Modulation of Phrenic Motoneuron Plasticity after Cervical Spinal Cord Injury

描述(申请人提供)：损伤的脊髓现在被认为具有强大的神经可塑性能力，希望通过治疗的方式利用这种能力来改善颈髓损伤(SCI)后的呼吸结果。康复和修复方法的基础是对脊柱呼吸回路和 慢性脊髓损伤后脊髓呼吸神经元的控制。原则上，损伤后的S脊髓实质上是一个“新脊髓”，其中影响到几乎所有功能领域的神经网络和控制机制都发生了显著变化。我们的研究小组最近对颈椎脊髓损伤后的脊柱呼吸回路进行了解剖学表征，但功能解剖相关性仍有待确定。我们提出了一系列的实验，这些实验将从神经生理学角度确定颈髓损伤后脊髓呼吸回路，检测关键神经调节剂5-羟色胺(5-HT)对该回路的影响，并确定一种有前景的脊髓移植方法对脊髓呼吸神经元和颈髓脊髓损伤后呼吸功能恢复的影响。本研究的总体假设是，慢性颈髓损伤后膈运动神经元(PMN)活动的调节受脊髓膈前中间神经元的影响，脊髓5-羟色胺是慢性脊髓损伤后脊髓呼吸系统的重要调节剂。高位颈椎SCI(侧位C2半切)的大鼠模型将被用来解决三个特定的目标。目标1将测试以下假设：慢性宫颈 脊髓损伤、中性粒细胞保持强大的可塑性，其爆裂模式受到部分调控 通过颈部的中间神经元。将使用多电极阵列测量PMN和颈部中间神经元的活性；将使用解剖学和免疫组织化学方法来评估脊髓呼吸回路。目的2将运用神经生理学、药理学、免疫化学和分子生物学技术，验证脊髓5-羟色胺受体激活是慢性颈髓损伤后膈神经运动恢复的重要组成部分的假说。最后，目标3将验证5-羟色胺能细胞移植可以增强或恢复脊髓呼吸神经元的5-羟色胺能调节，从而改善脊髓损伤后呼吸恢复的假设。C2半横断损伤后一周，成年大鼠将接受来自胎鼠中缝神经元的5-羟色胺能细胞脊髓内移植。移植物对膈运动系统的影响将通过行为、神经生理学、药理学、免疫组织化学和分子技术进行评估。这项建议将呼吸神经生理学、多单位记录方法、神经移植和颈椎脊髓损伤建模方面的独特和协同的专业知识结合在一起。创新方面包括：1)首次对慢性颈脊髓损伤后PMN爆发模式进行描述性和机械性研究；2)首次对颈SC后与呼吸相关的颈间神经元进行神经生理学研究；3)使用多阵列电极描述脊髓呼吸回路；以及4)首次使用移植策略来增强脊髓呼吸回路的5-羟色胺能神经支配。 公共卫生相关性：颈髓损伤对脊髓呼吸运动神经元(包括控制横隔膜的细胞)的神经控制的影响尚不清楚。我们提出了一系列实验来确定慢性颈脊髓损伤如何改变脊髓中呼吸运动神经元和中间神经元的神经调节。然后，这项工作将过渡到旨在通过以下方式促进呼吸恢复的翻译相关研究 将5-羟色胺能细胞移植到损伤的脊髓。

DESCRIPTION (provided by applicant): The injured spinal cord is now recognized to have a robust capacity for neuroplasticity, and it is desirable to therapeutically harness that in ways tht will enhance respiratory outcomes after cervical spinal cord injury (SCI). Fundamental to rehabilitation and repair approaches is a basic understanding of the spinal respiratory circuit and the control of spinal respiratory neurons after chronic SCI. In principle, the injured spinal cord s essentially a "new spinal cord" in which neural networks and control mechanisms affecting virtually every functional domain are significantly altered. Our group recently characterized the spinal respiratory circuit anatomically after cervical SCI, but functional-anatomical correlates remain to be determined. We propose a series of experiments which will neurophysiologically define the spinal respiratory circuit after cervical SCI, examine the influence of a key neuromodulator - serotonin (5-HT) - on the circuit, and determine the impact of a promising spinal cord transplantation approach on spinal respiratory neurons and recovery of ventilation after cervical SCI. The overall hypothesis guiding this proposal is that the regulation of phrenic motoneuron (PMN) activity following chronic cervical SCI is influenced by spinal pre-phrenic interneurons and that spinal 5-HT is a critically important modulator of the spinal respiratory circuitry following chronic SCI. A rat model of high cervical SCI (lateral C2 hemisection) will be used to address three specific aims. Aim 1 will test the hypothesis that following chronic cervical SCI, PMNs retain a robust capacity for plasticity, and their bursting patterns are partly regulated by cervical interneurons. PMN and cervical interneuron activity will be measured using a multi- electrode array; anatomical and immunohistochemical methods will be used to evaluate the spinal respiratory circuit. Aim 2 will use neurophysiological, pharmacological, immunochemical, and molecular techniques to test the hypothesis that spinal 5-HT receptor activation is an integral part of phrenic motor recovery after chronic cervical SCI. Lastly, Aim 3 will test the hypothesis that transplantation of serotonergic cells can enhance or restore serotonergic modulation of spinal respiratory neurons thereby improving respiratory recovery after SCI. One week following C2 hemisection injury, adult rats will receive an intraspinal transplant of serotonergic cells derived from fetal rat raph¿ neurons. The impact of the grafts on the phrenic motor system will be assessed using behavioral, neurophysiological, pharmacological, immunohistochemical, and molecular techniques. This proposal brings together a unique and synergistic combination of expertise in respiratory neurophysiology, multi-unit recording approaches, neural transplantation, and cervical SCI modeling. Innovative aspects include: 1) the first descriptive and mechanistic studies of PMN burst patterns after chronic cervical SCI; 2) the first neurophysiological studies of respiratory-related cervical interneurons after cervical SC; 3) the use of multi- array electrodes to describe the spinal respiratory circuitry, and 4) the firs use of transplant strategies to enhance serotonergic innervation of the spinal respiratory circuit. PUBLIC HEALTH RELEVANCE: The impact of cervical spinal cord injury on the neural control of spinal respiratory motoneurons - including the cells which control the diaphragm - is unclear. We propose a series of experiments to determine how chronic cervical spinal cord injury alters the neural regulation of respiratory motoneurons and interneurons in the spinal cord. The work will then transition to translationally relevant studies aimed at enhancing respiratory recovery by transplanting serotonergic cells into the injured spinal cord.