Tetsuji Nagao

Reproductive effects in male and female rats of neonatal exposure to genistein.

Sprague-Dawley rats were administered genistein orally at doses of 12.5, 25, 50, or 100 mg/kg on postnatal days 1 through 5 to examine its effects on reproductive function after puberty. In addition, preputial separation and vaginal opening as endpoints of sexual maturation, estrous cycling, sperm count, serum testosterone concentration, and histopathologic changes of reproductive organs of male and female rats were examined. Body weights of male and female rats exposed to genistein at any dose level examined were lower than those of controls. Timing of preputial separation in males and timing of vaginal opening were not affected by genistein treatment. The number of females showing estrous cycle irregularities was increased by genistein treatment. The fertility of female rats exposed neonatally to genistein at 100 mg/kg was disrupted, while neonatal exposure to genistein did not affect male fertility. Neither sperm counts nor serum testosterone concentration were changed by neonatal exposure to genistein. Female rats exposed neonatally to genistein at 100 mg/kg showed histopathologic changes in the ovaries and uterus, while male rats showed no histopathologic alterations in the gonads. The results of this study indicate that early neonatal exposure to genistein caused dysfunction of postpubertal reproductive performance as well as abnormal development of gonads in female but not in male rats.

Reproductive effects of nonylphenol in rats after gavage administration: a two-generation study.

The potential reproductive toxicity of nonylphenol (NP) was assessed in a two-generation reproductive toxicity study. Groups of 25 male and female Crj:CD (SD) IGS rats were given NP by gavage at levels of 2, 10, or 50 mg/kg, and 25 males and females were given corn oil as controls. No adverse changes in clinical signs were observed in any rats throughout the study. Significant increases in the liver, kidney and pituitary gland weights in males, and decreases in thymus weight in males and in ovary weight in females were observed in the 50 mg/kg group. NP did not affect sperm characteristics or the estrous cycle at any dose administered. A significant increase in the TSH level was observed in males in the 50 mg/kg group. No adverse effects of NP on reproduction were found. At necropsy, no treatment-related alterations were observed in any organs including the reproductive tissues in any group. Histopathologic changes were found in the liver of male and female rats and kidneys of males in the 50 mg/kg group. The viability of offspring from postnatal day 0 to 4 in the 50 mg/kg group was reduced as compared with that in the controls, although growth was not affected by NP administration. On postnatal day 22, an increase in the serum FSH level and decrease in T(3) level for males, and decreases in LH and TSH levels and an increase in T(3) levels for females were observed in the 50 mg/kg group. NP did not affect the timing of preputial separation, while vaginal opening was accelerated in the 50 mg/kg group. No adverse changes were found in behavior or learning in the offspring of NP-treated groups. There were no treatment-related changes in any reproductive parameter, including estrous cycle, mating, fertility, delivery, and lactation, except for significant decreases in the numbers of implantation sites and live pups, and a significant decrease in ovary weight in the 50 mg/kg group. Kidney and liver weight were increased in males in the 50 mg/kg group. Histopathologic examination revealed changes in the liver of males and females of the 50 mg/kg group. No treatment-related changes were observed in the sperm characteristics. Hormone data should be interpreted cautiously until the findings are repeated and confirmed by further studies. These results of NP suggested that the no observed adverse effect level (NOAEL) on reproductive capacity is 50 mg/kg/day or greater in parent animals, and 10 mg/kg/day in the next generation under the present experimental condition.

Reproductive function in rats exposed neonatally to bisphenol A and estradiol benzoate

The reproductive function in rats treated subcutaneously (s.c.) with 300 microg/g bisphenol A or 2 microg/g estradiol benzoate from postnatal Day 1 to 5 was examined after puberty as well as histolopathogic changes in reproductive organs. All male and female rats treated postnatally with estradiol benzoate showed poor reproductive capability, including adverse effects on masculine sexual behavior, and marked histopathologic alterations of the reproductive organs. In addition, estradiol benzoate markedly reduced the volume of the sexually dimorphic nucleus of the preoptic area (SDN-POA) in males. On the other hand, all male and female rats treated postnatally with bisphenol A showed normal reproductive function and no histopathologic abnormalities of reproductive organs. Bisphenol A did not affect the volume of the SDN-POA. These results indicated that neonatal exposure to estradiol benzoate affects reproductive function in male and female rats, and treatment with bisphenol A at a fairly high dose was ineffective if given postnatally to male and female rats.

