| The stage of fetal development is a critical period that affects an individual’s lifelong health.However,fetuses are susceptible to internal or external stimuli and cause long-term health hazards.Pregnant women who are premature in many countries are given synthetic glucocorticoid(s GC)medications,such as dexamethasone,between 24 and 33 weeks of pregnancy to promote fetal lung maturation and reduce respiratory distress syndrome or the risk of other diseases related to preterm birth.However,in recent years,more and more evidence shows that the use of multiple courses of s GC in the second and third trimester will bring a series of adverse reactions.Although there is currently no clinical data showing that fetuses treated with exogenous glucocorticoids in utero will develop hypothalamic-pituitary-adrenal(HPA)axis stress sensitivity,but there are animals’ experiments show that intrauterine dexamethasone exposure can lead to changes in HPA axis stress sensitivity of the offspring after birth.In multiple animal model studies,dexamethasone exposure during the second and third trimesters will cause the offspring to have significantly increased HPA axis basal activity and stress sensitivity during adolescence or adulthood.The HPA axis plays a vital role in the regulation of cardiovascular,metabolic,reproductive and nervous systems,and its imbalance in regulation can lead to some chronic diseases,such as metabolic syndrome and some mental diseases.Epidemiological investigations have found that the use of dexamethasone during pregnancy can cause intrauterine growth retardation(IUGR),which is related to the incidence of epilepsy after birth.According to the follow-up of 5,632 neonates who were treated with dexamethasone before delivery,it was found that these populations had a significantly increased risk of convulsions during childhood,and men were higher than women.The results of the above studies suggest that prenatal dexamethasone exposure(PDE)during pregnancy may cause increased sensitivity to HPA axis stress and epilepsy susceptibility of the offspring after birth,but the specific mechanism is not yet clear.The paraventricular nucleus(PVN)is the direct control site of the HPA axis.There is a class of neuroendocrine small cells which synthetic and secretable arginine vasopressin(AVP)and corticotropin-releasing hormone(CRH)in the PVN area.The hypothalamic PVN area is regulated by the higher center.The hippocampus is the high negative feedback control center of the HPA axis,which can promote the hyperactive HPA axis to return to normal levels under stress.There are two important neurotransmitters in the mammal’s brain:excitatory glutamate(Glu)and inhibitory gamma-aminobutyric acid(GABA),the dynamic balance of the two is maintained Brain regions(including hippocampus)play an important role in functional activities.When the body is stimulated by stress,the Glu energy neurons projected on the PVN area release Glu to excite CRH neurons,thereby activating the HPA axis.The negative feedback regulation of hippocampus after the increase of GC promotes the release of GABA from the synaptic terminals of GABAergic neurons projected into the PVN area of the hypothalamus to inhibit CRH neurons to avoid excessive excitation of the HPA axis.Glutamate decarboxylase(GAD)can convert Glu to GABA.It is the rate-limiting enzyme for GABA synthesis and plays a key role in regulating the balance between Glu and GABA.At present,it is believed that the target of HPA axis programming of the offspring caused by s GC treatment during pregnancy may be located in the hippocampus,hypothalamus,and pituitary,but the deep mechanism has not yet been clarified.It is currently believed that the mechanism of epilepsy is related to oxidative stress,inflammation,excitatory damage to neurons,and damage to glial cells.Among them,the excitatory injury of neurons caused by excessive discharge of hippocampal Glu energy neurons is considered to be one of the exact mechanisms of epilepsy.One of the reasons for the excessive excitement of Glu-neurons is that GABA-inhibitory signals cannot exert normal inhibition levels.Type 2 K+–Cl-cotransporter(KCC2)is a chloride ion transporter specifically expressed on the cell membrane of neurons.KCC2 couples outward with a potassium ion gradient to transport chloride ions outward from the cell against its electrochemical gradient.The expression of KCC2 in mature neurons is a necessary prerequisite for maintaining the hyperpolarized