbers of corpora lutea. Sexual receptivity, examined after ovariectomy and hormone replacement, was normal in all groups except those neonatally exposed to EB. FOS induction in hypothalamic gonadotropic (GnRH) neurons after hormone priming was impaired in the EB- and PPT-treated groups but neither of the BPA-treated groups. Our data suggest that BPA disrupts ovarian development but not the ability of GnRH neurons to respond to steroid-positive feedback.

Neonatal exposure to endocrine disrupting compounds (EDCs) can impair reproductive physiology, but the specific mechanisms by which this occurs remain largely unknown. Growing evidence suggests that kisspeptin (KISS) neurons play a significant role in the regulation of pubertal onset and ovulation, therefore disruption of KISS signaling could be a mechanism by which EDCs impair reproductive maturation and function. We have previously demonstrated that neonatal exposure to phytoestrogens decreases KISS fiber density in the anterior hypothalamus of female rats, an effect which was associated with early persistent estrus and the impaired activation gonadotropin releasing hormone (GnRH) neurons. The goals of the present study were to (1) determine if an ERalpha selective agonist (PPT) or bisphenol-A (BPA) could produce similar effects on hypothalamic KISS content in female rats and (2) to determine if male KISS fiber density was also vulnerable to disruption by EDCs. We first examined the effects of neonatal exposure to PPT, a low (50 microg/kg bw) BPA dose, and a high (50 mg/kg bw) BPA dose on KISS immunoreactivity (-ir) in the anterior ventral periventricular (AVPV) and arcuate (ARC) nuclei of adult female rats, using estradiol benzoate (EB) and a sesame oil vehicle as controls. AVPV KISS-ir, following ovariectomy (OVX) and hormone priming, was significantly lower in the EB and PPT groups but not the BPA groups. ARC KISS-ir levels were significantly diminished in the EB and high dose BPA groups, and there was a nonsignificant trend for lower KISS-ir in the PPT group. We next examined effects of neonatal exposure to a low (50 microg/kg bw) dose of BPA and the phytoestrogens genistein (GEN) and equol (EQ) on KISS-ir in the AVPV and ARC of adult male rats, using OVX females as an additional control group. None of the compounds affected KISS-ir in the male hypothalamus. Our results suggest that the organization of hypothalamic KISS fibers may be vulnerable to disruption by EDC exposure and that females might be more sensitive than males.

It is well established that estrogen administration during neonatal development can advance pubertal onset and prevent the maintenance of regular estrous cycles in female rats. This treatment paradigm also eliminates the preovulatory rise of gonadotropin releasing hormone (GnRH). It remains unclear, however, through which of the two primary forms of the estrogen receptor (ERalpha or ERbeta) this effect is mediated. It is also unclear whether endocrine disrupting compounds (EDCs) can produce similar effects. Here we compared the effect of neonatal exposure to estradiol benzoate (EB), the ERalpha specific agonist 1,3,5-tris(4-hydroxyphenyl)-4-propyl-1H-pyrazole (PPT), the ERbeta specific agonist diarylpropionitrile (DPN) and the naturally occurring EDCs genistein (GEN) and equol (EQ) on pubertal onset, estrous cyclicity, GnRH activation, and kisspeptin content in the anteroventral periventricular (AVPV) and arcuate (ARC) nuclei. Vaginal opening was significantly advanced by EB and GEN. By 10 weeks post-puberty, irregular estrous cycles were observed in all groups except the control group. GnRH activation, as measured by the percentage of immunopositive GnRH neurons that were also immunopositive for Fos, was significantly lower in all treatment groups except the DPN group compared to the control group. GnRH activation was absent in the PPT group. These data suggest that neonatal exposure to EDCs can suppress GnRH activity in adulthood, and that ERalpha plays a pivotal role in this process. Kisspeptins (KISS) have recently been characterized to be potent stimulators of GnRH secretion. Therefore we quantified the density of KISS immunolabeled fibers in the AVPV and ARC. In the AVPV, KISS fiber density was significantly lower in the EB and GEN groups compared to the control group but only in the EB and PPT groups in the ARC. The data suggest that decreased stimulation of GnRH neurons by KISS could be a mechanism by which EDCs can impair female reproductive function.

