Microglia In Dopaminergic Neurotoxicity Of MPTP

Background:Parkinson’s disease(PD)is the second most common anddebilitating neurodegenerative disorder that is characterizedby progressive loss of dopaminergic neurons andmotor dysfunctions.Among the first responders to neural injury,microgliaengage in intimate cross-talkwith other intrinsic brain cellsand leukocytes that infiltrate the brain from the peripherythrough the compromised blood–brain barrier.Depending timing and disease types,microglia produce varied substancessuch as chemokines,cytokinesand trophicfactors that influenceneuroinflammation and neural activity.However,the precise role of mciroglia in PDremains poorly understood.The survivalof microglia highly depends on colony-stimulating factor 1 receptor(CSF1R)signaling.CSF1 R inhibition using PLX3397 resultsin a nearly complete elimination of microglia from thebrain with no apparent impact on other brain cells such asneurons,astrocytes,or oligodendrocytes.In this study,we used PLX3397 to depletemicroglia,determined the influences of microglia depletionon neuroinflammation and dopaminergic neurotoxicity ofMPTP,and investigated the underlying mechanisms.Methods:We used a mouse PD model induced by MPTP exposure.Prior toMPTP injection,mice were given PLX3397 at a dose of 40 mg/kg/day for 21 days to deplete microglia and PLX3397 treatment was continued until the end of experiments.At day 7 after MPTP injection,evaluation of motor function wasperformed by rotarod test and pole test.Pathological damage of dopaminergic systemwasevaluated by immuno-histochemical staining.Flow cytometry was used to monitor the infiltration of immune cellsincluding T cells,B cells,NK cells,macrophages and neutrophils in the brain,and the activity of astrocytes.Immunofluorescence staining was usedto determine thealterationsof immune cells in the brain.To examinethe effects of microglia depletion on neruoinflammation after MPTP exposure,RT-PCR was used to measure the expression of IL-1β,TNF-α,IL-2,IL-6,IFN-γ and iNOSin the substantia nigra and striatum.In addition,the expression of BDNF was measuredby ELISA.To testwhether microglia directly protect against MPTP-induced neurotoxcity,SH-SY5 Y cells were co-cultured with microglia-like BV2 cells.MPP+-induced cytotoxicitywas measured usingMTT assay.Results:CSF1R inhibition using PLX3397 led to a nearly complete emilination of microglia in mice reciving MPTP exposure.Depletion of microglia exacerbatedthe impairment of locomotor activities and the loss of dopaminergic neurons in MPTP-treated mice.Further,depletion of microglia augmented the production of inflammatory mediators and infiltration of leukocytes in the brain after MPTP exposure.Microglia depletion-induced exacerbation of MPTP neurotoxicity was also seen in lymphocyte-deficient mice.In addition,depletion of microglia did not affect the production of brain-derived neurotrophic factor(BDNF),while dramatically augmented the production of inflammatory mediators by astrocytes after MPTP exposure.In contrast,microglia-like BV2 cells cannot directly protect SH-SY5 Y cells against MPP+-induced cytotoxicity,suggesting that microglia mightnot directly protect dopaminergic cells against MPTP-induced neurotoxicity.Conclusions:Conclusions:Our findings suggest a protective role of microglia against MPTP-induced neuroinflammation and dopaminergic neurotoxicity.