宫内GAD67高表达编程介导孕期乙醇暴露所致的子代下丘脑低基础活性

Lower basal activity of hypothalamic–pituitary–adrenal (HPA) axis is the major cause of many fetal origin diseases . However, the mechanism involved in the decreased activity of HPA axis has been not reported yet. Previous studies in our lab demonstrated that prenatal ethanol exposure (PEE) can cause over-exposure of maternal glucocorticoid (GC) and lower basal activity of HPA axis in offspring. Recently, we found that PEE can increase the expression of hypothalamic rate-limiting enzyme in the synthesis of GABA - Glutamic acid decarboxylase (GAD) 67 in fetal rats and also the adult rats. Based on literatures, we hypothesized that PEE cause high GC concentration of serum, GR binds to hypothalamic GC and translocates to the nucleus, through change early growth response factor 1 and DNA methyltransferase 1 expression, resulting in the epigenetic alteration of GAD67 promoters and enhance the expression of GAD67. Enhanced expression of GAD67 can increase the transformation of glutamate into GABA in the hypothalamus, which eventually led to the weakened function of hypothalamus and low basal activity programming. In the present study, we will use lab animals to confirm the important role of hypothalamic GAD67 intrauterine programming alteration in the decreased basal activity of hypothalamus induced by PEE. Further, we will clarify the epigenetic mechanism for the enhanced expression of fetal hypothalamic GAD67 in animal and cell levels, to clarify the intrauterine mechanism of decreased basal activity of hypothalamus induced by PEE.

下丘脑-垂体-肾上腺（HPA）轴低基础活性是多种胎源性疾病的主要诱因，然而其发生机制尚未明了。我们近期发现，孕期乙醇暴露可致子代大鼠过暴露母源性糖皮质激素（GC），且出生前、后下丘脑功能抑制及子代HPA轴低基础活性变化；进一步发现，出生前、后下丘脑GABA合成限速酶—左旋谷氨酸脱羧酶（GAD）67表达增加。综合文献，我们推测：孕期乙醇暴露下的胎血高GC可诱导下丘脑糖皮质激素受体活化转位至胞核，通过改变早期生长反应因子1和DNA甲基转移酶1的表达，导致GAD67启动子区表观遗传修饰异常及高表达，促使Glu向GABA转化，最终导致下丘脑功能减弱和低基础活性编程。本项目拟在整体水平证实下丘脑GAD67宫内编程改变在孕期乙醇暴露所致子代HPA轴低基础活性中的核心作用，进一步从整体和细胞水平证实GC上调胎下丘脑GAD67表达的表观遗传调控机制，以阐明孕期乙醇暴露所致下丘脑低基础活性的宫内发生机制。

下丘脑-垂体-肾上腺（HPA）轴低基础活性是多种胎源性疾病的主要诱因，然而其发生机制尚未明了。我们研究发现，孕期乙醇暴露（PEE）可致子代大鼠过暴露于母源性糖皮质激素（GC），胎血高GC可诱导下丘脑GC受体活化转位至胞核，导致下丘脑中GC代谢活化系统（11βHSDs/GR/C/EBPa）活化，活化的GR进一步增加左旋谷氨酸脱羧酶（GAD）67表达，从而促进下丘脑神经元的胞质和囊泡中谷氨酸（Glu）向γ-氨基丁酸(GABA)的生物转化，导致投射至下丘脑室旁核（PVN）区的Glu能/GABA能神经元发育失衡，介导PVN区Glu能/GABA能的重塑并持续到出生后甚至成年，最终导致下丘脑的小细胞神经元活性抑制和低功能表达，以及HPA轴低基础活性（表现为下丘脑CRH、AVP低表达和血ACTH、CORT水平降低）；而PEE雌性子代大鼠HPA轴低基础活性发生机制与雄性不同，可能是由于PEE所致母源性GC过暴露，诱导雌性胎下丘脑IGF1信号通路功能下调编程改变，导致其神经元发育不良，并以Glu能神经元显著，Glu能神经元发育不良及功能活性降低延续到了出生后，导致子代下丘脑基础状态下低功能表达及HPA轴低基础活性。. 为确证PEE所致母源性GC过暴露是导致雄性子代下丘脑GAD67宫内编程改变的主要因素，我们进一步观察到孕期地塞米松暴露（PDE）可致雄性IUGR子代下丘脑功能低下和HPA轴低基础活性，其机制也与PDE所致下丘脑GAD67宫内编程改变相关。本研究有助于阐明孕期外源物（乙醇/地塞米松）暴露所致下丘脑低功能表达及HPA轴低基础活性的宫内发生机制。同时，我们还发现，PEE子代大鼠成年期在受到刺激时表现为高应激敏感性（下丘脑CRH、AVP高表达及血CORT、ACTH水平增加）以及下丘脑潜在兴奋性增加（VGluT2/GAD65表达比增加），可能与PEE所致母源性GC过暴露，导致海马GAD67启动子区-1019~-691 bp去甲基化及GAD67表达上调，海马Glu能/GABA能神经元发育失衡，介导海马对下丘脑的负调控作用减弱及下丘脑VGluT2/GAD65表达比增加所致下丘脑潜在兴奋性增加相关，并最终表现为出生后高应激敏感性。

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