Very little is known about the impact of selective estrogen receptor modulators (SERMs) on the brain. We examined the effects of tamoxifen (TAMOX) and the synthetic estrogen 17alpha-ethynylestradiol (EE) on estrogen-dependent gene expression and receptor binding in the female rat brain. Both immediate and residual effects were examined in both the presence and absence of 17beta-estradiol. Two groups of adult, ovariectomized, female rats (n=30 per group) were injected with TAMOX (5 mg/kg), EE (0.1 mg/kg), or sesame oil daily for 14 days. Animals from the first group were implanted with blank or 17beta-estradiol Silastic capsules concurrently with the last three SERM injections (immediate, group 1). Animals from the second group received either blank or 17beta-estradiol implants 2 weeks after the last injection (residual, group 2). All animals were sacrificed 72 h after implantation. TAMOX increased uterine weight in the absence of estrogen, but inhibited uterine weight gain in the presence of estrogen in both groups 1 and 2. TAMOX and EE increased oxytocin receptor binding in the ventromedial nucleus of the hypothalamus (VMN) in the absence of estrogen in both groups 1 and 2. The estrogen-dependent induction of PR mRNA expression in the VMN was significantly attenuated by TAMOX in group 1. Finally, TAMOX and EE had opposite effects on ERbeta mRNA expression in the paraventricular nucleus in the absence of 17beta-estradiol in group 1. Neither had any effect in group 2 when 17beta-estradiol was present. These results suggest that TAMOX has mixed agonist/antagonist effects in the female rat brain, many of which persist at least 2 weeks after the administration ceases.

Isoflavone phytoestrogens are growing increasingly popular because of their reported cardiovascular and anticarcinogenic properties, but the effects of these compounds in the brain are largely unknown. In a previous study, we found that an isoflavone supplement, containing a mixture of soy phytoestrogens, inhibited estrogen-dependent female sexual behavior and was antiestrogenic for both ER alpha- and ER beta-dependent gene expression in the hypothalamus. Here we examined the impact of the soy isoflavone genistein, a major component of the supplement, on estrogen-dependent female sexual behavior and ER alpha- and ER beta-dependent gene expression in the rat brain. Genistein, at a dietary concentration of 100 or 500 ppm had no effect on lordosis behavior in rats. However, at 500 ppm genistein had differential activity through ER alpha and ER beta in the hypothalamus. Genistein had no effect, in either the presence or absence of 17 beta-E2, on oxytocin receptor density in the ventromedial nucleus of the hypothalamus, an estrogen-dependent action thought to be regulated via ER alpha. However, genistein increased ER beta mRNA expression in the paraventricular nucleus of the hypothalamus by 24%, whereas 17 beta-E2 decreased ER beta mRNA expression by 26%, a process likely mediated by ER beta itself. These results suggest that at this dose, genistein has antiestrogenic action through ER beta in the paraventricular nucleus but negligible activity through ER alpha in the brain.

Prevalence of neurodevelopmental disorders (NDDs) with social deficits is conspicuously rising, particularly in boys. Flame retardants (FRs) have long been associated with increased risk, and prior work by us and others in multiple species has shown that developmental exposure to the common FR mixture Firemaster 550 (FM 550) sex-specifically alters socioemotional behaviors including anxiety and pair bond formation. In rats, FRs have also been shown to impair aspects of osmoregulation. Because vasopressin (AVP) plays a role in both socioemotional behavior and osmotic balance we hypothesized that AVP and its related nonapeptide oxytocin (OT) would be vulnerable to developmental FM 550 exposure. We used the prairie vole (Microtus ochrogaste) to test this because it is spontaneously prosocial. Using siblings of prairie voles used in a prior study that assessed behavioral deficits resulting from developmental FM 550 exposure across 3 doses, here we tested the hypothesis that FM 550 sex-specifically alters AVP and OT neuronal populations in critical nuclei, such as the paraventricular nucleus (PVN), that coordinate those behaviors, as well as related dopaminergic (determined by tyrosine hydroxylase (TH) immunolabeling) populations. Exposed females had fewer AVP neurons in the anterior PVN and more A13 TH neurons in the zona incerta than controls. By contrast, in FM 550 males, A13 TH neuron numbers in the zona incerta were decreased but only in 1 dose group. These results expand on previous work showing evidence of endocrine disruption of OT/AVP pathways, including to subpopulations of PVN AVP neurons that coordinate osmoregulatory functions in the periphery. [ABSTRACT FROM AUTHOR]