GABAA receptor current(inhibitory effect).In recent years,people have gained a deeper understanding of the important role of abnormal KCC2 expression in epilepsy.In clinical cases and animal models,there is strong evidence that the inhibition of hippocampal KCC2 expression can increase the frequency or intensity of seizures,which fully shows that the abnormal expression of KCC2 is closely related to the occurrence of epilepsy.However,it is not clear whether PDE may cause progeny susceptibility to epilepsy by inhibiting the expression of KCC2 in the fetal hippocampus.Epigenetic modification has received increasing attention in the field of developmental programming.The process of epigenetic modification shows that environmental stimulation(endogenous or exogenous)of an organism can change its gene expression.Over the past decade,it has been demonstrated that epigenetic processes involve fetal programming of the nervous,endocrine,cardiovascular,and metabolic systems.For example,a series of HPA axis regulatory genes are affected by DNA methylation and histone acetylation.Experiments on humans and other animals have shown that exogenous GC given to the mother during pregnancy can cause epigenetic modifications of the offspring DNA,and these modifications are related to long-term programming changes in gene expression.The changes of epigenetic modification caused by early exogenous GC are very different from the changes of epigenetic modification caused by the natural increase of cortisol in the later period.Without intervention,the effects of early epigenetic modifications are likely to be maintained for life,which is widely accepted.In this study,we first observed the HPA axis high stress sensitivity and epilepsy susceptibility of PDE offspring rats,and paid attention to whether the HPA axis high stress sensitivity changes were transgenerational inherited from maternal line.Further,we explored the mechanism of intrauterine programming of HPA axis high stress sensitivity and epilepsy susceptibility caused by PDE.This study provides an important theoretical basis and experimental basis for elucidating the potential mechanism of long-term neurodevelopmental toxicity of PDE offspring,exploring early intervention strategies,and guiding the rational use of drugs during pregnancy to improve population quality.PART ONEPDE-induced HPA axis high stress sensitivity phenomenon and transgenerational genetic effectObjective: To observe the phenomenon of high stress sensitivity of HPA axis and potential increase of hypothalamic excitability of PDE-induced offspring after birth in PDE-induced offspring and its transgenerationalgenetic effects,Further,we explore the intrauterine origin of increased hypothalamic excitability.Methods: The pregnant rats were divided into control and PDE groups on 9 days of gestational day(GD9).They were treated with 0.2 mg/kg of dexamethasone every day on GD9-20 days.Partially(at least 12 rats in each group)GD20 rats were subjected to isoflurane anesthesia,and fetal rats were harvested by laparotomy,and the fetal hypothalamus was quickly removed under the microscope.Some rats were randomly divided into two groups(ice swimming group and control group).The ice-water swimming group was given 5 minutes of ice-water(5-7℃)swimming every day for 2 weeks.The last swim was performed with ether anesthesia after 1 hour,and the head was killed and blood was taken to remove the hypothalamus.Detection of adrenocorticotrophic hormone(ACTH)level by radioimmunoenzyme-linked method,detection of blood corticosterone(CORT)level by ELISA method,and real-time quantitative PCR detection m RNA expression of corticotrophin(CRH)in hypothalamus,Arginine vasopressin(AVP),vesicular glutamate transporter 2(VGlu T2),L-glutamic acid decarboxylase 65(GAD65),post-synaptic density 95(PSD95),Reelin,and Ca2+/ calmodulin-dependent protein kinase II-α(α-Ca MKII).The remaining part of the PDE group female pups and normal male rats were caged 2:1 to obtain F2 generation pups.F2 generation female offspring were caged with normal male rats at 12 weeks after birth to obtain F3 generation offspring.F2 and F3 generation female pups were given the same ice water swimming stimulation as the F1 generation in 10-12 weeks.After the last ice water swimming,the materials were tested and the same indicators as the F1 generation.Results:(1)Weight and IUGR rate: The birth weight of female and male offspring of F1 PDE group(0.2 mg/kg·d)was significantly lower than that of control group(P<0.01),and IUGR rate of offspring of PDE group was significantly higher In the control group(P<0.01).