Ex vivo gill assays used as an alternative approach to fish experimentation

DE LA TORRE, FERNANDO R.; LOZANO, ISMAEL E.; SCARCIA, PAOLA

Montevideo

Encuentro; SETAC Latin America 15th Biennial Meeting; 2023

Fish acute toxicity studies are one of the mostfrequently conducted vertebrate ecotoxicology tests where death is the intendedendpoint, therefore, opportunities to Replace, Reduce, and Refine Regulatory ofthose tests has become of increasing importance. Addressing the need foralternatives, sub-organismal responses proved to be useful to assess chemicalstress and were referred to biochemical/physiological changes (biomarkers)measured in vivo or by in vitro bioassays using isolated cell lines exposed to extractedand enriched surface water. Interestingly the freshwater fish gills are a focalpoint for studies that seek to understand deleterious effects of variouscontaminants of emerging concern (EC). In this context, predicting fish acutetoxicity of chemicals by means gill ex vivo assay is an alternative method tothe conventional approach in line with the 3Rs ideas. The aim of this study wasto assess the adverse response of selected oxidative stress biomarkers indifferent model fish species after ex vivo gill exposure to ECs (fipronil, Fp;triclosan, Tcs; nanosilver, AgNPs and ivermectin, Iv). Alternative studies wereconducted in order to compare: Fp in vivo and ex vivo responses; the mitigationeffect of humic acids on the toxicity of NPs and the photolytic degradation ofTCS. These short-term assays were conducted with native (Prochilodus lineatus,Cyphocarax voga, Piaractus mesopotamicus, Corydoras paleatus) and standardized species(Cyprinus carpio) as a preliminary screening approach where environmentalrepresentative concentrations of ECs were assessed. Briefly, four fish wereused and four pairs of gill arches were obtained that were randomly assigned tofour treatment groups (n=5): freshwater fish saline solution (Ctrl); methanolin saline solution (CtrlSv); and two nominal concentrations of C1 and C2prepared from a methanol stock solution of the EC. After incubating in glasscontainers for 1h at 25°C, branchial arches were removed and kept at −80 °Cuntil biochemical analysis. Results indicated that biomarker responses were dependenton fish species and the assayed ECs and alternative assays gave rise tovaluable information. The ex vivo model provided a versatile way of exposurewith a reduced number of fish and generate rapid information from a key targetorgan of toxicity. Besides, native test species were able to generate responseswith several ECs showing response capacity even under environmentally relevantconcentrations.