Department of Neurology, University of California, Irvine, CA 92697‐4292, USA.
Exp Neurol. 2012 Dec;238(2):93-102. doi: 10.1016/j.expneurol.2012.08.004. Epub 2012 Aug 19.
Excitotoxicity (caused by over-activation of glutamate receptors) and inflammation both contribute to motor neuron (MN) damage in amyotrophic lateral sclerosis (ALS) and other diseases of the spinal cord. Microglial and astrocytic activation in these conditions results in release of inflammatory mediators, including the cytokine, tumor necrosis factor-alpha (TNF-α). TNF-α has complex effects on neurons, one of which is to trigger rapid membrane insertion of α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) type glutamate receptors, and in some cases, specific insertion of GluA2 lacking, Ca(2+) permeable AMPA receptors (Ca-perm AMPAr). In the present study, we use a histochemical stain based upon kainate stimulated uptake of cobalt ions ("Co(2+) labeling") to provide the first direct demonstration of the presence of substantial numbers of Ca-perm AMPAr in ventral horn MNs of adult rats under basal conditions. We further find that TNF-α exposure causes a rapid increase in the numbers of these receptors, via a phosphatidylinositol 3 kinase (PI3K) and protein kinase A (PKA) dependent mechanism. Finally, to assess the relevance of TNF-α to slow excitotoxic MN injury, we made use of organotypic spinal cord slice cultures. Co(2+) labeling revealed that MNs in these cultures possess Ca-perm AMPAr. Addition of either a low level of TNF-α, or of the glutamate uptake blocker, trans-pyrrolidine-2,4-dicarboxylic acid (PDC) to the cultures for 48 h resulted in little MN injury. However, when combined, TNF-α+PDC caused considerable MN degeneration, which was blocked by the AMPA/kainate receptor blocker, 2,3-Dihydroxy-6-nitro-7-sulfamoylbenzo (F) quinoxaline (NBQX), or the Ca-perm AMPAr selective blocker, 1-naphthyl acetylspermine (NASPM). Thus, these data support the idea that prolonged TNF-α elevation, as may be induced by glial activation, acts in part by increasing the numbers of Ca-perm AMPAr on MNs to enhance injurious excitotoxic effects of deficient astrocytic glutamate transport.
兴奋性毒性（由谷氨酸受体过度激活引起）和炎症都有助于肌萎缩侧索硬化症（ALS）和脊髓的其他疾病中的运动神经元（MN）损伤。在这些情况下，小胶质细胞和星形胶质细胞的激活导致炎症介质的释放，包括细胞因子肿瘤坏死因子-α（TNF-α）。TNF-α 对神经元有复杂的影响，其中之一是触发 α-氨基-3-羟基-5-甲基-4-异恶唑丙酸（AMPA）型谷氨酸受体的快速膜插入，在某些情况下，特异性插入缺乏 Ca2+通透性的 AMPA 受体（Ca-perm AMPAr）。在本研究中，我们使用基于 kainate 刺激钴离子摄取的组织化学染色（“Co(2+) 标记”），首次直接证明在基础条件下成年大鼠腹角 MN 中存在大量 Ca-perm AMPAr。我们进一步发现，TNF-α 暴露通过磷脂酰肌醇 3 激酶（PI3K）和蛋白激酶 A（PKA）依赖性机制导致这些受体数量的快速增加。最后，为了评估 TNF-α 与缓慢兴奋性 MN 损伤的相关性，我们利用器官型脊髓切片培养物。Co(2+) 标记显示这些培养物中的 MN 具有 Ca-perm AMPAr。向培养物中添加低水平的 TNF-α或谷氨酸摄取抑制剂反式吡咯烷-2,4-二羧酸（PDC）48 小时，导致 MN 损伤很小。然而，当两者结合时，TNF-α+PDC 导致相当数量的 MN 退化，这可被 AMPA/kainate 受体抑制剂 2,3-二羟基-6-硝基-7-磺酰胺基苯并（F）喹喔啉（NBQX）或 Ca-perm AMPAr 选择性抑制剂 1-萘乙酰基 spermine（NASPM）阻断。因此，这些数据支持这样一种观点，即持续的 TNF-α 升高，如由小胶质细胞激活引起的升高，部分通过增加 MN 上 Ca-perm AMPAr 的数量来增强星形胶质细胞谷氨酸转运不足的有害兴奋性毒性作用。
