The role of astrocytes in Rett syndrome

描述（由申请人提供）：Rett综合征（RTT）是由编码甲基CpG结合蛋白2（MeCP 2）的X连锁MECP 2（一种基因转录的表观遗传调节因子）的功能缺失突变引起的。RTT和更普遍的倒退型自闭症有大量的表型重叠，包括自闭症行为。受影响的儿童在经过一段明显正常的生长和发育后，开始失去发达的技能（在本建议中称为“退化”）。RTT是一种由于表观遗传调节异常引起的突触可塑性疾病，如果突触功能障碍可以改善，则可能是可治愈的。对于我们理解RTT的关键问题包括什么机制触发消退，以及星形胶质细胞是否通过调节突触活动在RTT的发生和进展中发挥作用。我们的研究揭示了MeCP 2缺陷型星形胶质细胞的多种异常，包括无法支持树突状细胞生长，自发和诱发的Ca 2+信号传导减少，对ATP和谷氨酸挑战的异常反应，以及异常的D-丝氨酸释放等。我们推测RTT中的进行性星形胶质细胞异常可能会恶化神经元-胶质细胞的通讯并引发消退。为了生成支持这一假设的数据，我们的目标是：1。体外和原位研究RTT星形胶质细胞胞内钙信号异常的机制。我们将扩展我们现有的体外数据，并研究主要的细胞途径和调节细胞内钙离子动态的分子。我们还将使用海马切片制备物原位复制我们的结果。 2.确定星形胶质细胞MeCP 2缺乏对海马长时程增强（LTP）的影响，重点是D-丝氨酸的作用。我们将测量混合神经元-胶质细胞培养物和急性海马切片中的LTP，以确定MeCP 2缺陷型星形胶质细胞对RTT样本中观察到的异常LTP的贡献。随后，我们将确定添加D-丝氨酸（一种胶质递质，其由MeCP 2缺陷型星形胶质细胞释放显著减少）是否可以减轻LTP损伤。 3.确定星形胶质细胞MeCP 2缺陷在体内RTT样症状发作和进展中的作用。我们将生成小鼠模型，其中星形胶质细胞Mecp 2在野生型背景中选择性关闭或在选择的时间在Mecp 2-null背景中打开。我们将检查这些小鼠的行为表型、神经病理学和生化变化。这些结果有望促进我们对星形胶质细胞在发育过程中的生理和病理功能的理解，为未来的研究提供有用的模型，并确定RTT的潜在治疗靶点。雷：雷特综合征是一种大脑发育的遗传性疾病，折磨着非常年轻的女孩。星形胶质细胞是一种活跃的神经元伴侣脑细胞，在Rett综合征动物模型中高度异常。我们的目标是发现星形胶质细胞异常的分子机制，并产生新的小鼠模型来证明星形胶质细胞在Rett综合征中的重要作用。

DESCRIPTION (provided by applicant): Rett syndrome (RTT) is caused by loss-of-function mutations in the X-linked MECP2 encoding methyl-CpG-binding protein 2 (MeCP2), an epigenetic modulator of gene transcription. RTT and the more prevalent regressive type autism share a substantial phenotypic overlap, including autistic behaviors. The affected children start to lose developed skills (called "regression" in this proposal) after a period of apparently normal growth and development. RTT is a disease of synaptic plasticity due to abnormal epigenetic regulation, and is potentially curable if synaptic dysfunction can be ameliorated. Critical questions for our understanding of RTT include what mechanisms trigger the regression, and if astrocytes, by regulating synaptic activities, play a role in RTT onset and progression. Our studies revealed multiple abnormalities of MeCP2-deficient astrocytes, including failure to support dendritic growth, reduced spontaneous and evoked Ca2+ signaling, aberrant responses to ATP and glutamate challenges, and abnormal D-serine release, among others. We hypothesize that the progressive astrocytic abnormalities in RTT may deteriorate the neuron-glia communications and trigger the regression. To generate data in support of this hypothesis, our aims are: 1. Investigate the mechanism of abnormal intracellular Ca2+ signaling in RTT astrocytes in vitro and in situ. We will extend our existing in vitro data and examine major cellular pathways and molecules regulating intracellular Ca2+ dynamics. We will also replicate our results in situ using hippocampal slice preparations. 2. Determine the impact of astrocytic MeCP2-deficiency on hippocampal long-term potentioation (LTP), focusing on the role of D-serine. We will measure LTP in mixed neuron-glia cultures and acute hippocampal slices to determine the contribution of MeCP2-deficient astrocytes to abnormal LTP seen in RTT samples. Subsequently, we will determine if the addition of D-serine, a gliotransmitter the release of which by MeCP2-deficient astrocytes is substantially reduced, can alleviate the LTP impairments. 3. Determine the role of astrocytic MeCP2-deficiency in the onset and progression of RTT-like symptoms in vivo. We will generate mouse models in which astrocytic Mecp2 is selectively turned off in the wild-type background or turned on in the Mecp2-null background at the time of choice. We will examine the behavioral phenotype, neuropathology and biochemical changes of these mice. The results are expected to advance our understanding of the physiological and pathological functions of astrocytes during development, provide useful models for future studies, and identify potential therapeutic targets for RTT. Lay: Rett syndrome is a genetic disorder of brain development that afflicts very young girls. Astrocytes, a type of brain cells that are active partners of neurons, are highly abnormal in an animal model of Rett syndrome. The aims of our proposal are to discover the molecular mechanisms underlying astrocytic abnormalities, and to generate new mouse models to prove the important role of astrocytes in Rett syndrome.