(2)Change of HPA axis stress sensitivity of F1 generation adult pups: In the non-chronic stress group,the expression of CRH in hypothalamus of male pups in PDE group was lower than that of control group(P<0.01),but there was no significant change in females.The expression of AVP in the thalamus was significantly lower than that in the control group(P<0.01),the serum ACTH concentration in the offspring did not change significantly,and the blood CORT concentration was higher than that in the control group(P<0.01);while in the chronic stress group,the females in the PDE group The expressions of CRH and AVP in the hypothalamus of male offspring were higher than those in the control group(P<0.01),and the concentrations of serum ACTH and CORT were higher than those in the control group(P<0.05,P<0.01).(3)Changes in hypothalamic excitability of F1 generation adult pups: In the non-chronic stress group,compared with the control group,the expression of PSD95 in the hypothalamic excitatory functional protein of female pups in the PDE group increased while the expression of v Glu T2 decreased(P<0.01).The expression of sexual functional proteins Reelin and GAD65 decreased(P<0.01),while the expression ratio of v Glu T2/GAD65 increased(P<0.01),while the expression of PSD95 and GAD65 in the hypothalamus of male offspring in the PDE group decreased(P<0.01),v Glu T2/GAD65 The expression ratio increased(P<0.01);in the chronic stress group,compared with the control group,the expression of PSD95,v Glu T2,GAD65 and v Glu T2/GAD65 in the hypothalamus of female pups in the PDE group increased(P<0.05,P<0.01),the expression ratios of PSD95,v Glu T2 and v Glu T2/GAD65 in the hypothalamus of hippocampus of male pups in the PDE group increased(P<0.05,P<0.01).(4)The change of HPA axis activity in F2/F3 generation female offspring after chronic stress: There was no significant change in the expression of CRH and AVP in the hypothalamus of F2 generation PDE female offspring after chronic stress,and there was no significant change in serum ACTH and CORT concentration;and The expression of CRH and AVP in hypothalamus of female offspring in F3 generation PDE group after chronic stress was higher than that in control group(P<0.05,P<0.01).Serum ACTH concentration did not change significantly in non-stress group but in chronic stress group Higher than the control group(P<0.05),the blood CORT concentration did not change significantly in the non-stress and stress groups.(5)Changes in hypothalamic excitability after chronic stress in F2/F3 female adult pups: After F2 generation of chronic stress,the expression of hypothalamic excitability,inhibitory functional protein expression and v Glu T2/GAD65 expression in female PDE group were not significant change.After chronic stress in the F3 generation,the female offspring of the PDE group only had lower m RNA expression of the functional protein Reelin than the control group(P<0.05),other protein expression did not change significantly,and the expression ratio of v Glu T2/GAD65 was higher than that of the control group(P<0.05).(6)Changes in the potential excitability of hypothalamus in F1 generation fetuses: Compared with the control group,the expression of PSD95 and α-Ca MKII in the hypothalamus of female and male fetal rats in the PDE group was reduced(P<0.01),and the expression of v Glu T2 in the hypothalamus of the male fetal rats in the PDE group was reduced(P<0.01);the expression of the hypothalamic inhibitory gene GAD65 in the PDE female and male fetal rats was significantly reduced(P<0.01),and the expression of Reelin was not significantly changed;the expression ratio of v Glu T2/GAD65 in the hypothalamus of the female and male fetal rats in the PDE group was significantly increased(P<0.01),and the potential excitability of the hypothalamus was increased.Conclusion: PDE can cause high stress sensitivity of HPA axis in F1 generation of adult rats,and can be reproduced with F3 female rats with the maternal line,that is,there is a transgenerational genetic effect,but HPA axis stress sensitivity of F2 female rats has no significant change.It was further found that the F1/F3 generation of adult female rats had increased potential excitability in the thalamus and originated in the F1 generation.It is suggested that the potential increase in excitability of the hypothalamus