Unveiling enantioselective mechanisms of toxicity of butylone in Daphnia magna

Butylone (BTL) is a chiral synthetic cathinone widely abused due to its stimulant and hallucinogenic effects. It is available as a racemate and reported as a contaminant in wastewater. Our previous studies revealed BTL toxicity and of its enantiomers to non-target species. Nevertheless, the mechanisms underlying its toxicity remain poorly understood, particularly after sub-chronic exposure. This study aimed to expand the understanding of the ecotoxicity of BTL and its enantiomers by studying the potential mechanisms of toxicity after 15 days of exposure using Daphnia magna freshwater model. For that, neonates (<24 h) were exposed to an environmentally relevant concentration (0.10 μg L^-1) of the racemate (R,S)-BTL or each pure enantiomers, (R)-BTL or (S)-BTL, to study various key endpoints (morphophysiological, biochemical, reproductive, swimming activity, and gene expression). Results showed that both racemate and (S)-BTL exposure significantly induced triacylglycerol lipase (LIP4) gene expression linked to lipid metabolism, negatively affecting the growth and body size of the specimens exposed. Moreover, neurotoxicity was evidenced by the inhibitory response on acetylcholinesterase activity (AChE) and the upregulation of the expression of genes involved in the synthesis of dopamine and serotonin. Yet, swimming activity, reproduction and survival of D. magna remained unaffected. Enantioselective effects were observed in body growth (decrease (S)-BTL and increase (R)-BTL), heart size and area (increase (R)-BTL), and aromatic l-amino acid decarboxylase (AAAD) transcription levels (all BTL forms). Overall, the findings highlighted that environmentally relevant concentrations of BTL can lead to adverse effects in D. magna, confirming the potential threat of this illicit drug to this freshwater invertebrate, and can impact the primary consumers and aquatic food webs and their ecosystems.