Abstract Background It is well established that sexual differentiation of the rodent hypothalamic-pituitary-gonadal (HPG) axis is principally orchestrated by estrogen during the perinatal period. Here we sought to better characterize the mechanistic role the beta form of the estrogen receptor (ERβ) plays in this process. Methods To achieve this, we exposed neonatal female rats to three doses (0.5, 1 and 2 mg/kg) of the ERβ selective agonist diarylpropionitrile (DPN) using estradiol benzoate (EB) as a positive control. Measures included day of vaginal opening, estrous cycle quality, GnRH and Fos co-localization following ovariectomy and hormone priming, circulating luteinizing hormone (LH) levels and quantification of hypothalamic kisspeptin immunoreactivity. A second set of females was then neonatally exposed to DPN, the ERα agonist propyl-pyrazole-triol (PPT), DPN+PPT, or EB to compare the impact of ERα and ERβ selective agonism on kisspeptin gene expression in pre- and post-pubescent females. Results All three DPN doses significantly advanced the day of vaginal opening and induced premature anestrus. GnRH and Fos co-labeling, a marker of GnRH activation, following ovariectomy and hormone priming was reduced by approximately half at all doses; the magnitude of which was not as large as with EB or what we have previously observed with the ERα agonist PPT. LH levels were also correspondingly lower, compared to control females. No impact of DPN was observed on the density of kisspeptin immunoreactive (-ir) fibers or cell bodies in the arcuate (ARC) nucleus, and kisspeptin-ir was only significantly reduced by the middle (1 mg/kg) DPN dose in the preoptic region. The second experiment revealed that EB, PPT and the combination of DPN+PPT significantly abrogated preoptic Kiss1 expression at both ages but ARC expression was only reduced by EB. Conclusion Our results indicate that selective agonism of ERβ is not sufficient to completely achieve male-typical HPG organization observed with EB or an ERα agonist.

A wide range of health benefits have been ascribed to soya intake including a lowered risk of osteoporosis, heart disease, breast cancer, and menopausal symptoms. Because it is a hormonally active diet, however, soya can also be endocrine disrupting, suggesting that intake has the potential to cause adverse health effects in certain circumstances, particularly when exposure occurs during development. Consequently, the question of whether or not soya phyto-oestrogens are beneficial or harmful to human health is neither straightforward nor universally applicable to all groups. Possible benefits and risks depend on age, health status, and even the presence or absence of specific gut microflora. As global consumption increases, greater awareness and consideration of the endocrine-disrupting properties of soya by nutrition specialists and other health practitioners is needed. Consumption by infants and small children is of particular concern because their hormone-sensitive organs, including the brain and reproductive system, are still undergoing sexual differentiation and maturation. Thus, their susceptibility to the endocrine-disrupting activities of soya phyto-oestrogens may be especially high. As oestrogen receptor partial agonists with molecular and cellular properties similar to anthropogenic endocrine disruptors such as bisphenol A, the soya phyto-oestrogens provide an interesting model for how attitudes about what is ‘synthetic’ v. what is ‘natural,’ shapes understanding and perception of what it means for a compound to be endocrine disrupting and/or potentially harmful. This review describes the endocrine-disrupting properties of soya phyto-oestrogens with a focus on neuroendocrine development and behaviour. [ABSTRACT FROM PUBLISHER]

