Methcathinone disrupts glycerophospholipid metabolism and selectively activates the p38/COX-2 axis to induce anxiety-like behaviour in zebrafish.

Methcathinone, a synthetic cathinone derivative, is an emerging contaminant in aquatic environments, raising concerns about its neurotoxic effects on aquatic organisms. However, the fundamental molecular mechanisms underlying its anxiogenic and behavioural effects remain unclear. Here, adult zebrafish were administered sublethal doses of methcathinone intraperitoneally for three consecutive days, and behavioural changes were assessed using the novel tank, mirror attack, and social preference tests. Integrated metabolomic and transcriptomic analyses were conducted on brain tissues, followed by correlation network analysis and quantitative real-time PCR (qPCR) validation. Methcathinone exposure induced anxiety-like behaviour, increased aggression, and altered social preference. Metabolomic profiling revealed selective depletion of glycerophospholipids, whereas transcriptomic analysis revealed dysregulation of the MAPK signalling pathway. Correlation analysis demonstrated tight coupling between glycerophospholipid metabolites and genes involved in MAPK signalling and neuroactive ligand-receptor interactions. qPCR validation confirmed the downregulation of genes associated with glycerophospholipid synthesis, membrane remodelling, neurotrophic support, GABAA receptor subunits, and the CB1 receptor, alongside the upregulation of stress-responsive genes (mapk14b and ptgs2a). Pharmacological rescue with the p38 inhibitor SB203580 reversed the anxiety-like behavioural deficits and the associated downstream molecular changes, confirming the causal role of the p38/COX-2 pathway. Thus, methcathinone induces anxiety-like behaviour through a coordinated "lipid-signalling-neurotransmitter" axis. While strongly correlated with upstream glycerophospholipid depletion, the anxiety phenotype is directly driven by selective p38/COX-2 activation, causing impaired GABAergic and cannabinoid inhibitory neurotransmission. This study provides a foundational mechanistic framework for methcathinone-induced neurotoxicity and highlights potential molecular biomarkers for future evaluations of the ecological risks associated with synthetic cathinone contamination in aquatic systems.