A multi‑target adverse outcome pathway network for sub-chronic ketamine‑induced cognitive impairment in adult...

Ketamine (KET), an emerging contaminant, is increasingly detected in aquatic environments, yet its sub-chronic neurocognitive effects at environmentally relevant concentrations remain largely unknown. We exposed adult zebrafish to KET for 40 days and integrated network toxicology, transcriptomics, behavioral assays, and molecular simulations to investigate these effects. Sub-chronic KET exposure impaired spatial learning and memory at 0.4 μg/L, while anxiety‑like and social behavior were altered only at 40 μg/L. Intersecting transcriptomics with network toxicology yielded 9 candidate genes and 15 shared KEGG pathways, which were mainly associated with neurocognitive function. SHAP prioritized hmox1, camkk2, and trem2 and molecular simulations predicted stable binding of KET to these proteins, supporting their role as candidate molecular initiating events. Gene expression changes correlated with altered glutamate, and GABA levels. Based on these multi‑level findings, we constructed a proposed adverse outcome pathway (AOP) network in which KET is predicted to bind to HMOX1, CAMKK2, and TREM2, which may contribute to oxidative stress, neuroinflammation, and synaptic dysfunction converging on cognitive decline. This study provides evidence in adult zebrafish that sub-chronic exposure to environmentally relevant KET impairs cognition and proposes a multi‑target AOP network based on zebrafish data, supporting ecological risk assessment of KET as an emerging contaminant.