William J. Swartz

Effects of methoxychlor on the reproductive system of the adult female mouse 1. Gross and histologic observations

The purpose of this study is to examine the effects of the pesticide methoxychlor (MXC) on the reproductive system of the adult female mouse. Sexually mature (7- to 8-week) virgin female CD-1 mice were exposed to 1.25, 2.5, or 5.0 mg MXC (50% technical grade) via oral gavage for 5 consecutive days each week for either 2 or 4 weeks. Control groups received either 0.025 mg estradiol-17 beta (E-17 beta) or the sesame oil vehicle for the same time period. Vaginal smears were taken daily, and weights were recorded weekly. Twenty-four hours following the final exposure, animals were sacrificed. Ovaries and reproductive tracts were removed and weighed. One ovary from each animal was prepared for light microscopic evaluation. Results revealed a dose dependency of MXC in inducing persistent vaginal estrus (PVE). Ovaries of MXC-exposed and E-17 beta-exposed animals weighed significantly less than the sesame oil controls. In addition, there was an increase in the number of atretic large follicles in the E-17 beta group and in those mice treated with the two highest doses of MXC, indicating a potential reduction in the immediate fertility of the animal. Thus, this commonly employed pesticide appears to mimic closely those effects on the female reproductive system induced by estrogens.

Effects of methoxychlor treatment of pregnant mice on female offspring of the treated and subsequent pregnancies

This study was designed to assess whether exposure to the estrogenic pesticide methoxychlor (MXC) during pregnancy would affect reproductive parameters not only in female offspring exposed prenatally, but also in those of a subsequent litter. Mice were exposed via oral gavage to 7.5, 5.0, or 2.5 mg technical grade MXC (50%) or 0.025 mg estradiol-17 beta (E-17 beta) from days 6 to 15 of pregnancy. Following delivery, female offspring (F1a) were cross-fostered and sacrificed at 8 weeks of age. Mothers exposed during their first pregnancy were allowed to mate again and their second set of offspring (F1b) were similarly evaluated to detect any latent effects from the initial exposure. Mice exposed to 7.5 mg MXC were unable to carry their litters to term. Results revealed a significant increase in the length of gestation of mice exposed to both E-17 beta and 5.0 mg MXC. A larger percentage of atretic follicles appeared in the ovaries of F1a females exposed prenatally to 5.0 mg MXC when compared to controls. Females from the F1b litter displayed a significant advance in time of vaginal opening, an apparent residual effect of MXC from a mother exposed during a previous pregnancy.

Chlordecone-induced follicular toxicity in mouse ovaries

The effect of the pesticide, chlordecone, on murine follicular development was examined. Female CD-1 mice were exposed to chlordecone for 5 consecutive days for each of 4 consecutive weeks (0.25 mg/day). Controls received sesame oil vehicle or estradiol-17 beta (E-17 beta; 0.1 mg/day) since chlordecone has been ascribed estrogenic activity. Animals were sacrificed 24 h following the final exposure. Ovaries were removed, serially sectioned, and stained. Follicles were classified as small, medium, or large and were tabulated. Twice as many medium-sized follicles were found in the E-17 beta-treated mice as in both the chlordecone-exposed and sesame oil control groups. Both pesticide- and E-17 beta-exposed mice displayed a much higher percent of atresia in the large follicles; however, there were more actual healthy, large follicles in the E-17 beta group. Thus, both chlordecone and E-17 beta induced increased atresia among large follicles, which could be due to the estrogenicity of these agents. However, a decreased pool of healthy large- and medium-sized follicles occurred in chlordecone-treated mice, a condition not seen in E-17 beta-treated mice. Thus, the pool of potentially ovulatory follicles is reduced in the pesticide-treated animals.

Effects of methoxychlor on the reproductive system of the adult female mouse: 2. Ultrastructural observations☆☆☆

The purpose of this study is to examine the effects of the pesticide methoxychlor (MXC) on the ultrastructural appearance of the different cellular components of the mouse ovary. Sexually mature (7- to 8-week) virgin female CD-1 mice were exposed to 5.0 mg MXC (50% technical grade) via oral gavage for 5 consecutive days each week for 4 weeks. Control groups received either 0.025 mg estradiol-17 beta (E-17 beta) or the sesame oil vehicle for the same time period. Twenty-four hours following the final exposure, animals were sacrificed. Ultrastructural observations revealed increased lipid accumulation in interstitial cells and theca cells of both estradiol-treated and 5.0 mg MXC-treated mice. This would suggest that these cells are unable to synthesize and secrete steroids. Thus, this commonly employed pesticide appears to closely mimic those effects on the female ovary induced by estrogen.

Neonatal exposure to technical methoxychlor alters pregnancy outcome in female mice.

This study was designed to determine the ability of female mice who were exposed neonatally to the pesticide methoxychlor (MXC) to mate, ovulate, and become pregnant upon reaching sexual maturity. One-day-old female mice (5 to 8/group) were exposed daily by intraperitoneal (ip) injection for 14 d to either sesame oil or 10 microg estradiol-17beta or 0.1, 0.5 or 1.0 mg MXC suspended in sesame oil. The MXC exposures corresponded to 14 to 71, 68 to 357, or 135 to 714 mg/kg body weight, respectively. Three months later, female mice were placed with proven breeder males and checked daily for vaginal plugs. Mated female mice were sacrificed 18 d after the appearance of a vaginal plug to evaluate pregnancy. Uteri were examined for the presence of living fetuses and/or resorption sites. Ovaries were removed and prepared for histologic evaluation and tabulation of corpora lutea. All mice from all three MXC-treated groups did in fact mate, in comparison with only one of those exposed neonatally to estradiol. Increasing the dose of MXC produced a decreased number of pregnant animals at 18 d following mating. The mean number of live fetuses/litter was reduced in the 0.5 and 1.0 mg MXC-treated groups. Corpora lutea were significantly reduced in ovaries from only the 1.0 mg MXC group and the estradiol group. No effects of treatment were seen at 0.1 mg MXC. It is concluded that neonatal exposure to MXC does not interfere with mating. Instead, significant alterations are seen in initiating and/or maintaining pregnancy. The deleterious effects on pregnancy may be due to the influence of neonatal MXC treatments on the hypothalamic-pituitary-ovarian axis as well as on possible alteration of the uterine environment.

