RNA editing of AMPA receptor subunit GluR2 in ischemia

缺血性中风是发达国家的第三大死亡原因。这种疾病的一个关键特征是 一种高度选择性的神经元缺失模式;某些可识别的神经元子集，特别是CA 1 海马体中的锥体神经元严重受损，而其他神经元保持完整。这一步 选择性神经元损伤涉及Ca 2+通过Ca 2+可渗透的AMPA受体通道进入。AMPA 受体是谷氨酸受体（GluR）的主要亚型，其由GluR 1 -4亚基组装而成。 通道的Ca 2+渗透性由Q/R位点的GluR 2 RNA编辑主导;编辑的GluR 2（R） 亚基形成Ca 2+不可渗透的通道，而未编辑的GluR 2（Q）通道允许Ca 2+进入。在 大多数CA 1神经元的AMPA受体通道含有GluR 2（R），因此对Ca 2+流是不可渗透的。 最近，我们已经确定短暂的前脑缺血选择性地破坏GluR 2 Q/R位点编辑， 因此诱导损伤性Ca 2+通过AMPA受体通道进入脆弱的CA 1神经元。我们 还表明受损的GluR 2 Q/R位点编辑与以下基因的表达减少密切相关： ADAR 2（adenosine deaminase acting on RNA的缩写）基因，一种负责GluR 2 Q/R的核酶 网站编辑因此，我们假设ADAR 2基因表达的减少是导致受损的 GluR 2 Q/R位点编辑。为了直接解决这一假设，我们将确定ADAR 2基因的恢复是否 表达拯救GluR 2 Q/R位点编辑，进而阻断AMPA受体的Ca 2+渗透性 通道，导致缺血后大鼠中脆弱神经元的存活。总体而言，该项目将 有两个具体目标： 具体目的1：确定ADAR 2基因表达的恢复是否阻断Ca 2+通过 AMPA受体通道和拯救缺血后大鼠的脆弱神经元。 具体目的2：确定稳定ADAR 2基因沉默的产生是否诱导细胞变性。 缺血不敏感的神经元，如果ADAR 2缺陷神经元的变性是由RNA编辑引起的， 缺乏一个或多个谷氨酸受体亚单位。 总之，该项目将确定ADAR 2依赖性GluR 2 Q/R位点编辑决定了 神经元缺血。因此，这项工作将为斯托克治疗确定一个有希望的靶点。

Ischemic stroke is the third leading cause of death in developed countries. A critical feature of the disease is a highly selective pattern of neuronal loss; certain identifiable subsets of neurons, particularly CA1 pyramidal neurons in the hippocampus, are severely damaged while others remain intact. A step in this selective neuronal injury involves Ca2+ entry through Ca2+-permeable AMPA receptor channels. AMPA receptors are a major subtype of glutamate receptors (GluRs) that are assembled from GluR1-4 subunits. Ca2+ permeability of the channels is dominated by GluR2 RNA editing at the Q/R site; edited GluR2(R) subunits form Ca2+-impermeable channels, whereas unedited GluR2(Q) channels allow Ca2+ entry. In most CA1 neurons, AMPA receptor channels contain GluR2(R), and thus are impermeable to Ca2+ flow. Recently, we have identified that transient forebrain ischemia selectively disrupts GluR2 Q/R site editing and hence induces injurious Ca2+ entry through AMPA receptor channels into vulnerable CA1 neurons. We have also shown that impaired GluR2 Q/R site editing is closely correlated with reduced expression of ADAR2 (short for adenosine deaminase acting on RNA) gene, a nuclear enzyme responsible for GluR2 Q/R site editing. We thus hypothesize that reduced expression of ADAR2 gene is responsible for the impaired GluR2 Q/R site editing. To address this hypothesis directly, we will determine if restoration of ADAR2 gene expression rescues GluR2 Q/R site editing and in turn blocks Ca2+ permeability of AMPA receptor channels, leading to the survival of vulnerable neurons in the post-ischemic rats. Overall, this project will have two specific aims: Specific Aim 1: To determine whether restoration of ADAR2 gene expression blocks Ca2+ entry through AMPA receptor channels and rescues vulnerable neurons in the post-ischemic rats. Specific Aim 2: To determine if generation of stable ADAR2 gene silencing induces degeneration of ischemia-insensitive neurons, and if degeneration of ADAR2-deficient neurons results from RNA editing deficits of one or more glutamate receptor subunits. Together, this project will identify that ADAR2-dependent GluR2 Q/R site editing determines vulnerability of neurons to ischemia. Thus, this work will define a promising target for stoke therapy.