A comparison of behavior, reproductive parameters and response to pollutant between wild-type, transgenic and mutant zebrafish: could they all be considered the same "zebrafish" ?
Résumé
Regulatory assays available from the OECD for the testing of chemicals, such as endocrine disrupting chemicals (EDC) testing (TG229, TG230), can be performed with zebrafish without any recommendation regarding the strain that should be used. In addition to endpoints defined in the TG, transgenic fish could inform on mechanisms underlying EDC effects. By avoiding interference with pigmentation during imaging, use of casper background may improve tests sensitivity. In the present study, we compared 4 zebrafish lines (wild type AB, casper, cyp19a1a-eGFP and cyp19a1a-eGFP-casper) to determine whether transgene insertion and mutations could modify their basal physiology and their response to pollutant, thereby impeding their use for EDC testing. We thus compared the behavior (anxiety, sociability) and reproductive parameters (circulating vitellogenin (VTG) concentrations, female fecundity, egg viability) of control fish of the 4 lines and evaluated their responses to a model pollutant, e.g. clotrimazole. Sociability was evaluated by monitoring the time spent near congeners in a shuttle box while anxiety was evaluated using the novel tank diving test. No difference was observed between lines for either sociability or anxiety level. Concerning reproduction, no significant difference in the number of eggs laid per female, in the viability of eggs or in the female circulating VTG concentrations was noted between the 4 lines studied. Clotrimazole exposure (28 days; 250 µg/L) had similar effects in the 4 zebrafish lines despite a slightly higher sensitivity in wild type AB. Clotrimazole led to a masculinization of one part of the females, an inhibition of VTG, an inhibition of aromatase A expression (GFP measurement and qPCR) and an induction of 11-ketotestosterone. In summary, no critical difference in the behavior, reproductive parameters and response to pollutant was found between the 4 zebrafish lines, indicating that the transgene insertion and the mutations did not modify basal physiology of the fish. Therefore, their use in OECD tests for the study of EDC appears to be relevant. Further work will be conducted on strains sensitivity by evaluating dose-response effects of pollutants.