The rodent amygdaloid complex is composed of numerous subnuclei important for the sex-specific regulation of sociosexual behavior. Although estrogen receptors (ERs) are critical for organizing functional and cytoarchitectural sex differences in these subnuclei, a detailed developmental profile of ER expression in the amygdaloid complex is not available. Moreover, the kisspeptin gene (Kiss1) was recently identified in the adult amygdala, but it remains unknown if it is expressed during development. To fill these data gaps, rat brains (5-7/group) were assessed on postnatal days (PNDs) 0, 2, 4, 7, and 19 for ER alpha (ERα; Esr1), beta (ERβ; Esr2), and Kiss1 expression using in situ hybridization. Expression was quantified in the posterodorsal portion of the medial amygdala posterodorsal (MePD), lateral (PLCo), and medial (PMCo) components of the posterior cortical nucleus, and the amygdalohippocampal area (AHi). ERα expression was high throughout the amygdala at birth, but sexually dimorphic only in the AHi. ERα expression in the MePD and the PLCo showed a U-shaped expression pattern over time. In the PMCo, ERα expression decreased from PND 2 and remained low through PND 19. Sexually dimorphic expression of ERβ in the MePD was observed on PND 0, with higher levels in females, but reversed by PND 4 due to declining levels in females. No Kiss1 signal was observed in the postnatal amygdala, suggesting that expression arises after puberty. These data reveal that ER expression is region-specific within the neonatal amygdala. These differences likely contribute to sex differences in sociosexual behavior across the lifespan. J. Comp. Neurol. 521:465-478, 2013. © 2012 Wiley Periodicals, Inc. [ABSTRACT FROM AUTHOR]

Abstract: Phytoestrogens are plant derived compounds found in a wide variety of foods, most notably soy. A litany of health benefits including a lowered risk of osteoporosis, heart disease, breast cancer, and menopausal symptoms, are frequently attributed to phytoestrogens but many are also considered endocrine disruptors, indicating that they have the potential to cause adverse health effects as well. Consequently, the question of whether or not phytoestrogens are beneficial or harmful to human health remains unresolved. The answer is likely complex and may depend on age, health status, and even the presence or absence of specific gut microflora. Clarity on this issue is needed because global consumption is rapidly increasing. Phytoestrogens are present in numerous dietary supplements and widely marketed as a natural alternative to estrogen replacement therapy. Soy infant formula now constitutes up to a third of the US market, and soy protein is now added to many processed foods. As weak estrogen agonists/antagonists with molecular and cellular properties similar to synthetic endocrine disruptors such as Bisphenol A (BPA), the phytoestrogens provide a useful model to comprehensively investigate the biological impact of endocrine disruptors in general. This review weighs the evidence for and against the purported health benefits and adverse effects of phytoestrogens. [ABSTRACT FROM AUTHOR]

Abstract: Serotonin (5-HT) is known to play a role in the suppression of the lordosis response in males. We have previously shown that there is a sex difference in the density of 5-HT immunoreactive (5-HT-ir) fibers in the ventrolateral division of the adult ventromedial nucleus of the hypothalamus (VMNvl) and that neonatal administration of estradiol (E2) increases 5-HT-ir in the female VMNvl to male-typical levels. Here we demonstrate that postnatal administration of the ERα agonist 1,3,5-tris(4-Hydroxyphenyl)-4-propyl-1H-pyrazole (PPT), but not the ERβ agonist diarylpropionitrile (DPN), also masculinizes 5-HT-ir in the female VMNvl, suggesting a mechanistic role for ERα in this process. Sexual receptivity, as ascertained by the lordosis quotient, was unaffected by either PPT or DPN treatment but nearly abolished by estradiol benzoate (EB), a synthetic estrogen with high affinity for both ERα and ERβ. Collectively, these observations show that postnatal estrogens increase the density of 5-HT projections to the VMNvl via an ERα dependent mechanism, but that this increased inhibitory input is not sufficient to suppress the lordosis response. [Copyright &y& Elsevier]

Abstract: Changes in the volumes of sexually dimorphic brain nuclei are often used as a biomarker for developmental disruption by endocrine-active compounds (EACs). However, these gross, morphological analyses do not reliably predict disruption of cell phenotype or neuronal function. Therefore, an experimental approach that simultaneously assesses anatomical, physiological and behavioral endpoints is required when developing risk assessment models for EAC exposure. Using this more comprehensive approach we have demonstrated that the disruption of nuclear volume does not necessarily coincide with disruption of cellular phenotype or neuroendocrine function in two sexually dimorphic brain nuclei: the anteroventral periventricular nucleus of the hypothalamus (AVPV) and the sexually dimorphic nucleus of the preoptic area (SDN). These results demonstrate that nuclear volume is likely not an appropriate biomarker for EAC exposure. We further demonstrated that neonatal exposure to the EACs genistein (GEN) and Bisphenol-A (BPA) can affect sexually dimorphic brain morphology and neuronal phenotypes in adulthood with regional and cellular specificity suggesting that effects observed in one brain region may not be predictive of effects within neighboring regions. Finally, developmental EAC exposure has been shown to affect a variety of sexually dimorphic behaviors including reproductive behavior. These effects are likely to have a broad impact as maladaptive behavior could translate to decreased fitness of entire populations. Collectively, these findings emphasize the need to employ a comprehensive approach that addresses anatomical, functional and behavioral endpoints when evaluating the potential effects of EAC exposure. [Copyright &y& Elsevier]

