Alterations Of Synapse And Its Association With Neuroinflammation In The Offspring Of Maternal Immune Activation Rat

BackgroundMaternal immune activation(MIA)during gestation can increase later risk of schizophrenia(SZ)in adult offspring.It is believed that schizophrenia is a kind of neurodevelopmental disease.As synapses change along development,its role in the pathogenesis of schizophrenia has caught peoples’ attention.There are evidences which suggested that alterations in synaptic structure and function are central to the process of schizophrenia.Besides,neuroinflammation may have some connection with the expression of synaptic proteins and mediate the high risk of MIA offspring.Post-mortem brain studies have found alterations of neuro-immune system in patients with schizophrenia.However,little is known about dynamic changes of cerebral inflammation,synapse and behavioral performances along with the disease course.ObjectivesPregnant rats were administrated with polyinsinic acid-polycytidylic acid(Poly(I:C))in early gestation to lead to maternal immune activation and to establish the rat model of schizophrenia.Using this model,we observed the dynamic development of behavior,astrocytes and microglial cells in the brain,the levels of inflammatory cytokines(TNF-α,IL-6,IL-1β)as well as synaptic associated protein expression.In this study,different periods were analysed in the aspect of synaptic protein and the immune system as well as the association between them.Methods1.Specific pathogen free(SPF)Sprague-Dawley adult male and female rats were randomly selected at 9 weeks old for breeding.2.Female rats which were successfully pregnant were randomly divided into the Poly(I:C)model group(n=9)and the Vehicle control group(n=9).On gestational day9(GD9),two groups were respectively intravenous injected of 10mg/kg poly(I:C)and the same volume of 0.9% sterile saline.After the injection,the pregnant rats immediately returned to their former cages,waiting for 3 hours,and three rats from each group were randomly selected for quality control.The other rats lived under no interference until they produced calves.3.On postnatal day 21(PND21),the offsprings from both groups were weaned and housed three to four in one cage,some pups of each group remained undisturbed until peridolescence(PND40),the rest pups of each group were grew up to young adulthood(PND60),which respectively equal to puberty and young adulthood in human.4.The prepulse inhibition tests(PPI)was conducted in the two experimental groups at puberty and young adulthood.5.Brain immune alterations were correspondingly determined in the prefrontal cortex(PFC)and hippocampus(HC)at both time points.The status of microglia and astrocyte were detected through immunohistochemical staining of ionized calcium-binding adapter molecule 1(Iba1)and glial fibrillary acidic protein(GFAP).And levels of IL-6,IL-1β,TNF-α from both brain regions were evaluated by enzyme linked immuno-sorbent assay(ELISA).Western blot was used to evaluate the expression of synapse-related proteins.6.All of the data was presented with means±standard deviations.Different statistical methods were chosen according to different data types,such as,one-way analysis of variance(ANOVA)as well as independent-sample T test.SPSS 20.0 software system was adopted for data analysis.Results1.The levels of IL-1β,IL-6,and TNF-α in the plasma of successfully pregnant dams administrated with Poly(I:C)were significantly increased when comparing with the controlrats after injecting for 3 hours(p<0.01).2.MIA had no significant difference in all dB groups(70,75 and 80dB)at adolescence.PPI defects manifested in all dB groups(70,75 and 80dB)at adult offsprings of dams exposed to Poly(I:C)at GD9.3.As the results from WB manifested,the level of synapsin was elevated at PND40 in the PFC of the Poly(I:C)group.While the expression of PSD95 was increased in the PFC and HC of the Poly(I:C)group.While the expression of PSD95 was increased merely in PND60 of both targeted brain areas.The synaptic skeleton protein NFM and NFH decreased in the hippocampus and the puberty frontal lobes in puberty,while the p-cofilin that regulates the synaptic function increased in both targeted brain areas.4.Iba1-positive cells numbers was analyzed at two different time points and in the prefrontal cortex and hippocampus of the two groups(×100).Both in the PFC and the HC,more activated microglia were observed in Poly(I:C)offspring at the age of either 40 or 60 days old than the controls.The results clearly suggested increased number of microglia at both examined regions in Poly(I:C)offspring;moreover,the morphology of microglia in the two target regions was also apparently different from the vehicle offspring.In detail,activated microglias in offspring of MIA rats were characterized by enlarged cell bodies,and the processes became retracted and thickened which differed from the quiescent state in vehicle offspring with round cell body and thin processes as well as simple ramifications.5.While on PND40,astrocyte had no significant difference between two groups;on PND60,we can see elevated GFAP-positive astrocytes and hypertrophic astrocyte morphology in offspring of MIA rats.6.It suggested that the Poly(I:C)pubescent offspring showed increased concentrations of IL-1β and IL-6 in both PFC and HC compared with the controls.As for young adult offspring,no significant difference can be found in the HC,while in the PFC,the levels of TNF-α and IL-6 increased.Additionally,the effects of group treatment did not exist in theconcentration of TNF-α in the HC at two stages.7.There was a negative correlation between IL-6 and NFM protein expression in adult hippocampus,while TNF-α was positively correlated with PSD95 level.The density of astrocytes in puberty hippocampus was negatively correlated with the expression level of PSD95.ConclusionsThere are alterations of several synaptic protein levels and neuroinflammation in the early maternal immune activation rat generation before the manifestation of schizophrenia like symptoms,and there is a certain correlation between the synapse damage and the neuritis.Our research suggested that adolescence may be a more critical time point for disease intervention and treatment than adulthood.