CUMS-induced Depression Increased Susccptibility In An MPTP Mouse Model Of Parkinson’s Disease Through P2X7R Mediated Inflammation

Objectives:This study aimed to induce depression by this method to investigate whether depression increased the susceptibility in an MPTP PD mouse model and the increase in this susceptibility whether based on the effects of P2X7R promoting inflammation.Materials and methods:Six-week-old C57BL/6 male mice were used to induce depression-like symptoms after 14 days of CUMS random stress,and PD mouse model were induced by MPTP(20mg/kg,4 times/day,ip).The depression-like behavior of mice was tested by tail suspension experiment and sucrose preference test,and the motor function of mice was tested by rotating rod test and open field test.CUMS mice were given Brilliant Blue G(BBG),a P2X7R inhibitor,before MPTP.Behavioral changes of depression and motor function of CUMS mice after inhibiting P2X7R were observed.Immunofluorescence technique was used to detect the co-localization of P2X7R and Caspase-1 in microglia of the substantia nigra and the morphology of microglia.The expression of tyrosine hydroxylase(TH)positive dopaminergic neurons was detected by immunohistochemistry.The expression levels of P2X7R protein and inflammatory related proteins NLRP3 and Caspase-1 were detected by Western Blot,and the level of inflammatory factor IL-1beta were detected by ELISA.Results:The sucrose preference test showed that the preference of CUMS-induced mice to sucrose solution decreased(P<0.01),while MPTP mice unchanged.The tail suspension test showed that the immobility time of CUMS mice and MPTP mice increased(P<0.001).Rotating rod test showed that CUMS mice had no difference in rod time compared with Sham group,while acute MPTP mice had lower rod time(P<0.001).CUMS+MPTP group aggravated the dyskinesia than MPTP group(P<0.05),which was reversed by BBG administration and relieved the depressive behavior induced by CUMS(P<0.001).Immunohistochemical results showed that TH-positive neurons in CUMS+MPTP group lost more severely than those in MPTP group(P<0.05),which could be alleviated by BBG(P<0.001).Immunofluorescence images showed that P2X7R and Caspase-1 were co-localized in microglia of substantia nigra.Microglia in CUMS mice were activated and the expression of mature caspase-1 in microglia was increased.Western Blot results showed that the expression of P2X7R in CUMS group(P<0.001)and CUMS+MPTP group(P<0.05)was significantly higher than Sham group.The expression of P2X7R in CUMS+BBG+MPTP group was significantly lower than CUMS+MPTP group(P<0.05).Similarly,the expression of NLRP3 and Caspase-1 in CUMS group(P<0.01)and CUMS+MPTP group(P<0.05)increased significantly,while the expression of NLRP3 and Caspase-1 in CUMS+BBG+MPTP group decreased significantly compared with CUMS+MPTP group(P<0.001).The expression of IL-1beta in ELISA was similar to that in WB(P<0.05).These results suggested that CUMS aggravates the dyskinesia of MPTP model,promotes the activation of microglia in substantia nigra,increased the level of inflammation and the death of DA neurons,which can be reversed by BBG.Conclusion:Depression induced by CUMS increased the susceptibility in an MPTP mouse model,which is based on the activation of the inflammatory pathway mediated by microglia P2X7R.