Abstract: There is growing concern that naturally occurring and chemically manufactured endocrine-active compounds (EACs) may disrupt hormone-dependent events during central nervous system development. We examined whether postnatal exposure to the phytoestrogen genistein (GEN) or the plastics component bisphenol-A (BIS) affected sexual differentiation of the anteroventral periventricular nucleus of the hypothalamus (AVPV) in rats. The AVPV is sexually differentiated in rodents. The female AVPV is larger than the male AVPV and contains a higher number of cells expressing tyrosine hydroxylase (TH). Sexual differentiation of the AVPV results from exposure of the male nervous system to estrogen aromatized from testicular testosterone secreted in the first few days after birth. Thus, we hypothesized that exposure to EACs during this critical period could alter the sexually dimorphic expression of TH and the overall expression of estrogen receptor alpha (ERα) in the AVPV. Animals were given 4 subcutaneous injections of sesame oil (control), 50 μg 17β-estradiol (E2), 250 μg GEN, or 250 μg BIS at 12-h intervals over postnatal days (PND) 1 and 2 and sacrificed on PND 19. E2 treatment masculinized TH immunoreactivity (TH-ir) in the female AVPV while exposure to GEN or BIS demasculinized TH-ir in the male AVPV. In addition, we identified a population of neurons co-expressing TH and ERα located primarily in the medial region of the AVPV. Normally, females have nearly three times as many double-labeled cells as males, but their numbers were defeminized by E2, GEN or BIS treatment. These results suggest that acute exposure to EACs during a critical developmental period alters AVPV development. [Copyright &y& Elsevier]

There is perhaps no endocrine disrupting chemical more controversial than bisphenol A (BPA). Comprising a high‐volume production chemical used in a variety of applications, BPA has been linked to a litany of adverse health‐related outcomes, including effects on brain sexual differentiation and behaviour. Risk assessors preferentially rely on classical guideline‐compliant toxicity studies over studies published by academic scientists, and have generally downplayed concerns about the potential risks that BPA poses to human health. It has been argued, however, that, because traditional toxicity studies rarely contain neural endpoints, and only a paucity of endocrine‐sensitive endpoints, they are incapable of fully evaluating harm. To address current controversies on the safety of BPA, the United States National Institute of Environmental Health Sciences, the National Toxicology Program (NTP), and the US Food and Drug Administration established the Consortium Linking Academic and Regulatory Insights on BPA Toxicity (CLARITY‐BPA). CLARITY‐BPA performed a classical regulatory‐style toxicology study (Core study) in conjunction with multiple behavioural, molecular and cellular studies conducted by academic laboratories (grantee studies) using a collaboratively devised experimental framework and the same animals and tissues. This review summarises the results from the grantee studies that focused on brain and behaviour. Evidence of altered neuroendocrine development, including age‐ and sex‐specific expression of oestrogen receptor (ER)α and ERβ, and the abrogation of brain and behavioural sexual dimorphisms, supports the conclusion that developmental BPA exposure, even at doses below what regulatory agencies regard as "safe" for humans, contribute to brain and behavioural change. The consistency and the reproducibility of the effects across CLARITY‐BPA and prior studies using the same animal strain and almost identical experimental conditions are compelling. Combined analysis of all of the data from the CLARITY‐BPA project is underway at the NTP and a final report expected in late 2019. [ABSTRACT FROM AUTHOR]