mediates the occurrence of high stress sensitivity of the HPA axis in adult PDE offspring,which has an intrauterine origin.PART TWOHippocampal programming mechanism of high stress sensitivity of HPA axis of progeny rats induced by PDEObjective: Using animal experiments to explore the effects of PDE on the expression of GAD67,C-Jun N-terminal kinase(JNK)histone modification and expression in hippocampus of offspring before and after birth,and to clarify the mechanism of intrauterine programming of the high stress sensitivity of HPA axis induced by PDE in progeny rats.The fetal hippocampus cell line was further used,and through a series of interference experiments,the JNK histone modification and its expression were determined to mediate the effect of dexamethasone on GAD67 expression.Methods: Treatment of pregnant rats is the same as the first part.Partially(at least 12 rats in each group)pregnant rats are anesthetized on GD20,and fetal rats are harvested by laparotomy.Peripheral blood of the fetal rats is collected.Three fetal rats were taken for whole brain fixation.The other part is the same as the first part,after feeding the baby rats to 10 weeks after birth,some rats are randomly divided into two groups(Non-chronic stress group and chronic stress group).The chronic stress group was given 5 minutes of ice-water(5-7℃)swimming every day for 2 weeks.The last swim was performed with ether anesthesia after 1 hour,and the head was sacrificed to take blood,and the hypothalamus and hippocampus were removed.The three brains of the offspring were fixed.The remaining part of the PDE group female pups and normal male rats were caged 2:1 to obtain F2 generation pups.F2 generation female offspring were caged with normal male rats at 12 weeks after birth to obtain F3 generation offspring.F2 and F3 generation female pups were given the same ice water swimming stimulation as F1 generation in 10-12 weeks.After the last ice water swimming,the pups were taken from the hippocampus.Real-time quantitative PCR was used to detect the m RNA expression of CRH,AVP,VGlu T2,GAD65,PSD95,Reelin and α-Ca MKII in fetal rat hypothalamus;the m RNA expressions of GAD67,Sgk1,Fkbp5 and HDAC2 in the hippocampus of fetal and adult pups.Immunofluorescence staining was used to observe the activity changes of Glu/GABAergic neurons in the hippocampus of the fetal rat and the adult pups.Western blotting was used to detect the expression of GAD67 protein in the hippocampus of fetal and adult pups.Biochemical and ELISA techniques were used to detect the levels of neurotransmitters GABA and Glu in adult pups and fetal rat hippocampus tissues.Ch IP technology was used to detect the level of acetylation in the JNK promoter region H3K9/H3K14/H3K27.H19-7 fetal hippocampus cell line was treated with different concentrations of dexamethasone(0,0.02,0.1,0.5 μM)for 3 days.Further,before treatment with 0.5 μM concentration of dexamethasone,5 μM GR antagonist RU486 or 1 μM HDAC2 specific inhibitor Romidepsin was given.MTS method was used to detect cell viability,RT-q PCR was used to detect GR,GAD67,Sgk1,Fkbp5,HDAC2 m RNA expression,Ch IP technology was used to detect the acetylation level of JNK promoter region H3K14,and immunofluorescence staining was used to observe GAD67 expression in cells.Results:(1)Changes in GAD67 expression,excitability/inhibitory transmitter content in hippocampus of adult F1 generation offspring: In the non-chronic stress group,the m RNA/protein expression and GABA content of hippocampus GAD67 in female and male adult offspring in PDE group were higher than those in control group(P<0.05,P<0.01).In the chronic stress group,although the Glu content in the hippocampus of female and male adult rats in the PDE group was not significantly changed compared with the control group,the m RNA expression and GABA content of GAD67 were higher than the control group(P<0.05,P<0.01).(2)JNK histone acetylation and expression changes in hippocampus of F1 generation adult pups: In non-chronic stress group or chronic stress group,the expression of JNK in hippocampus of female and male adult pups in F1 generation PDE group was significantly lower than that of control group(P<0.01),And the level of H3K14 ac in the JNK promoter region was also lower than that in the control group(P<0.01).