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GESTATIONAL ATRAZINE EXPOSURE IN THE RAT: EFFECTS ON MAMMARY GLAND DEVELOPMENT AND FUNCTION IN MULTIPLE GENERATIONS
Citation:
FENTON, S. E. GESTATIONAL ATRAZINE EXPOSURE IN THE RAT: EFFECTS ON MAMMARY GLAND DEVELOPMENT AND FUNCTION IN MULTIPLE GENERATIONS. Presented at Atrazine Workshop, Iowa City, IA, April 26 - 27, 2005.
Description:
The chlorotriazine herbicides currently represent the most heavily used of all agricultural pesticides, with atrazine being the most common of these chemicals. Rodent toxicology studies indicate that atrazine can disrupt endocrine function and among its effects is an increased incidence of mammary tumor development in adult Sprague Dawley rats following lifetime dietary exposure. In fact, increased incidence of mammary tumors in this animal model was the first outcome that flagged atrazine for regulatory concern. Subsequent studies suggested that this effect was secondary to altered endocrine function resulting in premature reproductive senescence, during which time the persistent serum estrogen levels in the rat may act to promote tumors and enhance replication in estrogen-sensitive neoplastic cells of spontaneously initiated tumors. This type of reproductive aging does not occur in humans and therefore this mode of action of atrazine-induced tumors is limited to rat models. However, multiple laboratories have shown that atrazine disrupts the hypothalamic control of pituitary-ovarian function of rats exposed as adults, including a decrease in circulating prolactin and luteinizing hormone levels, an atrazine mode of action that is applicable to humans. The rodent and human mammary gland share many similarities in their early development: epithelial bud and primary ductal structures form during fetal life; outgrowth of that bud into the underlying mesenchymal fat pad takes place from birth to adolescence; and, development of an extensive ductal tree, ending in terminal end buds, occurs during puberty. Furthermore, these developmental events are regulated by hormones shown to be perturbed by environmental exposures. Because of these similarities, rodent models have been used for decades to evaluate the effects of endogenous (i.e., hormones, growth factors) and exogenous (i.e., pharmaceuticals, pollutants) components on normal mammary gland and mammary tumor development. It has been demonstrated by several laboratories that terminal end buds are sensitive to tumor initiation as they are undergoing rapid proliferation and differentiation and it is widely believed that the stage of development is key in susceptibility to carcinogenesis. Developing organisms have been shown to have increased susceptibility to tumor development if environmentally exposed during rapid growth and differentiation stages. We hypothesize that environmental pollutants capable of acting as endocrine disruptors perturb the normal development of the mammary gland if exposure coincides with sensitive periods of development for the gland. We present data, using atrazine as a model compound, to support this hypothesis. We have evaluated atrazine exposure parameters required for pubertal delays in Long Evans (LE) rats, including mammary gland development, using cross-fostering to determine if effects were strictly dam mediated (via milk) or were a direct effect (transplacental) on the pups. Timed pregnant LE rats (N=20/treatment group) were gavaged gestational days (GD) 15 19 with 100 mg atrazine/kg body weight (BW) or vehicle (controls, C). On postnatal day (PND)1, half of all litters were cross fostered, creating 4 treatment groups; C C, ATR C, C ATR, and ATR ATR (dam milk source, respectively). A significant delay in vaginal opening (VO) and increase in VO BW was seen only in the litters receiving milk from ATR exposed dams. However, mammary glands of female offspring (2/dam) in all groups exposed to ATR (ATR C, C ATR, and ATR ATR) displayed significant delays in epithelial development. These changes were detected as early as PND4 and stunted development was evident through PND40. Further, at all developmental stages examined, offspring in the ATR-ATR group exhibited the least developed glands. These delays in pubertal endpoints do not appear to be related to pup body weight or endocrine hormone concentrations. Our data suggest that the delay in VO of in utero ATR exposed offspring is mediated via the dam [milk], whereas direct exposure to atrazine in utero can cause delays in mammary gland development. Our data suggest that milk-derived factors (growth factors or hormones), in addition to transplacental exposure during mammary bud outgrowth may be involved in atrazine s mode of action on delayed mammary gland development. To determine the window of fetal mammary development most sensitive to the effects of atrazine, timed-pregnant LE dams (N=8/group) were prenatally dosed for 3 or 7-day intervals (GD13-15, 15-17, 17-19, or 13-19) with 100 mg atrazine/kg/d or vehicle (C), and their offspring evaluated for changes. Mammary glands taken from pups exposed to atrazine as early as PND4 displayed significant delays in epithelial development compared to C, with continued developmental delays at later time points that varied by time of exposure. However, the most persistent and severe delays were seen in the GD17-19 and GD13-19 atrazine exposure groups, demonstrating statistically similar growth retardation. Because mammary developmental deficits persisted into adulthood, we hypothesized that atrazine-exposed animals may have difficulties nursing their young. Females exposed prenatally to either atrazine (as defined) or C (N>4 litters/group) were bred and the F2 offspring from GD17-19 and GD13-19 exposure groups were significantly smaller in BW than C. The F2 offspring in these two atrazine exposure groups had significantly delayed mammary gland development compared to controls, but when mammary gland development data were analyzed using BW as a cofactor, glands from GD17-19 and 13-19 pups were no longer considered significantly delayed. This result in the F2 pups is thought to be a secondary effect of the delayed mammary development in the F1 dam, an outcome not affected by BW. In a separate study, it was determined that atrazine (8.7 to 100 mg/kg), delivered from GD15-19, did not decrease fetal body weights on GD20, even though the higher doses significantly decreased weight gain by the dosed dams. These data suggest that the consequences of brief ATR exposure, during a critical period of fetal mammary gland development (GD17-19), are both delayed mammary development of the offspring and inadequate nutritional support of F2 offspring, resulting in adverse effects on pup weight gain. Whether the developmental changes induced by exposure to relatively high doses of atrazine alter the offspring s susceptibility to mammary tumor development is not known.