Effects And Mechanism Of Clostridium Butyricum On Social Hierarchical Behaviors In Rats

Purposes: Gut microbiota disruption can cause neuronal function disorders and behavior deficits through the gut-brain axis.Clostridium butyricum,as probiotics,has the ability to regulate gut microbiota and alleviate neuronal injury.Social hierarchy is ubiquitous and plastic in the social organization of rats,which can be evaluated by behavioral experiments.The purpose of this study was to explore whether the rats' gut microbiota could influence hosts social hierarchical behaviors,and the role that Clostridium butyricum intervention on the gut microbiota played in the social hierarchical behaviors,then find out the gut-brain axis mediation pattern and the epigenetic mechanism.Methods: The male SD rats were divided into two groups: the dominant group and the subordinate group by offensive behavior test and water competition test.16 S r DNA sequencing was performed on the gut microbiota to analyze the bacterial differences between the two groups.In the following experiments,3-week-old male SD rats were divided into 5 groups: control group,gut microbiota disrupted group,dominant microbiota transplanted group,subordinate microbiota transplanted group,and disrupted microbiota transplanted group.After adolescence,behavioral experiments were conducted to evaluate the differences in social hierarchical behaviors,and 16 S r DNA sequencing was performed to analyze the composition of gut microbiota in each group.Subsequently,the contents of major fecal short-chain fatty acids such as butyric acid were analyzed by GC-MS,and the transcription and protein levels of histone deacetylase HDACs in different brain regions were evaluated by q PCR,western blotting and immunohistochemical methods.Based on butyric acid,the mediating mechanism of C.butyricum in the gut-brain axis were further evaluated.Finally,drug and specific genetic interventions were used to investigate the role of HDAC2 in social hierarchical behaviors.Results: 1)There were significant differences in gut microbiota between the dominant and subordinate groups,which was mainly manifested as different overall structure,and the relative abundance of Firmicutes in the dominant rats was significantly lower than that in the subordinate rats.Disruption of gut microbiota can induce subordinance,and effective microbiota transplantation can restore its dominant status.Intriguingly,the dominance of social hierarchical behaviors was transmitted with microbiome transplantation.The microbiota model based on cornerstone species can effectively influence and accurately predict the specific rank status;2)The contents of major short-chain fatty acids such as butyric acid were positively correlated with the hierarchical status of each group;The intestinal colonization of Clostridium butyricum showed a similar effect to that of butyric acid intervention,which could significantly improve the social status of mice with intestinal bacterial disruption;3)The transcription level and protein level of HDAC2 in rat mPFC region were negatively correlated with dominance status;The intervention TSA,an inhibitor of HDAC2 enzymatic activity,can significantly improve the social rank of rats;4)HDAC2 knockdown in the mPFC region of microbiota disrupted rats significantly improved their dominant status in social rank competition,while hippocampal knockdown did not significantly affect their social hierarchical behavior.Further studies showed that HDAC2-specific knockdown in neurons in the mPFC region had a dominance-enhancing effect,while similar genetic interventions in oligodendrocytes did not have a significant effect on hierarchical behavior.Conclusion: The results of this study indicate that gut microbiota is an important factor affecting social hierarchical behaviors in rats.The gut-brain axis pathway from gut microbiota to the central nervous system,namely intestinal flora-short-chain fatty acid-HDAC2,played an important role in gut bacterial intervention affecting hierarchy behavior,,which is the key pathway to mediate social hierarchical behaviors in rats.Intestinal colonization of the probiotics Clostridium butyricum has a significant repairing effect on the downgrading caused by gut microbiota disruption,which provides a theoretical basis for adjusting the hierarchical behaviors through the intake of probiotics,so as to alleviate the negative effects of different social hierarchy on health.