Derivation of GABAergic interneurons from mouse embryonic stem cells

描述（由申请人提供）：通过提供大脑皮层中抑制回路的主要来源，GABA能神经元需要同步活动，所述同步活动是在感知、协调运动、学习和记忆期间在网络内的神经元之间产生持续振荡所必需的。抑制性神经元传递的皮质缺陷与神经精神疾病如癫痫、自闭症和精神分裂症有关。产生无限供应的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应用潜力。该提案的目标是从小鼠胚胎干（ES）细胞中产生皮质GABA能中间神经元。使用已发表的协议的修改，我将首先优化生产端脑（FoxG1+）祖细胞，共表达Nkx2.1，转录因子，需要在啮齿类动物的皮质中间神经元的主要亚群的规范。由于Nkx2.1也产生了其他群体的皮层下产生的细胞，我已经产生了一个ES细胞系，表达绿色荧光蛋白的启动子区的控制下Lhx6，一个转录因子，其表达是丰富的Nkx2.1谱系中间神经元前体，并保持在大多数这些细胞通过出生后的发展。使用这个和其他工具，我将直接ES细胞对假定的interneuron祖细胞（有丝分裂）或前体（有丝分裂后，但尚未迁移到皮质）状态和测试他们的迁移能力，生存，并在出生后的小鼠皮质分化。这些实验可能对基于细胞的药物耐药性慢性局灶性癫痫治疗具有重要意义。此外，成功地从ES细胞中获得Nkx2.1谱系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.