Effect of butyl benzyl phthalate in Sprague-Dawley rats after gavage administration: a two-generation reproductive study

Butyl benzyl phthalate (BBP), a plasticizer, has been shown in in vitro studies to be weakly estrogenic, and in in vivo studies to possess testicular toxicity and teratogenicity, but few experimental data on BBP multigeneration effects on reproduction in mammals are available. The present two-generation reproductive study was conducted in male and female Sprague-Dawley rats using oral doses of 0, 20, 100, and 500 mg/kg/day BBP. Endpoints were chosen in order to evaluate both subchronic and reproductive toxicity. In the parent animals (F(0)), a decrease in body weight gain was observed in males in the 500 mg/kg/day group, although no significant decrease in food consumption was found. No dose-related changes were observed in estrous cyclicity, fertility, or lactation. A dose-dependent increase in kidney weight in rats of both sexes, an increase in liver weight in males, and a decrease in the weight of the ovaries in females were observed. No macroscopic or microscopic changes were found in the reproductive system of males or females. Oral administration of BBP caused a decrease in the serum concentration of testosterone, and an increase in FSH. In the next generation (F(1)), the body weight of male and female offspring at birth in the 100 and 500 mg/kg groups was significantly decreased, and the body weight in the 500 mg/kg group was lower throughout the study, while viability was not affected. Anogenital distance (AGD) at birth was decreased in male pups and was increased in female pups of the 500 mg/kg/day group. Preputial separation for male offspring in the 500 mg/kg/day group was delayed, while vaginal opening for female offspring in this group was not affected. BBP did not affect reproductive ability, including delivery and lactation, at any dose whereas macroscopic and microscopic changes of the testis, and decreased serum concentrations of testosterone were observed in male offspring of the 500 mg/kg/day group after puberty. From these data, it would appear that 20 mg/kg BBP is a no observed adverse effect level (NOAEL) for reproductive effects on parent animals and the next generation.

Maternal bisphenol A oral dosing relates to the acceleration of neurogenesis in the developing neocortex of mouse fetuses.

Bisphenol A (BPA), an endocrine-disruptor, is widely used in the production of plastics and resins. Human perinatal exposure to this chemical has been proposed to be a potential risk to public health. Animal studies indicate that postnatal exposure to BPA may affect neocortex development in embryos by accelerated neurogenesis and causing neuronal migration defects. The detailed phenotypes and pathogenetic mechanisms, especially with regard to the proliferation and differentiation of neural stem/progenitor cells, however, have not been clarified. C57BL/6J pregnant mice were orally administered BPA at 200μg/kg from embryonic day (E) 8.5 to 13.5, and the fetuses were observed histologically at E14.5. To clarify the histological changes, especially in terms of neurogenesis, proliferation and cell cycle, we performed histological analysis using specific markers of neurons/neural stem cells and cell cycle-specific labeling experiments using thymidine-analog substances. Cortical plate was hyperplastic and the number of neural stem/progenitor cells was decreased after the exposure to BPA. In particular, the maternal BPA oral dosing related to the effects on intermediate progenitor cells (IPCs, neural progenitor cells) in the subventricular zone (SVZ) of dorsal telencephalon. Exposure to BPA associated the promotion of the cell cycle exit in radial glial cells (RGCs, neural stem cells) and IPCs, and decreased the proliferation resulting from the prolong cell cycle length of IPCs in the SVZ. Our data show that maternal oral exposure to BPA related to the disruption of the cell cycle in IPCs and the effects of neurogenesis in the developing neocortex.

Low-dose bisphenol A does not affect reproductive organs in estrogen-sensitive C57BL/6N mice exposed at the sexually mature, juvenile, or embryonic stage

Bisphenol A (BPA) is used on a large scale in the manufacture of polycarbonate plastics. BPA has been shown to bind weakly to both estrogen receptor (ER) alpha and ER beta. The objective of this study was to evaluate the effects of low-dose BPA on male sexual development after exposure at various stages of development. Mice of the estrogen-sensitive strain C57BL/6N were exposed to BPA orally at doses of 2, 20, or 200 microg/kg at various stages, i.e. adulthood, the immature stage just after weaning, or the embryonic/fetal stage, to evaluate the effects of low-dose BPA on male reproductive organs. Body weight changes, weights of reproductive organs (testes, epididymides, seminal vesicles), cauda epididymal sperm density, and histology of reproductive organs including the ventral prostate were not affected by exposure to BPA at any dose examined. The results of this study indicate that exposure of estrogen-sensitive C57BL/6N mice to low-dose BPA did not reduce sperm density or disrupt development of the male reproductive organs.

Reproductive effects in male and female rats from neonatal exposure to p-octylphenol.

