The Role And Mechanism Of Microbiota-Derived Tryptophan Metabolites In Cognitive Impairments Induced By Chronic Neuropathic Pain

【 Background 】 Patients with chronic neuropathic pain(CNP)often suffered from cognitive impairments,but the underlying mechanism remains elusive.Gut microbiota and its derived tryptophan metabolites play vital roles in CNP and cognitive impairment via“microbiota-gut-brain” axis.However,the role of gut microbiota and its derived metabolites in CNP-induced cognitive impairments remains to be explored.【Methods】(1)Spared nerve injury(SNI)was applied to imitate CNP.SNI mice were clustered into cognitive impairment(CD)group and non-cognitive impairment(NCD)groups based on their performances in the Morris water maze test.Fecal and plasma samples were collected from Sham,CD and NCD group mice for 16 S r RNA sequencing,nontargeted and targeted metabolomics analysis.(2)SNI mice were treated with low(0.4 g/kg),normal(2 g/kg)or high(10 g/kg)-tryptophan diet respectively,and behavioral tests were used to evaluate the allodynia and cognitive function in mice.Mice were treated with hightryptophan diet after antibiotic compounds,and then behavioral tests,RT-PCR and targeted metabolomics analysis of tryptophan metabolites were performed to explore the role of gut microbiota in high-tryptophan diet modulating CNP-induced cognitive impairments.(3)Differential tryptophan metabolite(indole-3-propionic acid,IPA),Ah R agonist(FICZ)and Ah R antagonist(CH223191)were given to SNI mice and then behavioral tests were performed to evaluate its effects on pain and cognition in SNI mice.After administration of differential tryptophan metabolite and Ah R antagonist in SNI mice,the behavioral tests,ELISA,Western blot and RT-PCR were performed to detect changes of Ah R and central oxidative stress indicators in prefrontal cortex(PFC)and hippocampus.【Results】(1)The microbiota composition in CD mice was featured with higher abundance of Actinobacteria,Proteus and Bifidobacterium,and lower abundance of Bacteroides.Nontargeted metabolomics results showed tryptophan metabolites were significant signatures of CD group mice.Further results of tryptophan metabolites targeted metabolomics found that IPA decreased and 3-indoxyl sulfate increased in CD mice.(2)Results showed that hightryptophan diet(10 g/kg)could alleviate allodynia and cognitive impairment-like behaviors in SNI mice.However,the clearance of gut bacteria by antibiotics significantly reduced the effects of high-tryptophan diet(HTP)on pain and cognition improvement in SNI mice.HTP diet could reverse the proliferation of Actinobacteria and γ-Proteobacteria and lower IPA caused by SNI.However,pretreatment with antibiotics abolished the effects of HTP diet on IPA in PFC and hippocampus of SNI mice.(3)IPA,FICZ and CH223191 treatment showed no effect on mechanical pain hypersensitivity but attenuated cognitive impairment in SNI mice.IPA and FICZ attenuated cognitive impairment in SNI mice.The benefits of IPA on cognition could be completely blocked by CH223191 treatment previously.IPA reversed the expression levels of Ah R,Cyp1a1,Nuclear factor erythroid 2-related factor 2(Nrf2),and Nqo1,and improved the levels of antioxidant enzyme(such as superoxide dismutase,catalase and glutathione peroxidase)in PFC and hippocampus of SNI mice.However,CH223191 pretreatment reduced the regulatory effects of IPA on the levels of Ah R-Nrf2 signaling and antioxidant enzyme in SNI mice.【Conclusions】(1)Gut microbiota imbalance played a vital role in the development of cognitive impairment induced by CNP via influencing tryptophan metabolites in the host.(2)High tryptophan diet alleviated CNP and impairment cognitive via reshaping gut microbiota composition and increasing IPA in brain.(3)Tryptophan metabolite IPA attenuated impairment cognition induced by CNP via activating Ah R-Nrf2 signaling pathway in PFC and hippocampus to enhance antioxidative response.