Microinjection into zebrafish embryos (Danio rerio) - a useful tool in aquatic toxicity testing?

Abstract

Background: Microinjection was tested as a potentially powerful tool to introduce natural and anthropogenic pollutants directly into fish eggs to determine their toxicological impact on fish. With this technique, parental transfer of lipophilic contaminants may be mimicked. Here, we investigated the applicability of pollutant injection into the yolk of early zebrafish (Danio rerio) eggs with special regard on survival after vehicle injection. Tested vehicles were autoclaved tap water, dimethyl sulfoxide (DMSO), methanol, and triolein. Results: Highest mortality occurred after the injection of DMSO and methanol. The lethality rates were up to 40% higher than under control conditions. Best survival rates were obtained after triolein and water injections. However, the triolein droplet was not assimilated by the embryo within 96 h post fertilization suggesting an incomplete uptake of triolein-solubilized chemicals. Technical aspects concerning microinjection in zebrafish eggs are discussed with special emphasis on quantitative injection. Conclusions: Microinjection into the yolk cell of zebrafish eggs is feasible, but the application of exact volumes appears problematic. However, microinjection is a powerful tool for studies without the demand for high volume accuracy. Adopting microinjection for pollutant research requires further investigation.

Advancing the Zebrafish embryo test for endocrine disruptor screening using micro‐injection: Ethinyl estradiol as a case study

“…Neither canola oil nor DMSO affected any of the toxicity endpoints. Dimethylsulfoxide has already been used for the injection of zebrafish embryos in previous studies (Maes et al ; Schubert et al ; Li et al ), but triolein is the more commonly used solvent in injection studies with fish embryos (Edmunds et al ; Hano et al ; Nassef et al ; Schubert et al ; Xiao et al ). Our results, however, showed that the use of triolein can lead to effects on survival and heart rate.…”

“…Out of these 2 potential solvents, DMSO was selected as the optimal solvent for the present study because the high viscosity of canola oil caused practical problems. In addition, DMSO has already been used in similar micro-injection studies with zebrafish (Maes et al 2012;Schubert et al 2014;Li et al 2015). The variation among 3 different injection needles and within a single injection needle (3 replicates) is shown in Figure 2B.…”

“…From 2000 onward, a medaka fertilized egg nanoinjection assay has been in use to assess the toxicity of a variety of compounds, often with the aim of mimicking deposition in eggs through maternal transfer (Edmunds et al 1999;Villalobos et al 2000;Colman and Ramsdell 2003;Nassef et al 2010). More recently, studies have been published that applied micro-injection in zebrafish embryos to determine the toxicity of diethyl phthalate (DEP), perfluorooctanoic acid (PFOS), and triphenyltin (among other chemicals; Liu et al 2012;Schubert et al 2014;Kim et al 2015;Li et al 2015;Xiao et al 2017).…”

“…In the most recent studies using micro-injection in zebrafish embryos, different solvents have been used, including ethanol (Kim et al 2015), dimethyl sulfoxide (DMSO; Li et al 2015), phosphate-buffered saline (PBS; Liu et al 2012), and triolein (Xiao et al 2017). In the studies of Maes et al (2012) and Schubert et al (2014), the suitability of different solvents for micro-injection in zebrafish embryos was compared. Acetone, DMSO, and triolein were examples of solvents found to cause limited toxicity.…”

Biological evaluation of microbial toxin degradation by microinjected zebrafish (Danio rerio) embryos

“…Although, microinjection of substances into the yolk of zebrafish eggs is feasible, introduction of accurate volumes (e.g. constant volumes) through a series of injections seems to be problematic so nominal and real injected volumes may be different (Schubert et al, 2014).…”