Experimental Study Of Botulinum Toxin Type A In The Treatment Of Depression By Regulating BDNF-TrkB Signaling Pathway

Purposes1.Explore the molecular mechanism of botulinum toxin type A(BoNT/A)in the treatment of depression in mice;2.To explore the correlation between the Gαil/3 expression level in hippocampus of mice and depression behavior,and the effect of gene knockout Gαi1/3 on the efficacy of botulinum toxin type A(BoNT/A)in treating depression in mice;Methods1.Cultivation of wild type(WT)and Gαi1/3 double knockout mice(DKO)embryonic fibroblasts(MEFs),Gail single knockout MEFs cells,Gai3 single knockout MEFs cells,Gαi2 single knockout MEFs,Gαb1 knockout MEFs cells;2.Targeting knockdown Gail and/or Gai3 in MEFs using shRNA and siRNA methods;3.Reintroduction the plasmids containing the Gαi1 and Gai3 genes into DKO MEFs cells to obtain Resue 1 and Resue3 MEFs;4.Treated the above MEFs cells with botulinum toxin A(BoNT/A)respectively,and then detecte the activation level of the downstream adaptor protein Gabl and Akt-mTORC1 signaling pathway and Erk-MAPK signaling pathway;5.An animal model of depression was established by intraperitoneal injection of corticosterone(CORT)into C57 male mice and Gail/3 gene knockout mice were reared.Then,depression-like behaviors tests were performed in each group of mice.BoNT/A was injected into facial muscles of mice in Gail/3 gene knockout group and the model group.And then measured the depression-like behaviors of the mice in each group again.Result1.In MEFs,Gαi/3 double knock(DKO)blocked the activation of Akt mTOR/ERK-MAPK signaling pathway induced by BoNT/A.2.After BoNT/A stimulation,the MEFs of Gai-shRNA,Gai-siRNA,Gail-KO and Gai3-KO were slightly weakened compared with the wild type(WT).3.In resurel and resue3 MEFs,after stimulation with BoNT/A the downstream signal activation was obviously restored.4.After BoNT/A stimulation,Gab1 knockout MEFs showed a significant decrease or even absence of activation levels of downstream Akt mTORC1 signaling pathway(Akt,S6,S6K)and MAPK/ERK signaling pathway(ERK).5.Gail/3 gene knockout group and the depression model group mice showed depression-like symptoms;And facial muscle injection of BoNT/A could alleviate the depression-like symptoms of depression model group mice,but not the Gai/3 gene knockout group.Conclusion1.BoNT/A can increase the expression level of BDNF in the hippocampus and prefrontal cortex of depression model mice,and can relieve depression by activating BDNF-TrKB signaling pathway.2.Gαi/3 knockout mice have depression-like symptoms,and Gαi/3 knockout inhibited the antidepressant effect of BoNT/A;...