Therapeutic Effect Of GABABR2 Agonists On ASD-like Behavioral Deficits In Mice Prenatally Exposed To Valproic Acid

Part Ⅰ.GABABR2 Agonists STX209 Ameliorates Core ASD-like BehavioralDeficits in Mice Prenatally Exposed to Valproic AcidObjective Autism spectrum disorder(ASD)mice prenatally exposed to valproic acid were administered the GABAB receptor 2(GABAR2)agonist STX209 during early postnatal development.To observe its therapeutic potential on ASD-like behavior in VPA model mice,and to provide experimental basis for clinical treatment of autism.Methods Suitable age C57BL/6 mice were used for mating.The time of conception was determined by observing the vaginal suppository of female mice,and 300 mg/kg(12mg/m L)sodium valproate was intraperitoneally injected on the 10 th and 12 th days after conception,and the offsprings were VPA model mice(the first generation,F1).According to whether control mice and VPA model mice were treated with STX209(0.6 mg/kg,intraperitoneal injection,2 times a day),the offspring mice around 21 days old were divided into 4 groups: normal control group(CTRL)control group group(CTRL + STX209 group),VPA model group(F1-Con group),VPA model drug intervention group(F1 + STX209group).Previous studies have shown that male VPA mice have more stable ASD-like behavioral and pathological features than the female mice,so only male mice were selected for experiments.Meanwhile,in order to avoid the "litter effect",male mice in the litters were randomly and equally divided into the intervention and non-intervention groups.If a single male mouse remains,it is eliminated.In the end,four groups were established,each with 14 mice.VPA mice underwent behavioral testing after 2 weeks of continuous low-dose STX209 administration.In this study,quantifiable ASD core/related diagnostic behavior experiments were selected: open-field task and open-field habituation task,novel object recognition experiment,three-chamber social interaction test,and marble burying test.Continue administration of STX209 while the mice were tested for behavioral experiments.About 60 days after birth,mice were sacrificed and the total length and density of neuronal dendritic spines in their hippocampus(HC),as well as the expression of GABABR2 and glutamate decarboxylase 65/76(GAD65/67)were measured.Results This study investigated the effects of STX209 on VPA model mice via behavioral testing and revealed a significant reversal of core/associated autism-like behavior,including the sociability and preference for social novelty deficits,novelty recognition deficits,locomotion and exploration activity deficits and marble-burying deficits.This may be associated with STX209 correcting dendritic arborization,spine density and GABABR2 expression in hippocampus of VPA model mice.However,expression of glutamic acid decarboxylase 65/67 in the hippocampus were not altered by STX209.Conclusion STX209 administration ameliorated autism-like symptoms in mice exposed to VPA prenatally,suggesting that autism-like symptoms in children with a history of prenatal VPA exposure may also benefit from treatment with the GABABR2 agonist STX209.Part Ⅱ.GABAB Receptor 2 Agonists Baclofen Ameliorates Core ASD-like Behavioral Deficits in Mice Prenatally Exposed to Valproic AcidObjective GABABR2 agonist Baclofen was administered early in prenatal VPA-exposed mice to assess the therapeutic potential of GABABR2 agonist for ASD-like behavior in miceMethods The VPA mouse model was constructed in the same way as above.According to whether the VPA model group was intervened with baclofen,the male offspring mice were divided into three groups: normal control group(CTRL group),VPA model group(F1-Con group)and VPA model drug intervention group(F1 + baclofen)when they arer born around 21 days.The “litter effect” was also excluded as above.There were 14 mice in each of groups.After weaning,the mice in each group were given baclofen in drinking water at a concentration of 0.5mg/m L.After 2 weeks of continuous administration,the mice in each group were tested by behavioral experiment(same as in Part Ⅰ),that were open-field task and open-field habituation task,novel object recognition experiment,three-chamber social interaction test,and marble burying test.During the experiment,baclofen was continued to be given.Results Behavioral experiments showed that chronic administration of baclofen significantly improved core/relevant ASD-like deficits in VPA model mice,including(1)increased the parameters of socail interaction and social novelty preference in three-chamber social interaction test;(2)increased the exploration of novel objects,(3)increased locomotor and exploratory activities in the open-field habituation task test,and(4)increased the number of buried marbles and burying actions.Conclusion Oral administration of the GABAB receptor 2 agonist baclofen ameliorated ASD-like core/relevant symptoms in mice prenatally exposed to VPA.Part Ⅲ.Prenatal GABAB Receptor Agonist Administration Corrects the Inheritance of Autism-like Core Behaviors in Offspring of Mice Prenatally Exposed to Valproic AcidObjective The offspring of littermates of VPA model mice still exhibit ASD-like behaviors.We treated VPA model mice with baclofen during mating and pregnancy to evaluate the therapeutic potential of the GABABR2 agonist baclofen on the inheritance of ASD-like behaviors in VPA model mice.Methods The age-appropriate VPA model mice(F1)in the same litter were used for mating,and baclofen(0.5 mg/m L)was / was not administered orally to F1 male and female mice three days before mating,the female mice continued to be / be not given baclofen until delivery,and the offsprings were the second-generation mice(F2).Based on prenatal use of baclofen,the offsprings were divided into two groups: second-generation control group(F2-Con group)and second-generation baclofen intervention group(F2-Int group).The male offsprings were selected for the experiment,and the “litter effect” was also excluded as above.Five weeks after the birth of F2 mice,the mice in F2-Con group and F2-Int group were tested through behavioral experiment(same as the Part Ⅰ and Part Ⅱ,namely open-field task and open-field habituation task,novel object recognition experiment,three-chamber social interaction test,and marble burying test).Finally,when comparing the differences between groups,the mouse data of CTRL group in Part Ⅱ were added,and the three groups were analyzed and compared.Results Prenatal administration of baclofen can significantly improve the inheritance of ASD-like behavior in VPA model mice.Compared with F2-Con group mice,ASD-like behavior in F2-Int group mice was significantly improved.The results were similar to those in F1 mice and included the following changes: prenatal baclofen treatment in the F2 generation(1)increased the social time and index,(2)increased the exploration of new objects,(3)increased locomotor and exploratory activities in the open-field test,and(4)increased the number of buried marbles and burying actions.It is possible that prenatal administration of baclofen corrected the total/mature dendritic spines density of pyramidal neurons in the hippocampus(HC)and medial prefrontal cortex(m PFC)of F2 mice,normalizing their synaptic plasticity.Conclusion Prenatal administration of baclofen can significantly corrected the inheritance of core ASD-like behavior and neuropathological defects in VPA model mice,suggesting that GABABR2 agonist intervention in early life may block the inheritance of ASD-like behavior in clinical patients,which has some implications for the treatment and prevention of ASD.