Using Tg(Vtg1:mcherry) Zebrafish Embryos to Test the Estrogenic Effects of Endocrine Disrupting Compounds.

There are many endocrine disrupting compounds (EDC) in the environment, especially estrogenic substances. The detection of these substances is difficult due to their chemical diversity; therefore, increasingly more effect-detecting methods are used, such as estrogenic effect-sensitive biomonitor/bioindicator organisms. These biomonitoring organisms include several fish models. This protocol covers the use of zebrafish Tg(vtg1: mCherry) transgenic line as a biomonitoring organism, including the propagation of fish and the treatment of embryos, with an emphasis on the detection, documentation, and evaluation of fluorescent signals induced by EDC. The goal of the work is the demonstration of the use of the Tg(vtg1: mCherry) transgenic line embryos to detect estrogenic effects. This work documents the use of transgenic zebrafish embryos Tg(vtg1: mCherry) for the detection of estrogenic effects by testing two estrogenic substances, α- and ß-zearalenol. The described protocol is only a basis for designing assays; the test method can be varied according to the test endpoints and the samples. Moreover, it can be combined with other assay methods, thereby facilitating the future use of the transgenic line.

Investigation of Fertilizing Capacity of Zebrafish (Danio rerio) Sperm Exposed to Heavy Metals.

The objective of our study was to investigate the effects of heavy metals on the fertilizing capacity of exposed zebrafish sperm, on embryonic survival, and on occurrence of embryonic deformities following fertilization with exposed sperm. It is important to test heavy metals because they are well-known pollutants. Sperm of externally fertilizing species can get in contact with pollutants found in aquatic environment. Zebrafish sperm, despite its advantages, has seldom been used in in vitro toxicological studies and no reports are available regarding the fertilizing capacity of exposed sperm. Zebrafish sperm was stripped and exposed to concentrations of the tested heavy metals (Zn2+, Cd2+, Cr3+, Cu2+, Ni2+, Hg2+, As3+) for 30 or 120 minutes. Calculated half-maximal effective concentration (EC50) values do not differ significantly from those calculated for motility for any of the tested heavy metals, which means fertilization rate can indicate the toxicity of the given substance following exposure of sperm. Thus, its application as in vitro toxicological end point is reasonable. The survival of embryos and embryonic development have not been affected by the exposure of spermatozoa, which means all alterations in spermatozoa caused by heavy metals have been expressed before 24 hours post fertilization.