Huang Q, Zhou D, Sapp E, Aizawa H, Ge P, Bird E D, Vonsattel J P, DiFiglia M
Laboratory of Cellular Neurobiology, Massachusetts General Hospital, Boston 02114, USA.
Neuroscience. 1995 Mar;65(2):397-407. doi: 10.1016/0306-4522(94)00494-p.
In Huntington's disease striatal neurons undergo marked changes in dendritic morphology and coincidently exhibit an increase in immunoreactive calbindin D28k (calbindin), a cytosolic calcium-binding protein which is highly abundant in these neurons. Previous studies in the rat striatum have shown that excitotoxic injury, which is linked to a rise in intracellular Ca2+, mimics many of the neurochemical and neuropathological characteristics of Huntington's disease. We speculated, therefore, that the apparent increase in calbindin labeling in Huntington's disease spiny neurons may signal the response to an excitotoxic process. To investigate this possibility, we compared the cellular features of calbindin immunoreactivity in grade 1-4 Huntington's disease cases with those seen in rat striatal neurons in vivo and in vitro following treatment with N-methyl-D-aspartate (NMDA) receptor agonist, quinolinic acid. In human post mortem control cases calbindin immunoreactivity was seen primarily in the somata and proximal dendrites of striatal neurons. In the Huntington's disease cases, calbindin labeling was markedly increased throughout the second and third order dendrites and in spines, and this change was more prevalent in advanced cases (grades 3-4). In the rat brain, two weeks after intrastriatal injection of quinolinic acid (6-20 ng), surviving medium-spiny neurons in the transition zone around the lesion core exhibited a marked increase in calbindin immunoreactivity similar to that seen in Huntington's disease spiny neurons. In more peripheral areas away from the lesion and on the contralateral unlesioned side, calbindin immunostaining was confirmed to somata and proximal dendrites. In situ hybridization histochemistry with an 35S-labeled oligonucleotide probe showed no change or a decrease in calbindin mRNA levels in neurons within the transition zone, suggesting that the observed increase in calbindin staining was not the result of increased transcription. In 12 day old postnatal striatal cultures, 2-6 h exposures to quinolinic acid (0.5 mM) significantly increased the length of neurites exhibiting calbindin immunoreactivity when compared to untreated controls. This effect was blocked by the selective NMDA receptor blocker (+/-)-2-amino-5-phosphonopentanoic acid (AP-5), indicating that an NMDA receptor-mediated mechanism contributed to the change in staining pattern. Results in rats suggest that the subcellular redistribution of calbindin immunoreactivity observed in Huntington's disease spiny neurons may be related to an NMDA receptor-induced excitotoxic process. An increased availability of calbindin protein at dendrites and spines may reflect a greater demand for Ca2+ buffering precipitated by an abnormal rise in in intracellular Ca2+.
在亨廷顿舞蹈症中，纹状体神经元的树突形态会发生显著变化，同时，一种细胞溶质钙结合蛋白——免疫反应性钙结合蛋白D28k（钙结合蛋白）的表达增加，该蛋白在这些神经元中含量很高。先前对大鼠纹状体的研究表明，与细胞内Ca2+升高有关的兴奋性毒性损伤模拟了亨廷顿舞蹈症的许多神经化学和神经病理学特征。因此，我们推测，亨廷顿舞蹈症棘状神经元中钙结合蛋白标记的明显增加可能预示着对兴奋性毒性过程的反应。为了探究这种可能性，我们将1-4级亨廷顿舞蹈症病例中钙结合蛋白免疫反应性的细胞特征与用N-甲基-D-天冬氨酸（NMDA）受体激动剂喹啉酸处理的大鼠纹状体神经元在体内和体外的细胞特征进行了比较。在人类死后对照病例中，钙结合蛋白免疫反应性主要见于纹状体神经元的胞体和近端树突。在亨廷顿舞蹈症病例中，钙结合蛋白标记在二级和三级树突以及棘中显著增加，并且这种变化在晚期病例（3-4级）中更为普遍。在大鼠脑中，纹状体内注射喹啉酸（6-20 ng）两周后，病变核心周围过渡区存活的中等棘状神经元的钙结合蛋白免疫反应性显著增加，类似于在亨廷顿舞蹈症棘状神经元中观察到的情况。在远离病变的更外周区域以及对侧未病变侧，钙结合蛋白免疫染色定位于胞体和近端树突。用35S标记的寡核苷酸探针进行原位杂交组织化学显示，过渡区内神经元的钙结合蛋白mRNA水平没有变化或下降，这表明观察到的钙结合蛋白染色增加不是转录增加的结果。在出生后12天龄的纹状体培养物中，与未处理的对照相比，暴露于喹啉酸（0.5 mM）2-6小时可显著增加显示钙结合蛋白免疫反应性的神经突长度。这种作用被选择性NMDA受体阻滞剂（+/-）-2-氨基-5-膦酸戊酸（AP-5）阻断，表明NMDA受体介导的机制促成了染色模式的变化。大鼠实验结果表明，在亨廷顿舞蹈症棘状神经元中观察到的钙结合蛋白免疫反应性的亚细胞重新分布可能与NMDA受体诱导的兴奋性毒性过程有关。树突和棘处钙结合蛋白的可用性增加可能反映了细胞内Ca2+异常升高引发的对Ca2+缓冲的更大需求。
