| Background Autism spectrum disorder（ASD）is a neurobiological-based developmental disorder in which social impairment and repetitive-stereotyped behavior are its core symptoms.Autism is often accompanied by a variety of diseases,such as cognitive impairment,emotional impairment,and learning and memory impairment.Abnormal synaptic structure and function is an important pathogenesis of autism.Exercise rehabilitation training can improve abnormal behavior of ASD.However,few studies have been conducted on the role of exercise in ASD synaptic remodeling,and the specific molecular mechanisms remain unclear.PurposeThe purpose of this study was to investigate the effects of 4-week moderate-intensity running wheel exercise combined with swimming on social behavior,repetitivestereotyped behavior,anxiety,short-term working memory,cognitive ability in valproic acid（VPA）-induced ASD model rats,and further explore whether these effects are related to the synaptic structural plasticity of the medial prefrontal cortex and the BDNFTrk B-Cdc42 signaling pathway in the medial prefrontal cortex,in order to provide a new experimental basis for the clinical formulation and implementation of exercise rehabilitation treatment plans for ASD patients and theoretical support.MethodsIn this experiment,Sprague-Dawley rats were injected intraperitoneally with a dose of300 mg/kg of valproic acid at two-time points on the 10 th and 12 th day of pregnancy.Pregnant mice with equal doses of normal saline were used as normal control rats to obtain male offspring after giving birth.4-5 week old normal control rats and ASD model rats were randomly divided into four groups: control group（NS）,control exercise group（NSE）,ASD model group（ASD）,ASD exercise group（ASDE）,7 in each group.The NSE group and the ASDE group underwent one-week adaptive exercise training,followed by formal exercise training: running training 30 min each time;swimming training 20 min each time.Exercise 5 days a week,2 days off,for 4 weeks.No intervention was given to the NS group and the ASD group.After the behavioral test（open field test and repetitive behavior test,Y-maze test,three-box social test,and novel object recognition test）at the end of the last training period,3 rats were randomly selected from each group for perfusion fixation,and their brains were removed.The prelimbic cortex（Pr L）area and the infralimbic cortex（IL）area in the medial prefrontal cortex were separated,and the synaptic ultrastructure of each group was observed and imaged by electron transmission microscopy.Three rats were randomly selected from each group to isolate fresh medial prefrontal cortex,and western blotting was used to detect the protein expression changes of BDNF,Trk B,phosphorylated Trk B and total Cdc42 in the medial prefrontal cortex of rats in each group.Results（1）The results of behavioral testing showed that compared with the NS group,the ASD model rats had obvious repetitive behaviors（significantly increased self-grooming time,P<0.05）,anxiety state（active time in the central area of the open field,entering The number of times and the central activity distance were significantly reduced,P<0.05）,social behavior disorder（decreased interest in exploring unfamiliar rats,P<0.05）and cognitive impairment（the exploration preference for new objects was significantly decreased,P<0.05）,while there was no significant difference in short-term working memory ability and activity ability（P>0.05）;after 4 weeks of running wheel exercise combined with swimming exercise intervention,compared with the ASD group,the anxiety state,repetitive-stereotyped behavior of the rats in the ASDE group,social impairment and cognitive impairment were significantly improved（P<0.05）.（2）The results of electron transmission microscopy showed that compared with the NS group,the synaptic density in the ASD group showed an opposite trend in the Pr L and IL subregions: Pr L was significantly increased while IL was significantly decreased,which were significantly reversed after exercise intervention.Further analysis of synaptic morphological changes found that in the Pr L area,compared with the NS group,the ASD group had significantly increased Flat-type and Frown-type synapses（P<0.01）.After exercise intervention,compared with the ASD group,the number of Frown-type synapses decreased significantly in the ASDE group（P<0.05）,while the Flat type had a downward trend but no significant difference,and there was no significant difference between the four groups of Smile type synapses.In the IL area,compared with the NS group,the Smile-type synapses in the ASD group were significantly decreased（P<0.05）;after exercise intervention,the number of Smile-type synapses in the ASDE group was significantly increased compared with the ASD group（P<0.01）.However,there was no difference between the four groups of Flat-type synapses and Frown-type synapses in the IL region.Further analysis of the synaptic ultrastructure in each group showed that in the Pr L region,compared with the NS group,the width of the synaptic cleft in the ASD group was significantly reduced and the thickness of the postsynaptic denser was significantly thickened（P<0.05）.Compared with the ASD group,the width of the synaptic cleft in the ASDE group increased（P<0.05）and the thickness of the postsynaptic denser was significantly reduced（P<0.01）,but the length of the synaptic active area and the synaptic curvature showed no significant difference among the four groups.In the IL area,compared with the NS group,the width of the synaptic cleft in the ASD group was significantly reduced and the length of the synaptic active area increased（P<0.05）,After exercise intervention,the width of synaptic cleft increased and the length of synaptic active area decreased in the ASDE group compared with the ASD group（P<0.05）,but there was no significant difference in PSD thickness and synaptic curvature among the groups in the IL area.（3）Western blotting results showed that: the expression of BDNF and Cdc42 in the medial prefrontal cortex of the ASD group was lower than that of the other groups,but there was no significant difference;After exercise training,the expression of BDNF and Cdc42 in the medial prefrontal cortex in ASDE group was slightly increased compared with other groups,but there was no significant difference.The results of protein expression of Trk B and phosphorylated Trk B（P-Trk B）showed that the ASD group was higher than other groups,but there was no significant difference between the groups.ConclusionExercise training can effectively improve social impairment,repetitive stereotyped behavior,anxiety and cognitive ability in ASD rats.The neurostructural basis for this improved effect may be the differential density and morphological plastic changes of synaptic ultrastructure in the Pr L and IL subregions of the medial prefrontal cortex,and its molecular basis may not be significantly related to the BDNF-Trk B-CDC42 signaling pathway in the medial prefrontal cortex. |
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