Zebrafish (Danio rerio) as a model to study developmental effects of endocrine disruption: molecular mechanisms, as well as persistence and reversibility of effects

The present study aimed to investigate the reversibility of endocrine disruption in zebrafish (Danio rerio) at different effect levels. For this purpose, three different endocrine disrupting chemicals (EDCs) were chosen according to their different modes of action. 17!- Ethinylestradiol (EE2), a semi-synthetic estrogen is the most popular substance in oral contraceptives and is regularly measured in surface waters. 17ß-trenbolone is an anabolic steroid, which binds with high affinity to the androgen receptor. The substance is used in industrial animal farming and sports as muscle growth promoter. Prochloraz is a popular fungicide with multiple modes of action. Its main effect on the endocrine system is that it inhibits the enzyme aromatase, which is essential for the conversion of androgens into estrogens. The effects of those three EDCs were assessed by performance of exposure experiments with developing zebrafish. After exposure of 60 days (from fertilized egg to sexual differentiation), half of the fish were continuously exposed until 100 days post hatch and the other half was held in clean water. In addition, this project focuses on assessing the correlation between different levels of biological organization. For this purpose, five effect levels with different ecological relevance were investigated: (1) population level: sex ratio; (2) individual level: growth; (3) organ/cell level: histology of gonads; (4) protein level: vitellogenin (VTG) induction and (5) mRNA level: aromatase (cyp19b) expression in brain. The three different EDCs investigated in this study showed strong impact on the sexual development of zebrafish at all effect levels at environmentally relevant concentrations. For trenbolone and prochloraz we could only find tendencies of reversibility, most effects remained unchanged after 40 days of depuration in clean water. Despite different underlying mechanisms, these substances produce an irreversible and considerable drift of the sex ratio towards males, as well as permanent effects on growth, VTG and aromatase levels. A clear reversibility of those effects could only be shown for EE2. Even at population level the impact of the semi-synthetic estrogen was reversible. These results show that the sexual development of zebrafish is a fragile process that can easily be disrupted permanently by substances that are found in the environment. Moreover, the results indicate that even a periodic exposure to those EDCs can cause severe impairment for wildlife and humans