A number of alkylphenolic compounds are used in a variety of commercial products and have been shown in in vitro studies to be weakly estrogenic, but in vivo data are not available addressing this issue in mammals. Human exposure to alkylphenols may occur not only from these environmental contaminants but also through contact with manufactured and metabolic breakdown products. In this study, Sprague-Dawley rats were exposed to octylphenol by oral gavage at doses of 0 (vehicle: corn oil), 12.5, 25, 50, or 100 mg/kg once daily on postnatal days 1 through 5 to examine its effects on male and female reproductive function after puberty. In addition, preputial separation and vaginal opening as endpoints of sexual maturation, estrous cycling, sperm count, serum testosterone concentration, and histopathologic changes of the reproductive organs of male and female rats were examined. Male reproductive organs were weighed at necropsy. Body weights of male and female rats exposed to octylphenol at 50 and 100 mg/kg throughout the study after the administration period, those of both sexes at 7 and 9 weeks of age in the 25 mg/kg group, and that of females at 9 weeks of age in the 12.5 mg/kg group were lower than those of controls. Significant delays in acquisition of puberty in males and females exposed to octylphenol at 50 and 100 mg/kg were observed. Estrous cycle, copulation and fertility, sperm count, and serum testosterone concentration were not affected by neonatal exposure to octylphenol. Significant decrease in absolute and relative prostate weight in the 12.5, 25, 50, and 100 mg/kg groups, and absolute epididymal weight in the 100 mg/kg group, increase in relative testes weight in the 100 mg/kg group, and relative seminal vesicle weights in the 50 and 100 mg/kg groups were found. Histopathologic analyses of reproductive organs in male and female rats exposed neonatally to octylphenol revealed no marked alterations. The results of this study indicate that early neonatal exposure to octylphenol by oral gavage did not cause dysfunction of reproductive performance (mating and fertility) in male or female rats, and no disruption of development of the reproductive tract was observed in male or female rats, while significant decreases in body weights in the 25 mg/kg and more groups, delays of sexual maturation in the 50 mg/kg and greater groups, and decrease in ventral prostate weights in all octylphenol-treated groups were found. Therefore, it is concluded that NOAEL (no-observed adverse effect level) for systemic toxicity was < or =12.5 mg/kg/day and that for reproductive toxicity was 100 mg/kg/day under the present experimental condition.

Behavioral Characteristics of Rats Selectively Bred for High and Low Avoidance Shuttlebox Response

Abstract Selective breeding of rat strains showing high and low avoidance behavior in the shuttlebox test has been performed since 1985. To elucidate the postnatal behavior of the high and low avoidance strains, animals from both lines were subjected to an open‐field test, wheel cage activity test and Biel maze test in the 13th, 14th and 16th generations, respectively. No significant differences in any parameter in the open‐field test were observed between high and low avoidance lines. However, animals from the high avoidance line showed higher motor activity in the wheel cage and fewer errors in the Biel maze than the low avoidance line.

Newborn mice exposed prenatally to bisphenol A show hyperactivity and defective neocortical development

The central nervous system is especially susceptible to toxic insults during development. Prenatal administration of bisphenol A (BPA) induces histologic anomalies in the dorsal telencephalon of the embryo. Whether these anomalies affect the morphogenesis and maturation of neuronal function of the newborn neocortex, however, is unknown. To evaluate the neurodevelopmental and behavioral effects of prenatal BPA exposure at 20 and 200μg/kg/day in newborn mice, we performed a detailed histologic analysis of the neocortex and tested for the presence of behavioral abnormalities in newborn mice prenatally exposed to BPA using our newly developed behavioral test. Observations of newborn mice prenatally exposed to BPA revealed abnormal neuronal distribution and layer formation, hypoplasia of layer 6b, and abnormal dopaminergic neuronal projections in the neocortex. Further, the newborn mice exhibited hyperactivity. These findings suggest that prenatal BPA exposure induces neurobehavioral toxicity associated with abnormal dopaminergic neuronal projections, and abnormal corticogenesis and lamination. Histologic and behavioral analyses of newborn mice are considered useful for assessing the neurodevelopmental and behavioral toxicity of chemicals.

Comparative study of behavioral development between high and low shuttlebox avoidance rats

Previously, we reported that high- and low-avoidance animals (HAA and LAA, respectively), selectively bred for different avoidance response rates in a shuttlebox avoidance test, showed additional behavioral differences in wheel cage activity and in water maze performance after weaning. In the present study, physical and behavioral development were examined in HAA and LAA during the preweaning period. As compared to HAA, LAA offspring showed lower body weight, delayed eye opening, poorer performance in pivoting and negative geotaxis, and increased open-field activity. A fostering study indicated that these differences observed in eye opening, pivoting, negative geotaxis, open-field activity, swimming speed, shuttlebox avoidance and wheel cage activity were independent of maternal care. Only the pup weight was strongly dependent on the maternal line. These results indicate that behavioral differences between HAA and LAA observed in the pre- and postweaning periods may be linked to the avoidance genotype, but the difference in pup weight may be caused by maternal care.