Gut Microbiota Dysbiosis Mediates Podophyllotoxin-Induced Neurotoxicity in Zebrafish by Modulating Purine Metabolic and Synaptic Dysfunction

Podophyllotoxin (PPT), a natural plant-derived insecticide, has raised concerns due to its significant cytotoxicity and potential neurobehavioral toxicity to non-target organisms, despite its widespread use. The mechanisms underlying this toxicity are not fully understood. This study employed zebrafish as a vertebrate model to comprehensively evaluate the behavioral impacts and mechanistic basis of PPT-induced neurotoxicity using integrated behavioral tests, histopathological analysis, and multi-omics approaches. Zebrafish exposed to various concentrations of PPT (0.997, 1.95, 3.91, 7.81, 15.6, and 31.2 ug/mL) for two days exhibited marked intestinal and neurotoxicity. Behavioral assays showed that PPT significantly impaired cognitive performance. Histological analysis revealed notable structural disruptions in both brain and intestinal tissues. 16S rRNA sequencing indicated that PPT exposure led to reduced gut microbial diversity and richness, with significant increases in the relative abundances of Acinetobacter and paraperlucidibaca. Metabolomic profiling showed PPT altered the purine metabolism pathway, reducing neuroprotective metabolites (adenosine and inosine) and increasing neurotoxic compounds (quinolinic acid and oxalic acid). Transcriptomic analysis revealed differentially expressed genes linked to neuronal/synaptic damage and inflammation pathways. qPCR confirmed that PPT suppressed synaptic functional related genes (GAD1b, DRD2b) and upregulated complement pathway-related genes (c3a.1, c1qc) and pro-inflammatory cytokines (IL-6, IL-1beta, TNF-alpha). Correlation analysis highlighted strong associations between Acinetobacter and paraperlucidibaca abundances and altered purine metabolite levels, as well as genes involved in synaptic and inflammatory signaling.