CNS Myelination Co-Culture Microsystem for Axon-Gila Signaling

描述(申请人提供)：由少突胶质细胞产生的髓鞘对大脑发育和功能是必不可少的。少突胶质细胞和髓鞘功能障碍现在被认为是与多发性硬化症、精神分裂症、抑郁症和阿尔茨海默病相关的许多神经缺陷的原因。然而，髓鞘形成的少突胶质细胞及其控制髓鞘形成过程的轴突之间的信号在很大程度上仍不清楚。这在一定程度上是由于缺乏有效的体外髓鞘形成模型，该模型可以通过中枢神经系统(CNS)中的少突胶质细胞重新捕获髓鞘轴突的关键事件。我们研究的长期目标是开发和利用微系统，以了解中枢神经系统髓鞘形成/脱髓鞘的分子和细胞基础，并开发促进损伤后髓鞘修复的策略。本研究的目的是开发和实施一种新型的体外中枢神经系统髓鞘形成系统，该系统能够精确控制髓鞘形成过程中的各种信号因子，旨在揭示髓鞘形成的分子基础，促进髓鞘修复。通过一种创新的、多学科的方法，我们最近开发了一种新的分区的髓鞘共培养微系统，在该系统中，皮质神经元的轴突被引导从细胞体生长到流体隔离的微环境中，在那里它们与少突胶质细胞相互作用并被包裹。在这项计划中，我们将进一步发展我们的初步微设备，使其成为一种新型的体外中枢神经系统髓鞘微系统，该系统能够以单一细胞分辨率精确控制多种细胞类型的位置和密度，能够以精确的空间和时间分辨率施加局部化学刺激，并且能够以高通量施加局部电刺激。我们将首先描述髓鞘微系统中的髓鞘生成。我们将可视化髓鞘形成的过程，并具体研究细胞与细胞之间的相互作用、生长和抑制因子、模拟神经元活动的电刺激在调节髓鞘少突胶质细胞发育中的作用。轴突神经调节蛋白-1决定了周围神经系统髓鞘形成的开始和程度，但其在中枢神经系统髓鞘形成中的作用尚不清楚。因此，我们将专门确定neuRegin-1和睫状神经营养因子家族成员在调控髓鞘少突胶质细胞发育中的作用。最后，集成的多电极阵列将用于选择性刺激神经元，并研究轴突放电对髓鞘形成的影响。这些研究将对中枢神经系统轴突髓鞘形成的机制提供重要的新见解，以及可用于髓鞘研究和药物开发和测试的强大的中枢神经系统髓鞘形成模型系统。公共卫生相关性：在这项提议完成后，我们希望建立一个新的模型来研究髓鞘形成，这是一个对大脑功能至关重要的过程，并发现促进髓鞘形成的信号，从而促进脱髓鞘疾病的髓鞘修复。我们细胞培养微系统的独特功能还使其能够用于筛选和测试与髓鞘功能障碍相关的神经系统疾病的药物，如多发性硬化症、阿尔茨海默病和精神分裂症。

DESCRIPTION (provided by applicant): Myelin produced by oligodendrocytes is essential for brain development and function. Dysfunction of oligodendrocytes and myelin is now thought to contribute to many neurological deficits associated with multiple sclerosis, schizophrenia, depression and Alzheimer's disease. However, the signals between myelinating oligodendrocytes and their underlying axons that control the myelination process remain largely unknown. This is in part due to lack of efficient in vitro myelination models that recapture key events of myelinating axons by oligodendrocytes in the central nervous system (CNS). The long-term goal of our research is to develop and utilize microsystems toward understanding the molecular and cellular basis of CNS myelination/demyelination and to develop strategies that promote myelin repair after injury. The purpose of this proposal is to develop and implement a novel in vitro CNS myelination system that enables precise control over various signaling factors in myelinogenesis aimed at uncovering the molecular basis of myelination and promoting myelin repair. Through an innovative and multidisciplinary approach, we have recently developed a novel compartmentalized myelination co-culture microsystem where axons from cortical neurons are guided to grow away from cell bodies into a fluidically isolated microenvironment where they interact with and are ensheathed by oligodendrocytes. In this proposal, we will further develop our preliminary microdevice into a novel in vitro CNS myelination microsystem that can accurately control the positions and densities of multiple cell types with single cell resolution, can apply localized chemical stimuli with precise spatial and temporal resolution, and can apply localized electrical stimuli, all at high throughput. We will first characterize myelinogenesis in the myelination microsystem. We will visualize the myelination process and examine specifically the role of cell to cell interactions, growth and inhibition factors, electrical stimulation mimicking neuronal activity in regulating the development of myelinating oligodendrocytes. Axonal neuregulin-1 determines the onset and the extent of myelination in the peripheral nervous system, but its role in CNS myelination is not clear. Therefore, we will specifically determine the role of neuregulin-1 and members of ciliary neurotrophic factor family in regulating the development of myelinating- oligodendrocytes. Finally, the integrated multielectrode arrays will be used to selectively stimulate neurons and study the effect of axonal firing on myelinogenesis. These studies should provide important new insights into the mechanisms responsible for CNS axon myelination as well as a powerful CNS myelination model system that can be exploited for myelin researches and drug development and testing. PUBLIC HEALTH RELEVANCE: Upon completion of this proposal, we hope to establish a novel model for studying myelination, a process essential to brain functions, and to uncover signals that promote myelination and thus facilitate myelin repair in demyelinating diseases. The unique feature of our cell culture microsystem also allows it to be used for screening and testing drugs for neurological diseases associated with myelin dysfunction such as multiple sclerosis, Alzheimer's disease and schizophrenia.