Mechanisms of Central Synaptic Dysfunction in SMA

描述（由申请人提供）：脊髓性肌萎缩症（SMA）是一种遗传性神经退行性疾病，以运动神经元丧失和骨骼肌萎缩为特征。SMA是婴儿死亡最常见的遗传原因，但目前尚无有效的治疗方法。虽然对这种疾病的遗传原因了解很多，但对解释受影响个体表现出的运动症状严重程度的生理变化的信息却很少。特定的、脆弱的神经元群的功能障碍可能导致神经回路的继发性变化，从而加剧神经元功能障碍。在脊髓中，运动神经元接受来自局部中间神经元、来自大脑的下行通路和感觉神经元的直接突触输入。在之前的一项研究中，我们报道了SMA小鼠感觉初级传入神经和运动神经元之间的单突触连接强度在疾病过程的早期，在检测到大量运动神经元细胞损失之前大大降低。在SMA小鼠中，这种功能的丧失部分是由运动神经元上初级传入按钮的丧失介导的。本研究的目的是确定运动回路的哪些输入或部分特别受到疾病的影响，以及这是由于运动神经元功能障碍还是这些神经元回路中SMN缺乏的内在原因。在Aim 1中，我们将分析在SMA小鼠的脊髓运动神经元的躯体和树突上直接形成突触的神经元群的功能影响。此外，我们将通过绘制和量化SMA小鼠运动神经元上的突触密度来关联功能和结构缺陷。这些研究将纵向延伸，以确定疾病过程中缺陷的时间进程。在Aim 2中，我们将使用利用Cre-lox技术的新型小鼠来研究SMN蛋白调控在逆转疾病严重表型中的作用。我们将采用行为、生理和形态分析来确定这些方法的有效性。激光捕获显微解剖也将用于从对照和SMA小鼠中分离选定的与疾病相关的神经元类型，用于RNA分析。这些将包括腰椎前角的运动神经元和其他一些神经元和非神经元群。在目标3中，我们将研究SMA中受影响的运动回路中突触丢失的机制。我们将采用免疫组织化学标记，通过比较SMA和野生型脊髓，确定疾病不同阶段受影响的突触的起源。总的来说，这些实验有可能阐明突触缺陷在SMA小鼠疾病进展中的重要性。

DESCRIPTION (provided by applicant): Spinal muscular atrophy (SMA) is an inherited neurodegenerative disease characterized by motor neuron loss and skeletal muscle atrophy. SMA is the most common genetic cause of death in infancy, but no effective treatment is currently available. While much is known about the genetic causes of the disease, less information is available on the physiological alterations that explain the severity of motor symptoms displayed by affected individuals. Dysfunction of specific, vulnerable neuronal populations may precipitate secondary changes in neural circuits that could exacerbate neuronal dysfunction. In the spinal cord, motor neurons receive direct synaptic inputs from local interneurons, descending pathways from the brain, and sensory neurons. In a previous study we reported that the strength of monosynaptic connections between sensory primary afferents and motor neurons in SMA mice is greatly reduced early in the course of the disease, before substantial motor neuron cell loss can be detected. This loss of function is mediated in part by the loss of primary afferent boutons on motor neurons in SMA mice. The goals of this study is to identify which inputs or parts of the motor circuit are particularly affected by the disease and whether this is due to motor neuron dysfunction or intrinsic to SMN deficiency in these neuronal circuits. In Aim 1, we will analyze the functional effects of a neuronal population that makes direct synapses on the somata and dendrites of spinal motor neurons in SMA mice. In addition we will correlate functional and structural defects by mapping and quantifying the synaptic density on motor neurons in SMA mice. These studies will extend longitudinally to determine the time course of the defects in the course of the disease. In Aim 2, we will use novel mice taking advantage of the Cre-lox technology to study the effects of regulation of SMN protein in reversing the severe phenotype of the disease. We will employ behavioral, physiological and morphological assays to determine efficacy of these approaches. Laser capture microdissection will also be employed to isolate selected, disease-relevant neuronal types from control and SMA mice for RNA analysis. These will include motor neurons in the ventral horns of the lumbar spinal cord and several other neuronal and non-neuronal populations. In Aim 3, we will investigate mechanisms involved in synaptic loss in the motor circuits affected in SMA. We will employ immunohistochemical markers to identify the origin of the synapses affected at different stages of the disease by comparing SMA and wild type spinal cords. Collectively, these experiments have the potential to elucidate the importance of synaptic defects in the progression of the disease in SMA mice. PUBLIC HEALTH RELEVANCE: SMA is an incurable motor neuron disease and the leading genetic cause of death in infancy. We will establish whether central synaptic defects are causally involved in the severe phenotype in a mouse model of SMA, employing behavioral, physiological and morphological assays. Different mechanisms of synaptic elimination will be investigated by utilizing novel transgenic mouse models and determine their potential beneficiary effects in reversing the SMA phenotype.