In vivo studies of fishes exposed to xenoestrogens have reported vitellogenin (Vtg) induction, ovatestes, altered sex ratios, and impaired reproductive capacity. The objective of this study was to determine concentration dependent effects of a weak estrogen receptor agonist, 4-nonylphenol (NP) and a potent estrogen receptor agonist, 17alpha-ethinylestradiol (EE) on sex ratios, gonad morphology, Vtg induction and breeding success in zebrafish (Danio rerio). Fish were exposed from 2 to 60 days post-hatch (dph) to NP (10, 30, or 100 microg/l nominal), EE (1, 10, or 100 ng/l nominal), or solvent control (acetone; 0.2% v/v) in a static-renewal system with replacement every 48 h. At 60 dph, 20 fish from each treatment were euthanized for histological examination of gonads and Western blotting for Vtg using pooled heart homogenates. Remaining fish were reared in clean water until adulthood (120 dph) for breeding studies. Due to high mortality in the 100 ng/l EE group, insufficient fish were available for analyses. The percentage of males at 60 dph changed from 45% (9/20) in solvent controls to 0% at 10 ng/l EE and 10% at 100 microg/l NP. A concentration dependent increase in the number of fish with undeveloped gonads at 60 dph was observed in the EE exposure group. Two fish with ovatestes were observed at 100 microg/l NP, while one was observed at 30 microg/l NP. Western blotting showed induction of Vtg at 30 and 100 microg/l NP and 10 ng/l EE. The sex ratios of adults determined at 160 dph revealed no significant departure from 1 male:1 female, suggesting that exposure of zebrafish to estrogenic chemicals during sexual differentiation and early gametogenesis did not irreversibly alter phenotypic sex. The condition factor of adult fish and ovo-somatic index of adult female fish were also unaffected by developmental exposure to NP or EE. Despite this, breeding trials conducted in adult fish from 120 to 160 dph revealed significant reductions in the percent of viable eggs, hatchability, and swim-up success at 10 ng/l EE and 100 microg/l NP. Our results suggest that functional reproductive capacity (breeding success) may be more sensitive than gross morphological endpoints (condition, ovo-somatic index, sex ratio) in adult zebrafish exposed to xenoestrogens during sexual differentiation and early gametogenesis.

Animals↗

PubMed2003

Developmental estrogenic exposure in zebrafish (Danio rerio): II. Histological evaluation of gametogenesis and organ toxicity.

Aquatic species can be exposed to endocrine disrupting chemicals (EDCs) in wastewater that often includes the weak estrogen, 4-nonylphenol (NP) and the potent estrogen, 17alpha-ethinylestradiol (EE). The goal of the present study was to determine concentration-dependent effects of developmental exposure to NP and EE on gametogenesis, as well as gonad, kidney and liver pathology using quantitative histological evaluation of hematoxylin/eosin-stained saggital sections of zebrafish (Danio rerio). The major finding of the present study was that exposure to NP (>/=100 microg/l nominal) and EE (>/=1 ng/l nominal) from 2 to 60 days post-hatch (dph) caused concentration-dependent suppression of gametogenesis in both male and female zebrafish. Severe kidney pathology was observed in 60 dph zebrafish, specifically glomerular dilation or degeneration, fibrosis, tubule enlargement and tubule necrosis, at a threshold of 10 ng/l EE. However, minor kidney histopathology indicated by increased pyknotic nuclei in kidney tubule and interstitial (hematopoietic) cells was detected at lower estrogenic exposures (>/=10 microg/l NP nominal) than delayed gametogenesis. Considering all histological parameters in the current study, the rank order of potency for pathological effects in 60 dph zebrafish was 10 ng/l EE>1 ng/l EE=100 microg/l NP>30 microg/l NP>10 microg/l NP10 (nominal concentrations). Zebrafish from the same cohort examined in the current study that had been placed in clean water from 60 to 300 dph had histologically normal testes and no kidney or liver histopathology. However, increased ovarian follicle atresia was detected at 300 dph in zebrafish exposed developmentally to 100 microg/l NP. Therefore, we conclude that functional rather than morphological changes may be more important for future evaluations of developmental exposure to estrogens in fish, and that negative effects in female rather than male gonads may contribute to prolonged breeding impairment.

Animals↗

PubMed2002