AMPA RECEPTOR EXPRESSION AND SELECTIVE NEURONAL DEATH

选择性破坏某些神经元标志着许多神经系统疾病，包括小脑退行性变和运动神经元疾病，如ALS。大量证据表明，这些疾病的退行性变可能是由于兴奋性毒性引起的，其中谷氨酸受体的过度激活导致神经元损伤。此外，最好的模型表明，在这些疾病中破坏的两种神经元类型--小脑浦肯野细胞和脊髓运动神经元--的损害是由谷氨酸受体的AMPA亚型介导的。本项目将通过在分子水平上将AMPA受体的特殊表达模式与与毒性相关的功能受体特性联系起来，来研究这些神经元选择性易损性的机制。特别是，相对抵抗脱敏的AMPA受体的表达，或对钙离子具有较高渗透性的受体的表达，可能使这些细胞选择性地容易受到谷氨酸毒性的影响。这些问题将在以下特定目的中进行研究：1结合膜片钳电生理研究，结合单细胞PCR扩增，评估浦肯野神经元AMPA受体的功能特性是否由AMPA亚单位mRNA表达模式决定。2.探讨Purkinje细胞表达的AMPA受体的脱敏和钙通透性与其选择性易感性的关系。3.明确AMPA受体的功能特性及其在运动神经元中的分子表达模式。同样，我们将使用单细胞聚合酶链式反应和膜片钳技术研究亚基表达对细胞脱敏和钙离子通透性的影响。4.检测在器官型脊髓切片培养中脊髓运动神经元选择性易损性的受体亚型，确定导致选择性易损性的AMPA受体的特性。了解AMPA受体的特性如何导致浦肯野细胞和脊髓运动神经元的选择性损伤，可能是了解其退变机制和识别小脑退行性变和ALS神经保护的最佳分子靶点的关键。

Selective destruction of certain neurons marks many of the neurological diseases, including the cerebellar degenerations and motor neuron diseases such as ALS. Considerable evidence suggests that the degeneration in these diseases may result from excitotoxicity, in which excessive activation of glutamate receptors leads to neuronal damage. Furthermore, the best models suggest that the damage to the two neuronal types destroyed in these diseases, cerebellar Purkinje cells and spinal motor neurons, is mediated by the AMPA subtype of glutamate receptors. This project will study the mechanism of the selective vulnerability of these neurons by relating their particular patterns of AMPA receptor expression at the molecular level to functional receptor properties relevant to toxicity. In particular, expression of AMPA receptors that are relatively resistant to desensitization, or of receptors that have higher permeability to Ca2+ may render these cells selectively vulnerable to glutamate toxicity. These issues will be investigated in the following specific aims: 1 To assess whether the functional properties of AMPA receptors in Purkinje neurons are determined by the AMPA subunit mRNA expression pattern using single cell PCR amplification in conjunction with patch-clamp electrophysiological studies. 2. To explore the relationship of the desensitization and Ca2+ permeability of AMPA receptors expressed in Purkinje cells to their selective vulnerability in electrophysiological studies correlated with specific toxicity in culture. 3. To define the functional properties of AMPA receptors and their molecular expression pattern of motor neurons. Again, the determination of desensitization and Ca2+ permeability by subunit expression will be studied using single cell PCR and patch-clamp studies. 4. To examine the receptor subtypes responsible for selective vulnerability in spinal motor neurons in organotypic spinal cord slice cultures, identifying the properties of AMPA receptors responsible for selective vulnerability. Understanding how the properties of AMPA receptors can lead to selective damage to Purkinje cells and spinal motor neurons may hold the key to understanding the mechanisms of their degeneration and to recognition of the best molecular targets for neuroprotection in cerebellar degenerations and ALS.