(3)Changes of JNK/GAD67 expression and JNK epigenetic modification in hippocampus after chronic stress of F2/F3 generation offspring rats: Histone acetylation level and JNK and GAD67 m RNA expression of JNK promoter in F2 offspring female PDE group offspring after chronic stress There was no significant difference from the control group;however,the level of JNK promoter H3K14 ac of female pups in the F3 generation PDE group after chronic stress was significantly lower than that of the control group(P<0.01),and the m RNA expression of JNK was lower than that of the control group(P<0.01),GAD67 m RNA expression was higher than the control group(P<0.05).(4)F1 generation fetal rat hippocampus GAD67 expression,excitatory/inhibitory transmitter content changes: PDE group female and male fetal hippocampus Glu content decreased but GAD67 expression increased(P<0.05,P<0.01),while PDE group fetal rat hippocampus GAD67 MRNA and protein expression were also significantly higher than the control group(P<0.05,P<0.01).It was further found that the Glu content in the fetal hippocampus of female and male offspring in the PDE group was lower than that in the control group but the GABA content was higher than that in the control group(P<0.05,P<0.01).(5)F1 generation fetal rat hippocampus GR activation status,HDAC2 expression and JNK histone acetylation and expression changes: PDE group female and male fetal hippocampus Sgk1,Fkbp5 m RNA expression were significantly higher than the control group(P<0.01),while fetal HDAC2 expression in the hippocampus was higher than that in the control group and JNK expression was lower than that in the control group(P<0.05,P<0.01).The levels of H3K14 ac in the JNK promoter region of female and male fetal hippocampus in the PDE group were lower than those in the control group(P<0.01),while the levels of H3K9 ac and H3K27 ac did not change significantly.(6)Cell-level verification GR/HDAC2 mediated dexamethasone-induced JNK histone acetylation and expression changes: three days after administration of different concentrations(0,0.02,0.1,0.5 μM)of dexamethasone treated cells,fetal hippocampal cells GR,The m RNA expression of HDAC2 and GAD67 increased(P<0.05,P<0.01),while the m RNA expression of JNK decreased in a concentration-dependent manner(P<0.01).Furthermore,pretreatment of cells with 5 μM RU486 can reverse the increase in m RNA expression of Fkbp5,Sgk1,HDAC2,and GAD67,which are downstream targets of GR caused by 0.5 μM dexamethasone(P<0.01),and H3K14 ac levels in JNK promoter And JNK m RNA expression decreased(P<0.01).Similarly,pretreatment of cells with 1 μM Romidepsin can reverse the level of H3K14 ac in the JNK promoter region caused by 0.5 μM dexamethasone,decrease the m RNA expression of JNK and increase the expression of GAD67 m RNA(P<0.01).Immunofluorescence detection results also showed that RU486 and Romidepsin pretreatment can reverse the increase in GAD67 expression caused by dexamethasone,making it close to the control group.Conclusion: PDE can activate the GR of fetal rat hippocampus,and up-regulation of HDAC2 expression causes the decrease of histone acetylation level in the JNK promoter region which inhibits its expression.The inhibition of JNK can increase the expression of GAD67 in the hippocampus,further promote the conversion of Glu to GABA,and cause excitement in the hippocampus.Inhibitory neurotransmitter imbalance,weakened excitatory output of hippocampal pyramidal neurons to BST,enhanced inhibitory output,weakened Glu-GABA signal transmission in the BST area,and finally caused the GABAergic inhibition signal projected to the PVN area of the hypothalamus to weaken.The negative regulatory effect of the thalamus is weakened,resulting in increased potential excitability of the hypothalamus.In the PDE group,female F3 hippocampus still maintain low levels and low expression of histone acetylation,GAD67 expression is still higher than the control group and the hypothalamus potential excitability is increased.This may be the reason why the HPA axis high stress sensitivity changes are inherited from the maternal line to the F3 generation.PART THREEPDE-induced offspring susceptibility to epilepsy and intrauterine programming mechanismObjective: Using animal experiments to confirm the susceptibility to epilepsy in PDE offspring rats after birth,and explore the effects of PDE on the acetylation modification and expression of histone hippocampal KCC2 histones.Using the experiment of fetal hippocampus cells,the mechanism of intrauterine programming of susceptibility to epilepsy in progeny rats induced by PDE was clarified.Methods: Partial treatment of pregnant