BackgroundSeveral chemicals in the environment have the potential to inhibit aromatase, an enzyme critical to estrogen synthesis.ObjectivesThe objective of this study was to provide a detailed characterization of molecular and biochemical responses of female fathead minnows to a model aromatase inhibitor, fadrozole (FAD).MethodsFish were exposed via water to 0, 3, or 30 μg FAD/L for 8 days and then held in clean water for 8 days, with samples collected at four time points during each 8-day period. We quantified ex vivo steroid production, plasma steroids, and plasma vitellogenin (Vtg) concentrations and analyzed relative transcript abundance of 10 key regulatory genes in ovaries and 3 in pituitary tissue by real-time polymerase chain reaction.ResultsEx vivo 17β-estradiol (E2) production and plasma E2 and Vtg concentrations were significantly reduced after a single day of exposure to 3 μg or 30 μg FAD/L. However, plasma E2 concentrations recovered by the eighth day of exposure in the 3-μg/L group and within 1 day of cessation of exposure in the 30-μg/L group, indicating concentration- and time-dependent physiologic compensation and recovery. Concentration-dependent increases in transcripts coding for aromatase (A isoform), cytochrome P450 side-chain cleavage, steroidogenic acute regulatory protein, and follicle-stimulating hormone receptor all coincided with increased E2 production and recovery of plasma E2 concentrations.ConclusionsResults of this research highlight the need to consider compensation/adaptation and recovery when developing and interpreting short-term bioassays or biomarkers or when trying to predict the effects of chemical exposures based on mode of action.

This study explored the possibility that environmental estrogens in sewage effluent may reduce the reproductive fitness of adult male fish by suppressing their reproductive behaviors, including their ability to compete for nests and females. Male fathead minnows (Pimephales promelas) were exposed for three weeks to either blank control, effluent released by a sewage treatment plant (STPE), waterborne estradiol (E2), or a synthetic androgen (methyltestosterone [MT]). Afterward, fish were placed with females and a nest, and their behavior was monitored for 5 d in either the presence or the absence of a competing (unexposed control) male. Males exposed to either the STPE or E2 (approximately 50 ng/L, a level chosen to mimic the estrogenic content of the STPE) had elevated levels of circulating vitellogenin (p < 0.05) and lower levels of 11-ketotestosterone (KT; p < 0.05). Nearly all STPE- and E2-exposed males spawned successfully in the absence of a competing male, but in both cases, exposed males suffered nearly total reproductive failure when they had to compete. Conversely, males exposed to MT (approximately 50 ng/L) outcompeted control males. Behavioral observations suggested that subtle differences in agonistic behaviors, typically associated with circulating androgens (i.e., KT), were responsible. We speculate that male fish exposed to estrogenic effluent in the field are less likely to reproduce successfully within large populations of wild fish, thereby causing abnormal and potentially detrimental patterns of gene flow within those populations.

This report presents the study design, environmental data, and quality-assurance data for an integrated chemical and biological study of selected streams or lakes that receive wastewater-treatment plant effluent in Minnesota. This study

The objective of the present study was to characterize responses of the reproductive endocrine system of the fathead minnow (Pimephales promelas) to the fungicide vinclozolin (VZ), using a 21-d reproduction assay, and a shorter-term (approximately two weeks) test in which fish were cotreated with the VZ (a putative anti-androgen) and the androgen 17beta-trenbolone (TB). Effects on fecundity, gonadal histology, secondary sexual characteristics, reproductive hormones, and relative abundance of androgen receptor (AR) and 11beta-hydroxysteroid dehydrogenase (11betaHSD) mRNA transcripts were evaluated in one or both of these studies. Fecundity of VZ-exposed fish was decreased in a concentration-dependent manner in the 21-d test, culminating in complete reproductive failure at a concentration of 700 microg/L. Exposure to VZ decreased expression of male secondary sexual characteristics -- an effect typical of anti-androgens. The finding that exposure of females to TB-induced expression of prominent, male-like tubercles, which could be effectively blocked with VZ, provides powerful evidence of the anti-androgenic activity of VZ in vivo. In the two experiments VZ produced several responses possibly indicative of compensation or adaptation of the fish to the anti-androgen, including increases in gonad weight, AR and 11 betaHSD mRNA transcript abundance, and ex vivo gonadal production of testosterone and 11-ketotestosterone. Overall, our results demonstrate that the model anti-androgen VZ, which also is an environmental contaminant, impairs reproductive success of fathead minnows and elicits endocrine responses consistent with an anti-androgenic mode of action.