Training Novel Host-Graft Interfaces to Enhance Spinal Cord Repair

描述(由申请人提供)：神经元替换显示出促进脊髓损伤(SCI)后恢复的希望，并被认为是干细胞介导的脊髓修复的主要目标。然而，实现宿主-移植物神经元的有效沟通是神经元替换的一个重大挑战，目前还没有进行过研究。这一提议的总体假设是，呈现新建立的宿主-移植物神经元网络具有生理模式的活动将增强功能连接。基于我们在脊髓损伤模型以及呼吸和计算神经生物学方面的丰富经验，我们建议通过将大鼠胚胎脊髓(FSC)组织移植到临床相关的成年大鼠高位颈椎(C3)挫伤中来验证这一假设。初步的跨神经元神经解剖示踪数据显示，FSC供体细胞与宿主膈运动神经元和颈部中间神经元之间存在突触连接。此外，FSC移植物从宿主接受广泛的5-羟色胺能输入，一些移植物神经元表现出对缺氧敏感的放电模式，包括明显的吸气相关的爆发。因此，一个必要的解剖学-功能框架已经到位，可以通过以下特定目的使用FSC移植到成年大鼠的C3挫伤中来检验我们的中心假设：目的1)测试FSC来源的神经元将在解剖和生理上与宿主灰质整合的假设，以及目的2)测试通过间歇低氧(IH)刺激的“训练”将增强FSC来源的神经元的解剖和生理整合的假设。为了验证这些假设，我们将使用多学科方法，包括移植物和宿主脊髓之间的连接的神经解剖学研究，清醒大鼠的呼吸测量，以及移植物的神经生理学研究。我们建议的一个创新技术特征是，这将是首次使用微电极阵列来监测脊髓中移植相关的神经丛。将使用FSC移植方法，因为这样的移植物会发育成含有大量类似于中间灰质中间神经元的细胞的有髓组织。一种独特的康复模式-每天暴露在轻度间歇性低氧(IH)-将被使用，因为它导致脊髓可塑性与脊髓损伤动物呼吸输出的持续增加相关。同样重要的是，IH提供了一种工具，可以在移植物周围(可能是在移植物内部)引入或增加适当的图案。这项建议汇集了呼吸神经生理学、计算神经生物学、神经移植和脊髓损伤方面的独特专业知识，努力促进在理解和治疗脊髓损伤方面取得变革性的进展。 公共卫生相关性：呼吸损害是颈髓损伤后的一个重要问题。一种可能促进脊髓损伤后运动恢复的策略是“神经替代”疗法，即将细胞移植到脊髓损伤处。在这些实验中，我们将检验一种新的康复范例是否可以提高脊髓损伤后神经移植的有效性。

DESCRIPTION (provided by applicant): Neuronal replacement shows promise for enhancing recovery after spinal cord injury (SCI), and is considered a major objective for stem cell-mediated spinal cord repair. Achieving effective host-graft neuronal communication, however, represents a major challenge for neuronal replacement which has not been investigated. The overall hypothesis of this proposal is that presenting newly established host-graft neuronal networks with physiologically-patterned activities will enhance functional connectivity. Based on our extensive experience with SCI modeling and both respiratory and computational neurobiology, we are proposing to test this hypothesis by transplanting rat fetal spinal cord (FSC) tissue grafts - a source of neuronal progenitors - into clinically-relevant, high cervical (C3) contusion injuries in adult rats. Preliminary transneuronal neuroanatomical tracing data show synaptic connectivity between FSC donor cells and host phrenic motoneurons and cervical interneurons. In addition, FSC grafts receive extensive serotonergic inputs from the host, and some graft neurons exhibit hypoxia-sensitive discharge patterns including apparent inspiratory- related bursting. A prerequisite anatomical-functional framework is thus in place to test our central hypothesis via the following specific aims using FSC grafts placed into C3 contusion injuries in adult rats: Aim 1) to test the hypothesis that FSC-derived neurons will become anatomically and physiologically integrated with host gray matter, and Aim 2) to test the hypothesis that "training" via intermittent hypoxia (IH) stimulation will enhance the anatomical and physiological integration of FSC-derived neurons. To test these hypotheses, we will use a multi-disciplinary approach including neuroanatomical studies of connectivity between the graft and host spinal cord, measurement of breathing in awake rats, and neurophysiological studies of the graft. An innovative technical feature of our proposal is that this will be the first use of microelectrode arrays to monitor graft- associated neural ensembles in the spinal cord. A FSC grafting method will be used because such grafts develop into myelinated tissue containing a large contingent of cells resembling intermediate gray matter interneurons. A unique rehabilitative paradigm - daily exposure to mild, intermittent hypoxia (IH) - will be used because it leads to spinal cord plasticity associated with persistent increases in respiratory output in spinal injured animals. Equally important, IH provides a tool to introduce or increase appropriately patterned bursting around, and possibly within, the graft. This proposal brings together unique expertise in respiratory neurophysiology, computational neurobiology, neural transplantation, and SCI in an effort to facilitate transformative advances in the understanding and treatment of SCI. PUBLIC HEALTH RELEVANCE: Respiratory compromise is a significant problem after cervical spinal cord injury. A strategy that may enhance motor recovery after spinal injury is "neural replacement" therapy in which cells are transplanted into the spinal cord lesion. In these experiments, we will examine if a novel rehabilitation paradigm can enhance the effectiveness of a neural transplant following spinal cord injury.