Derivation of GABAergic interneurons from mouse embryonic stem cells

描述（由申请人提供）：gaba能神经元是大脑皮层抑制性电路的主要来源，在感知、协调运动、学习和记忆过程中，gaba能神经元需要在网络内的神经元之间产生持续振荡的同步活动。抑制神经元传递的皮质缺陷与癫痫、自闭症和精神分裂症等神经精神疾病有关。产生无限供应的gaba能神经元将允许这些细胞在各种疾病模型中广泛使用，以评估它们在基于细胞的治疗中的应用潜力。这一提议的目标是从小鼠胚胎干细胞（ES）细胞中产生皮层gaba能中间神经元。通过对已发表方案的修改，我将首先优化共表达Nkx2.1的端脑（FoxG1+）祖细胞的生产，Nkx2.1是啮齿动物皮质中间神经元主要亚群的指定所必需的转录因子。由于Nkx2.1也会产生其他皮层下生成的细胞群体，因此我已经生成了一个在Lhx6启动子区域控制下表达绿色荧光蛋白的ES细胞系，Lhx6是一种转录因子，其表达在Nkx2.1谱系的神经元间前体中富集，并通过出生后的发育在大多数这些细胞中维持。利用这个和其他工具，我将引导胚胎干细胞进入假定的中间神经元祖细胞（有丝分裂）或前体细胞（有丝分裂后，但尚未迁移到皮层）状态，并测试它们在出生后小鼠皮层内迁移、存活和分化的能力。这些实验可能对耐药性慢性局灶性癫痫的细胞治疗具有重要意义。此外，成功地从胚胎干细胞中衍生出nkx2.1谱系的中间神经元将为研究调节这些关键神经元发育的内在和外在因素提供宝贵的系统。

DESCRIPTION (provided by applicant): By providing the major source of inhibitory circuitry in the cerebral cortex, GABAergic neurons are required for the synchronous activity necessary to generate sustained oscillations among neurons within a network that are essential during perception, coordinated movement, learning and memory. Cortical deficits in inhibitory neuronal transmission have been implicated in neuropsychiatric disorders such as epilepsy, autism, and schizophrenia. Generating a limitless supply of GABAergic neurons from would allow for the extensive use of these cells in a variety of disease models in order to evaluate their potential for use in cell based therapies. The goal of this proposal is to generate cortical GABAergic interneurons from mouse embryonic stem (ES) cells. Using modifications of published protocols, I will first optimize the production of telencephalic (FoxG1+) progenitors that co-express Nkx2.1, a transcription factor that is required for the specification of major subgroups of cortical interneurons in rodents. Since Nkx2.1 also gives rise to other populations of subcortically generated cells, I have generated an ES cell line that expresses green fluorescent protein under control of the promoter region of Lhx6, a transcription factor whose expression is enriched within Nkx2.1-lineage interneuron precursors and maintained in most of these cells through postnatal development. Using this and other tools, I will direct ES cells towards putative interneuron progenitor (mitotic) or precursor (postmitotic, but not yet migrated into cortex) states and test their capacity to migrate, survive, and differentiate within postnatal mouse cortex. These experiments could have important implications for cell based therapies for medication resistant chronic focal epilepsy. In addition, successful derivation of Nkx2.1-lineage interneuron's from ES cells would provide an invaluable system for the study of intrinsic and extrinsic factors regulating the development of these critical neurons.