Decreased superovulation in adult mice following neonatal exposures to technical methoxychlor

To examine the effects of technical methoxychlor (MXC) on superovulation, neonatal mice received intraperitoneal (i.p.) injections of either sesame oil, 10 micrograms of estradiol 17 beta, or 0.1, 0.5, or 1 mg of technical MXC. At 2 and 4 months, half of the mice received a superovulatory regimen of 10 IU pregnant mares serum gonadotropin followed by 10 IU human chorionic gonadotropin. The mice were sacrificed 15 to 20 h later, the number of ovulated oocytes were counted, and the ovaries were removed for histology. In the lowest MXC dose, the ovaries appeared normal and at 2 months, ovulated the same number of oocytes as controls. Estradiol or the highest two MXC doses induced ovarian atrophy. Following gonadotropin injections, these ovaries also ovulated oocytes. However, the number of oocytes recovered from experimental mice exhibited a time- and dose-dependent decline, and by 4 months, their number was significantly reduced. Neonatal exposures to MXC reduces ovulatory rates and ovarian functions in adults.

Enzyme histochemical investigation of glycol methacrylate embedded chick embryonic tissue.

SynopsisThe advantages of the water-soluble glycol methacrylate (GMA) embedding procedure make it highly applicable for use with fragile early embryonic material. Not only can one obtain tissue sections containing excellent histological detail, but numerous enzymes are retained for subsequent histochemical localization. For the purpose of establishing a methodology whereby concomitant histology and histochemistry could be obtainable, various fixatives and fixation times have been evaluated on GMA embedded chick embryonic mesonephros and gonad. It was found that fixing the tissues for 1 h in a solution of 95% ethanol, 5% acetic acid and 10% neutralbuffered formalin resulted in the retention of not only excellent histology but also alkaline and acid phosphatase. Thus, with this procedure, more specific investigations of early embryonic tissue can be performed.

Response of adult murine uterine epithelium to 50% methoxychlor

This study was designed to assess the response of uterine epithelia of adult mice to a 4-week exposure of 50% methoxychlor (MXC) to ascertain whether significant changes were induced by 50% MXC that might compromise future implantation. Sexually mature virgin female mice were exposed to 0.1, 0.5, 1.0, 2.5, or 5.0 mg MXC via oral gavage for 5 consecutive days for 4 weeks. Controls received either sesame oil or 25 micrograms estradiol-17 beta (E-17 beta) also by gavage. At sacrifice, segments from each uterine horn were prepared for morphometric studies or for transmission electron microscopy. Results revealed a dose-dependent increase in the heights of uterine epithelial cells. Epithelial cell heights of the two groups treated with the highest doses of the pesticide were similar to that of the E-17 beta-treated group. Electron microscopy revealed increased vacuolization and swelling of mitochondria in cells of the 2.5 and 5.0 mg treated groups when compared to either of the control groups. In addition, there were effects on the number and size of microvilli in the uterine epithelial cells. The present study clearly demonstrates that a 4-week exposure of adult female mice to 50% MXC elicits significant estrogenic and toxic effects on the uterine epithelium.

Histologie Investigation Of Glycol Methacrylate Embedded Chick Embryonic Tissue

Specific light microscopic investigations (i.e. histochemical) of early embryonic material have always been beset by difficulties in processing and obtaining tissue sections of good quality. The advent of glycol methacrylate (GMA) as an embedding medium now provides a means to overcome these inherent problems with this tissue. Investigations were carried out to assess the histological results produced by different fixatives and times of fixation of GMA embedded 5‐day chick embryonic tissue. Optimum cellular preservation of all tissues occurred following fixation in a mixture of acetic acid, 95% ethanol and neutral buffered formalin (AAF). With the procedures described in this study, a new method is available for more comprehensive examination of all types of early embryonic material.

Long- and short-term effects of carbaryl exposure in chick embryos

Abstract The effects of carbaryl on chick embryos following 5 and 12 days of exposure were examined. Carbaryl was dissolved in two vehicles (acetone and sesame oil) and the roles of these two vehicles in avian embryogenesis were assessed. The dosages of carbaryl used were 10, 5, 2.5, and 1.0 mg. All injections were administered prior to incubation. Groups of embryos exposed for 5 days to carbaryl in sesame oil and acetone showed a significant increase in mortality over their respective vehicle controls. There were no indications of external abnormalities in any of the 5-day surviving embryos regardless of how they were exposed. Those embryos exposed to carbaryl for 12 days demonstrated a significant increase in mortality over similarly treated 5-day groups; however, a greater embryotoxicity still occurred in those embryos exposed to carbaryl in sesame oil. Subcutaneous edema was a common defect seen in embryos receiving the higher dosages of carbaryl in acetone. Lower-limb abnormalities were less commonly observed. Results of this study demonstrate an increased embryotoxicity of carbaryl in sesame oil as opposed to acetone at all dosages employed. It is also apparent that carbaryl can become incorporated into the embryo from the yolk sac within 5 days, but a longer period of exposure is necessary for external abnormalities to occur.