Study On Behavioral Abnormalities And Regulatory Mechanisms Of Hypothalamus In Catch-up Growth IUGR Rats

Objective: CG-IUGR fetuses have a significantly increased risk of developing metabolic problems and cardiovascular diseases in the long term,but research on neurodevelopment was limited and focused mainly on cognitive and learning abilities.In order to clarify whether CG-IUGR individuals exhibit abnormalities in emotional and social behavior,this study constructed a CG-IUGR rat model to explore the types of behavioral abnormalities that occurred during development and the underling mechanisms,providing support for future prevention and treatment.Methods: The IUGR rat model was constructed by maternal nutrient restriction,and the CG-IUGR rat model was constructed by limiting litter amount.Male CG-IUGR and normal individuals were selected and their weight were recorded and plotted.At representative development stages of 4th(juvenile),6th(adolescent),9th(sexual maturity),12th(early adulthood),and 16 th week(adulthood),further behavioral examinations were conducted.The weight of food consumed continuously over 72 hours was monitored and the relative expression levels of hypothalamic appetite-regulating neuropeptides were quantified by RTq PCR and Western blot to reflect the feeding behavior of CG-IUGR rats.The marble burying test,open field test,three-chamber test,and forced swimming test were used to determine whether CG-IUGR male individuals exhibited repetitive stereotyped behavior,anxiety-like behavior,social impairments,and depressive-like behaviors.The brain weight/body weight ratio of CTRL and CG-IUGR rats at each age was compared.And RTq PCR was used to measure the genes expression levels of Nestin(marker of neural stem cell),Bax/Bcl2(marker of apoptosis),Notch1(marker of neuron proliferation and differentiation),NRP2 and NCAM1(marker of synapse plasticity),and MBP(marker of myelin sheath formation)in the hypothalamus of rats at different stages to confirm the neurodevelopmental status.Primary hypothalamic cells were extracted from neonatal rats,and immunofluorescence staining was used to evaluate the neurogenesis of the hypothalamus in IUGR fetuses.RT-q PCR was used to detect the changes in gene expression levels of MAGEL2,OXT,and OXTR in the hypothalamus of rats,and serum ELISA was used to detect the level of oxytocin in serum.Both of them were used to confirm whether the MAGEL-OXT pathway was expressed normally in hypothalamus of CG-IUGR rats.RTq PCR was used to detect changes in the gene expression levels of methylation,deacetylation,the WASH complex and their regulatory complexes downstream of MAGEL2 in the hypothalamus,to confirm whether these processes were related to the abnormal behavioral performances of CG-IUGR rats,Results: The increase in food intake in CG-IUGR rats was only observed during the early postnatal period(around 4th week),which coincides with the main period of catch-up growth under conditions of abundant nutrition.Repetitive stereotyped behaviors,social novelty preference deficits,and depressive-like behaviors persisted from infancy to adulthood in CG-IUGR rats,and anxiety-like behaviors were only observed around the adolescent period.What’s more,the decline in motor ability was not observed.CG-IUGR rats showed persistent developmental delay in the hypothalamus,including delayed neuronal proliferation and differentiation,decreased synaptic plasticity,and impaired myelination,which parallel their persistent abnormal behavioral patterns.The behavioral abnormalities in CG-IUGR rats before 4th week may also be associated with the overall downregulation of the MAGEL2-OXT pathway.While the expression levels of the MAGEL2-OXT pathway were normalized after 4th week,the behavioral abnormalities still persisted.The possible mechanisms may be related to the dysfunctional distribution and activity of OXTR,or long-term effects of the decrease in m RNA expression of MAGEL2 in the short term before 4th week.The persistent developmental delay in the hypothalamus and the overall downregulation of the MAGEL2-OXT pathway before 4th week in CGIUGR rats may be related to the methylation regulated by DNMT and TET enzymes,rather than to the abnormal function of the deacetylation or the dysfunctional downstream WASH complex of MAGEL2.Conclusions: CG-IUGR rats exhibited persistent repetitive stereotyped behaviors,social novelty preference deficits,and depressive-like behaviors from infancy to adulthood,accompanied by increased food intake and appetite before 4th week and anxiety-like behaviors around the adolescent period.These abnormal behavioral patterns correspond to the persistent hypothalamic neural developmental delay that has lasted from birth to 16 th week.And the abnormal behavioral patterns before 4th week may also be associated with the overall downregulation of the MAGEL2-OXT pathway in the hypothalamus.The developmental delay of the hypothalamus and the overall downregulation of the MAGEL2-OXT pathway in CG-IUGR rats before 4th week may be related to changes in the methylation process during development,rather than to the deacetylation process or the formation of the downstream WASH complex of MAGEL2.