• Bisphenol A (BPA) is a well characterized endocrine disruptor that can affect brain sexual differentiation and behaviour. • Oxytocin receptor expression is sexually dimorphic and sensitive to steroid hormones. • We hypothesized that developmental BPA exposure would alter oxytocin receptor expression in the juvenile rat brain. • Effects, including loss of sexually dimorphic expression, were found in four brain regions. • These data support prior work showing that BPA can disrupt brain sexual differentiation. Bisphenol A (BPA) is a well-characterized endocrine disrupting chemical (EDC) used in plastics, epoxy resins and other products. Neurodevelopmental effects of BPA exposure are a major concern with multiple rodent and human studies showing that early life BPA exposure may impact the developing brain and sexually dimorphic behaviors. The CLARITY-BPA (Consortium Linking Academic and Regulatory Insights on BPA Toxicity) program was established to assess multiple endpoints, including neural, across a wide dose range. Studies from our lab as part of (and prior to) CLARITY-BPA have shown that BPA disrupts estrogen receptor expression in the developing brain, and some evidence of oxytocin (OT) and oxytocin receptor (OTR) disruption in the hypothalamus and amygdala. While BPA disruption of steroid hormone function is well documented, less is known about its capacity to alter nonapeptide signals. In this CLARITY-BPA follow up study, we used remaining juvenile rat tissues to test the hypothesis that developmental BPA exposure affects OTR expression across the brain. Perinatal BPA exposure (2.5, 25, or 2500 μg/kg body weight (bw)/day) spanned gestation and lactation with dams gavaged from gestational day 6 until birth and then the offspring gavaged directly through weaning. Ethinyl estradiol (0.5 μg/kg bw/day) was used as a reference estrogen. Animals of both sexes were sacrificed as juveniles and OTR expression assessed by receptor binding. Our results demonstrate prenatal exposure to BPA can eliminate sex differences in OTR expression in three hypothalamic regions, and that male OTR expression may be more susceptible. Our data also identify a sub-region of the BNST with sexually dimorphic OTR expression not previously reported in juvenile rats that is also susceptible to BPA. [ABSTRACT FROM AUTHOR]

Bisphenol A (BPA) is a high volume endocrine disrupting chemical found in a wide variety of products including plastics and epoxy resins. Human exposure is nearly ubiquitous, and higher in children than adults. Because BPA has been reported to interfere with sex steroid hormone signaling, there is concern that developmental exposure, even at levels below the current FDA No Observed Adverse Effect Level (NOAEL) of 5 mg/kg body weight (bw)/day, can disrupt brain sexual differentiation. The current studies were conducted as part of the CLARITY-BPA (Consortium Linking Academic and Regulatory Insights on BPA Toxicity) program and tested the hypothesis that perinatal BPA exposure would induce morphological changes in hormone sensitive, sexually dimorphic brain regions. Sprague-Dawley rats were randomly assigned to 5 groups: BPA (2.5, 25, or 2500 μg/kg bw/day), a reference estrogen (0.5 μg ethinylestradiol (EE 2 )/kg bw/day), or vehicle. Exposure occurred by gavage to the dam from gestational day 6 until parturition, and then to the offspring from birth through weaning. Unbiased stereology was used to quantify the volume of the sexually dimorphic nucleus (SDN), the anteroventral periventricular nucleus (AVPV), the posterodorsal portion of the medial amygdala (MePD), and the locus coeruleus (LC) at postnatal day 28. No appreciable effects of BPA were observed on the volume of the SDN or LC. However, AVPV volume was enlarged in both sexes, even at levels below the FDA NOAEL. Collectively, these data suggest the developing brain is vulnerable to endocrine disruption by BPA at exposure levels below previous estimates by regulatory agencies. [ABSTRACT FROM AUTHOR]

During development, microglia, the resident immune cells of the brain, play an important role in synaptic organization. Microglial colonization of the developing brain is sexually dimorphic in some regions, including nuclei critical for the coordination of social behavior, suggesting steroid hormones have an influencing role, particularly estrogen. By extension, microglial colonization may be vulnerable to endocrine disruption. Concerns have been raised regarding the potential for endocrine disrupting compounds (EDCs) to alter brain development and behavior. Developmental exposure to Bisphenol A (BPA), a ubiquitous EDC, has been associated with altered sociosexual and mood-related behaviors in various animal models and children. Through a comparison of the promiscuous Wistar rat ( Rattus norvegicus ) and the socially monogamous prairie vole ( Microtus ochrogaster ), we are the first to observe that developmental exposure to the synthetic estrogen ethinyl estradiol (EE) or BPA alters the sex-specific colonization of the hippocampus and amygdala by microglia. [ABSTRACT FROM AUTHOR]