The Protective Effect And Molecular Mechanism Of Folic Acid On Autism Model Induced By Valproic Acid In Rats

Autism spectrum disorder(ASD)is a neurodevelopmental disorder that occurs in early childhood and can be diagnosed before the age of 3.It is a lifelong disease influenced by genetic and environmental factors.The disease incidence is about 1.5%with an increasing trend,the incidence has a significant gender difference,the ratio between male and female is about 4:1.The core characteristics of autism are abnormal social communication abilities and increased repetitive stereotypes accompanied by other behavioral impairments and dysfunctions,which cause heavy physical and psychological burdens on families and society.The pathogenesis of autism is complicated and remains unclear,current research indicates that the pathogenesis of autism is associated with a variety of molecular and biological changes,including the change of synaptic-associated proteins expression and synaptic structural plasticity,in which the changes of excitatory synaptic and inhibitory synaptic markers and dendritic spine density play an important role,the cellular signaling pathways involved are MAPK signal pathway and AKT signal pathway.Valproic acid(VPA),a drug for the treatment of epilepsy and bipolar disorder,exerts neuroprotective effects by reducing the excitability of neurons.VPA exposure during pregnancy increases the risk of autism in offspring with abnormal behaviors,such as delayed growth and development,increased repetitive stereotypes,and social communication disorders.Currently,research has successfully established an animal model of autism by intraperitoneal injection of VPA in rodents during pregnancy,which has good surface validity,construct validity and predictive validity and provides an important basis for better understanding of the pathogenesis and prevention of autism.Folic acid(FA)is a water-soluble B vitamin.Tetrahydrofolate is the active form and it is an important methyl donor involved in metabolic regulation in the body.Epidemiological studies have shown that folic acid supplementation in early pregnancy is neuroprotective.It has the ability to reduce the probability of autism in offspring,but the mechanism is still not clear.This study will focus on the protective effects and related mechanisms of folic acid on VPA-induced autism rat modelsIn this study,rat model of autism was established by exposing rats to VPA during pregnancy,folic acid group was continuously orally administered with folic acid(1 mg/kg,light folic acid,4 mg/kg,high folic acid)from Eld to E12.5d,we explored the protective effect of folic acid on autism and its molecular mechanism,providing theoretical and experimental basis for the prevention of autism.We assessed growth and development level,exercise capacity and anxiety level,and social interaction ability of 5w male offspring by growth and development test,the open field test(OFT)and the three-chamber test.The effective dose of the folic acid group was screened for subsequent molecular experiments.We detected the dendritic spine density in the medial prefrontal cortex(mPFC)by Golgi staining.We used Western Blot to quantify the excitatory synaptic and inhibitory synaptic marker proteins,BDNF,and MAPK and AKT signaling pathways protein expression on the orbital frontal cortex(OFC),medial prefrontal cortex(mPFC)and hippocampus(HP).Results:1.Established VPA-induced rat autism model.Compared with the control group,the model group showed significant delayed growth and development ability,increased anxiety level and abnormal social communication ability,this shows the successful establishment of an animal model of autism.2.Folic acid has protective effect on autism.Compared with the model group,the folic acid groups had different degrees of reversal of behavior,and the high-dose folic acid(4mg/kg)group had a more obvious reversal effect.The growth and development test showed that the VPA+HFA group significantly reversed the delayed growth development compared with VPA group;the results of the open field experiment showed that movement including time and frequency of the VPA+HFA group in the central area of the field were significantly higher than that of the model group(P<0.05),demonstrating a significant reversal in the increased anxiety level in the model group.The results of the three-chamber test showed that the social index(SI)and social preference index(SPI)of the VPA+HFA group were significantly higher than the model group(P<0.05),significantly reversing the abnormal social communication in autism observed in the model group.3.Folic acid alters synaptic plasticity in autism rat model.The density of dendritic spines in the mPFC-PrL region of the model group increased significantly(P<0.00/).Compared with the model group,the density of dendritic spines in the VPA+HFA group was significantly lower(P<0.001),which might be associated with the change in BDNF expression.4.Folic acid alters the expression of synaptic markers in autism rat model.The expression of excitatory synaptic marker CaMKII protein in the frontal cortex(OFC)increased significantly(P<0.001),and the expression of inhibitory synaptic protein marker GAD65 protein in hippocampus(HP)decreased significantly(P<0.05),compared with the VPA group,these effects in the VPA+HFA group were reversed.For the phosphorylation level of ERK1/2 and AKT proteins,there was no significant change in each brain region.Conclusion:Maternal folic acid supplementation in early pregnancy can significantly improve the autistic behavior of rodents obtained from VPA-induced autism models,and its related neurobiological mechanisms may be related to the expression of BDNF and the abnormal development of excitatory synapses and inhibitory synapses.