Ouardouz Mohamed, Coderre Elaine, Basak Ajoy, Chen Andrew, Zamponi Gerald W, Hameed Shameed, Rehak Renata, Yin Xinghua, Trapp Bruce D, Stys Peter K
Ottawa Health Research Institute, University of Ottawa, Ontario, Canada.
Ann Neurol. 2009 Feb;65(2):151-9. doi: 10.1002/ana.21533.
The deleterious effects of glutamate excitotoxicity are well described for central nervous system gray matter. Although overactivation of glutamate receptors also contributes to axonal injury, the mechanisms are poorly understood. Our goal was to elucidate the mechanisms of kainate receptor-dependent axonal Ca(2+) deregulation.
Dorsal column axons were loaded with a Ca(2+) indicator and imaged in vitro using confocal laser-scanning microscopy.
Activation of glutamate receptor 6 (GluR6) kainate receptors promoted a substantial increase in axonal [Ca(2+)]. This Ca(2+) accumulation was due not only to influx from the extracellular space, but a significant component originated from ryanodine-dependent intracellular stores, which, in turn, depended on activation of L-type Ca(2+) channels: ryanodine, nimodipine, or nifedipine blocked the agonist-induced Ca(2+) increase. Also, GluR6 stimulation induced intraaxonal production of nitric oxide (NO), which greatly enhanced the Ca(2+) response: quenching of NO with intraaxonal (but not extracellular) scavengers, or inhibition of neuronal NO synthase with intraaxonal Nomega-nitro-L-arginine methyl ester, blocked the Ca(2+) increase. Loading axons with a peptide that mimics the C-terminal PDZ binding sequence of GluR6, thus interfering with the coupling of GluR6 to downstream effectors, greatly reduced the agonist-induced axonal Ca(2+) increase. Immunohistochemistry showed GluR6/7 clusters on the axolemma colocalized with neuronal NO synthase and Ca(v)1.2.
Myelinated spinal axons express functional GluR6-containing kainate receptors, forming part of novel signaling complexes reminiscent of postsynaptic membranes of glutamatergic synapses. The ability of such axonal "nanocomplexes" to release toxic amounts of Ca(2+) may represent a key mechanism of axonal degeneration in disorders such as multiple sclerosis where abnormal accumulation of glutamate and NO are known to occur.
谷氨酸兴奋性毒性对中枢神经系统灰质的有害影响已有充分描述。尽管谷氨酸受体的过度激活也会导致轴突损伤，但其机制尚不清楚。我们的目标是阐明红藻氨酸受体依赖性轴突Ca(2+)失调的机制。
用Ca(2+)指示剂加载背柱轴突，并在体外使用共聚焦激光扫描显微镜成像。
谷氨酸受体6（GluR6）红藻氨酸受体的激活促进了轴突[Ca(2+)]的显著增加。这种Ca(2+)积累不仅是由于细胞外空间的流入，而且很大一部分来自于ryanodine依赖性细胞内储存，而这又依赖于L型Ca(2+)通道的激活：ryanodine、尼莫地平或硝苯地平可阻断激动剂诱导的Ca(2+)增加。此外，GluR6刺激诱导轴突内一氧化氮（NO）的产生，这极大地增强了Ca(2+)反应：用轴突内（而非细胞外）清除剂淬灭NO，或用轴突内Nω-硝基-L-精氨酸甲酯抑制神经元型一氧化氮合酶，均可阻断Ca(2+)增加。用模拟GluR6 C末端PDZ结合序列的肽加载轴突，从而干扰GluR6与下游效应器的偶联，可大大减少激动剂诱导的轴突Ca(2+)增加。免疫组织化学显示轴膜上的GluR6/7簇与神经元型一氧化氮合酶和Ca(v)1.2共定位。
有髓脊髓轴突表达功能性含GluR6的红藻氨酸受体，形成了类似于谷氨酸能突触后膜的新型信号复合物的一部分。这种轴突“纳米复合物”释放有毒量Ca(2+)的能力可能是轴突变性的关键机制，在诸如多发性硬化症等疾病中，已知会发生谷氨酸和NO的异常积累。
