Derivation of GABAergic interneurons from mouse embryonic stem cells

描述(申请人提供)：通过提供大脑皮层抑制电路的主要来源，GABA能神经元需要同步活动，以在网络中的神经元之间产生持续的振荡，这在感知、协调运动、学习和记忆过程中是必不可少的。抑制神经元传递中的皮质缺陷与癫痫、自闭症和精神分裂症等神经精神障碍有关。从细胞中产生无限的GABA能神经元将允许在各种疾病模型中广泛使用这些细胞，以评估它们在基于细胞的治疗中的潜力。这项提议的目标是从小鼠胚胎干细胞中产生皮质GABA能中间神经元。利用已发表的方案的修改，我将首先优化共表达Nkx2.1的端脑(FoxG1)前体细胞的生产，Nkx2.1是一种转录因子，是指定啮齿动物皮质间神经元主要亚群所必需的。由于Nkx2.1还产生了其他皮质下产生的细胞群，我已经产生了一个在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，其表达丰富在Nkx2.1系神经元间前体细胞中，并在出生后发育期间在大多数这些细胞中保持表达。使用这一工具和其他工具，我将引导ES细胞进入假定的神经元间祖细胞(有丝分裂)或前体(有丝分裂后，但尚未迁移到皮质)状态，并测试它们在出生后小鼠皮质内迁移、存活和分化的能力。这些实验可能对药物耐药的慢性局灶性癫痫的基于细胞的治疗具有重要的意义。此外，成功地从ES细胞中获得Nkx2.1系中间神经元，将为研究调控这些关键神经元发育的内在和外在因素提供一个宝贵的系统。

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.