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Abstract The aim of this study was to explore the relationships among gut microbiota disturbances and serum and spinal cord metabolic disorders in neuropathic pain. 16S rDNA amplicon sequencing and serum and spinal cord metabolomics were used to identify alterations in the microbiota and metabolite profiles in the sham rats and the chronic constriction injury (CCI) model rats. Correlations between the abundances of gut microbiota components at the genus level, the levels of serum metabolites, and pain-related behavioural parameters were analysed. Ingenuity pathway analysis (IPA) was applied to analyse the interaction networks of the differentially expressed serum metabolites. First, we found that the composition of the gut microbiota was different between rats with CCI-induced neuropathic pain and sham controls. At the genus level, the abundances of Helicobacter, Phascolarctobacterium, Christensenella, Blautia, Streptococcus, Rothia and Lactobacillus were significantly increased, whereas the abundances of Ignatzschineria, Butyricimonas, Escherichia, AF12, and Corynebacterium were significantly decreased. Additionally, 72 significantly differentially expressed serum metabolites and 17 significantly differentially expressed spinal cord metabolites were identified between the CCI rats and the sham rats. Finally, correlation analysis showed that changes in the gut microbiota was significantly correlated with changes in serum metabolite levels, suggesting that dysbiosis of the gut microbiota is an important factor in modulating metabolic disturbances in the context of neuropathic pain. In conclusion, our research provides a novel perspective on the potential roles of the gut microbiota and related metabolites in neuropathic pain.