Approved Abstracts

DNA damage and malformations in zebrafish exposed to psychoactive metaphedrone



Author(s): Ribeiro O.; Saavedra E.; Gaivão I.; Félix L.; Ribeiro C.; Tiritan M.E.; Carrola J.S.;
Presenter: João Soares Carrola

Synthetic cathinones (SCat) are psychostimulants substances that emerged onto drug markets as “legal” alternatives to illicit drugs. Their abuse still represents a serious public health issue and a threat to both terrestrial and aquatic wildlife. In fact, due to their incomplete removal in wastewater treatment plants, SCat can be found in aquatic ecosystems. Despite the low levels usually found, they can negatively impact non-target organisms like fish. Metaphedrone (3-MMC) is a synthetic cathinone, initially synthesized to replace mephedrone, due to its prohibition in European countries. The assessment of fish genetic damage is important as it can have individual and populations level consequences even at relevant environmental concentrations under chronic exposure and/or early life stages. Thus, the aim of this work was to evaluate the toxic effect of metaphedrone in zebrafish, "Danio rerio", focusing on DNA damage and malformations. For this, 50 embryos with 2-3 hours post-fertilization (hpf) were distributed in 5 replicates and exposed to various concentrations of 3-MMC (0.1, 1, 10 and 100 μg/L). Zebrafish malformations and DNA damage were analysed at 72 and 96 hpf, respectively. Malformations were assessed using a stereo microscope. For genetic damage evaluation, 2 slides (4 gels) were performed per treatment to realize the comet assay technique (single-cell gel electrophoresis). The slides were analysed by fluorescence microscopy and 100 aleatory comets were sorted in five classes: 0 to 4 arbitrary units (AU) to determine the genetic damage indicator (GDI). The data were subjected to an analysis of variance and respective comparison of means to assess the effect of the concentration of 3-MMC on the malformations and DNA damage. No statistically significant effects were observed for zebrafish malformations however, a significant increase in genetic damage was detected in animals exposed to the higher concentration of 3-MMC compared to the control. This increase in DNA damage may be related to oxidative stress, DNA adducts, alkylation of bases or other factors. Besides, addiction and behavior alteration was recently found in brown trout "Salmo trutta" at environmentally relevant concentrations (1 µg L−1) exposed to psychostimulants, namely, methamphetamine.
Our data evidenced that 3-MMC is not teratogenic for zebrafish but can lead to genotoxic problems for the higher concentration tested (100 μg/L) presenting potential negative consequences like neurological and behavioural problems, and also carcinogenicity. However, more studies are needed to confirm these data and investigate deeper the impacts of 3-MMC in fish including the potential transmission of human societal problems to aquatic ecosystems.
Funding: This work was supported by National Funds by FCT - Portuguese Foundation for Science and Technology, under the project UIDB/04033/2020 and PTDC/CTA-AMB/6686/2020.
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Keywords: Genotoxicity; zebrafish (Danio rerio); metaphedrone

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