rats is the same as the first and second parts.Anesthetize a part of GD20 rats after isoflurane anesthesia,take the fetal rat hippocampus quickly under the microscope and fix 3 fetal rat whole brains in each group.The other part of the pregnant rats was born naturally until the offspring were weaned at PW4.Three male offspring were randomly selected from each litter and divided into 2 groups(control group and PDE group)and 3 batches.The first batch rats were anesthetized with 2% isoflurane and sacrificed at PW12.Three randomly selected whole brains were fixed in 4% paraformaldehyde for immunofluorescence detection.The remaining hippocampal samples were collected and immediately frozen in liquid nitrogen and stored at-80℃;the second batch of rats underwent Lithium pirucapine chloride(Li PC)epilepsy modeling at PW12,and Electroencephalogram(EEG)and video monitoring were performed;the third batch of rats underwent Li PC epilepsy modeling at PW12 and then a week-long behavioral test in PW18.The m RNA expressions of KCC2,Sgk1,Fkbp5 and HDAC2 in the hippocampus of fetal and young rats were detected after birth.Immunofluorescence was used to detect the expression of KCC2 in the hippocampus of the fetal and hippocampus after birth.Western blotting was used to detect the expression of KCC2 protein in the hippocampus of fetal and adult pups.Ch IP technology was used to detect the histone acetylation level in the KCC2 promoter region of fetal rats and adult hippocampus.The H19-7 fetal hippocampal cell line was treated with different concentrations of dexamethasone(0,0.02,0.1,0.5 μM)for 3 days.Further,before treatment with 0.5 μM concentration of dexamethasone,5 μM GR intervention agent RU486 or 1 μM HDAC2 specific inhibitor Romidepsin was given.MTS method was used to detect cell activity,RT-q PCR was used to detect GR,KCC2,Sgk1,Fkbp5,HDAC2 m RNA expression,Ch IP technology was used to detect the acetylation level of KCC2 promoter region H3K14 ac,immunofluorescence staining was used to observe the expression of KCC2 in cells.Results:(1)The susceptibility to epilepsy and cognitive impairment of the offspring after birth: Compared with the control group,the seizure grade of male offspring of the PDE group increased in the Li PC-induced epilepsy model(P<0.05),and the time of first seizure decreased(P<0.01),EEG showed epilepsy activation(P<0.01)and increased seizure-like electrical activity.It was further found that during the 6 consecutive days of training,the escape latency and movement distance of male offspring in the PDE group were significantly prolonged(P<0.05,P<0.01).In the hidden platform test,the spatial positioning ability of male rats in PDE group decreased(P<0.05).In the detection test,the spatial positioning ability of male offspring rats in the PDE group was reduced(P<0.01),and the recognition rate of new things and new positions was significantly reduced(P<0.05).(2)KCC2 expression/epigenetic changes after birth: the m RNA and protein expression of hippocampus KCC2 in male offspring of PDE group were significantly lower than that of control group(P<0.01).The histone acetylation level of H3K14 in the hippocampus KCC2 promoter region of male progeny rats in the PDE group was lower than that in the control group(P<0.01).(3)GR activation status of fetal rat hippocampus,acetylation and expression of KCC2 protein,intracellular chloride ion concentration: m RNA expression of male fetal rat hippocampus GR and its downstream target Fkbp5/Sgk1 in PDE group was higher than that of control group(P<0.01),HDAC2 m RNA expression was also significantly higher than the control group(P<0.01),KCC2 m RNA expression was lower than the control group(P<0.01).The expression of KCC2 protein in male fetal hippocampus of PDE group was higher than that of control group(P<0.01).The acetylation level of KCC2 promoter region H3K14 in male fetal hippocampus of PDE group was lower than that of control group(P<0.01).The chloride ion concentration in male fetal hippocampus cells of PDE group was higher than that of control group(P<0.01).(4)KCC2 expression and histone acetylation changes in fetal hippocampal neuronal cell line: after 3 days of dexamethasone treatment,the expression of Fkbp5/Sgk1 m RNA was higher in the treatment group than in the control group(P<0.01).The expression of HDAC2 in the group was also higher than that in the control group(P<0.01),and the expression of KCC2 in the 0.1 μM and 0.5 μM dexamethasone-treated group was lower than that in the control group(P<0.05).The acetylation level of H3K14 in KCC2 of 0.5 μM dexamethasone-treated group was lower than that of the control group(P<0.05).After 3 days of treatment with 0.5 μM dime,the chloride ion concentration in the medium did not change significantly,while the intracellular chloride ion concentration in fetal hippocampal neurons was significantly higher than that in the control group(P<0.01).(5)Cell-level counter-evidence test: cells were given 0.5 μM concentration of dexamethasone before treatment,5 μM RU486 was given,Fkbp5,Sgk1 and HDAC2 m RNA expression decreased to the control level(P<0.01);KCC2 m RNA expression level was also increased significantly than dexamethasone group(P<0.01),and H3K14 ac level in the KCC2 promoter region was higher than that of the dexamethason treatment group and the control group was close(P<0.01).Administration of 0.5 μM concentration of dexamethasone to cells before administration of 1 μM Romidepsin can reverse the effect of dexamethasone treatment on H3K14 ac in the KCC2 promoter region(P<0.05),and restore m RNA expression of KCC2 to control level(P<0.05).Immunofluorescence showed that both RU486 and RO treatment could reverse the effect of dexamethasone-induced KCC2 expression reduction,making it close to the control group.Conclusion: PDE can cause increased susceptibility to epilepsy in male offspring.The mechanism of hippocampal programming is: PDE can activate fetal hippocampus GR,upregulate the expression of HDAC2,and reduce histone acetylation in the hippocampus KCC2 promoter region.Modification and expression.KCC2 is one of the key proteins that affect the internal and external balance of chloride ions during the development of neurons.The decreased expression will cause the intracellular chloride ion concentration in neurons to be too high,and the inhibitory effect of GABAergic nerve signals can be weakened.These changes can last after birth,hippocampal neurons are more likely to be overexcited in epilepsy modeling,increasing the susceptibility to epilepsy.Summary:This study confirmed that PDE can cause high stress sensitivity of HPA axis in offspring adult rats,and has a maternal intergenerational genetic effect.The occurrence of high stress sensitivity on the HPA axis of PDE adult offspring rats is related to the increased local excitability of the adult hypothalamus(specifically the expression ratio of v Glu T2/GAD65),which is partly due to the change in potential excitability in the uterus.The hippocampus acts as a high negative feedback regulation center for the HPA axis,and participates in the occurrence of PDE-induced high stress sensitivity of the HPA axis in adult offspring rats.The level of H3K14 ac in the JNK promoter region is related to m RNA expression,which in turn leads to increased expression of the downstream target gene GAD67.The high expression of GAD67 in hippocampus can promote the excessive conversion of Glu to GABA in the local tissues,causing the weakening of the GABA inhibition signal projected into the PVN area of the hypothalamus,resulting in the weakening of the negative regulatory effect of the hippocampus on the hypothalamus.This high expression of GAD67 in the hippocampus mediated by epigenetic mechanism has an intrauterine programming effect,which can continue to the F1 generation after birth or even to the F3 generation,resulting in increased potential excitability of the hypothalamus in the adult offspring of the F1/F3 generation,thereby increasing its stress sensitivity of the HPA axis.This study confirmed the existence of hippocampal developmental programming and steady-state changes(enhanced stress sensitivity)in dexamethasone,and further confirmed the existence of epilepsy susceptibility after birth in PDE offspring rats through the Li PC epilepsy model(second shock).The mechanism is related to intrauterine dexamethasone exposure through activation of fetal hippocampus GR and up-regulation of HDAC2 expression,thereby reducing KCC2 expression,which in turn leads to excessively high chloride ion concentration in neuronal cells and weakened GABAergic inhibition signal.This low expression of hippocampal KCC2 mediated by epigenetic mechanism has an intrauterine programming effect,which can last after birth,causing hippocampal neurons to be overexcited and abnormally discharged,thereby increasing the susceptibility of the offspring to epilepsy after birth. |
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