Semi-quantitative analysis of α-synuclein in subcellular pools of rat brain neurons: An immunogold electron microscopic study using a C-terminal specific monoclonal antibody
Semi-quantitative analysis of α-synuclein in subcellular pools of rat brain neurons: An immunogold electron microscopic study using a C-terminal specific monoclonal antibody
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DOI：
10.1016/j.brainres.2008.08.067
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发表时间：
2008-12
期刊：
影响因子：
2.9
通讯作者：
Ling Zhang;Chunyan Zhang;Yuangang Zhu;Qing Cai;P. Chan;K. Uéda;Shun Yu;Hui Yang
中科院分区：
文献类型：
--
作者：
Ling Zhang;Chunyan Zhang;Yuangang Zhu;Qing Cai;P. Chan;K. Uéda;Shun Yu;Hui Yang
α-突触核蛋白（α-Syn）是一种由140个氨基酸组成的脑富集蛋白。尽管有强有力的证据表明该蛋白在几种神经退行性疾病的发病机制中的作用，但其生理功能仍然知之甚少。为了研究α-Syn的生理功能，必须对其进行精确的亚细胞定位。虽然α-Syn在脑中的表达已经使用几种不同的抗体进行了广泛的研究，但其在神经元中的精确亚细胞定位仍然是难以捉摸的。本研究采用免疫金电镜技术，用一种新制备的能识别α-Syn C端115-121个氨基酸的单克隆抗体3D 5，研究了α-Syn在大鼠脑神经元中的亚细胞定位。此外，在几个脑区的神经元的不同亚细胞池中的蛋白质的相对量进行了评估和比较。结果表明，α-Syn阳性的金颗粒在神经元的轴突、突触前终末、细胞质和细胞核中分布不均匀，其中突触前终末和细胞核中的金颗粒密度大于其他亚细胞池。在细胞质区域，在一些线粒体中看到相对致密的金颗粒。在同一亚细胞池中，不同脑区神经元的金颗粒密度不同。皮质神经元突触前终末和突触核内的金颗粒密度明显高于纹状体、海马和黑质神经元，但其线粒体内的金颗粒密度明显低于纹状体、海马和黑质神经元。海马、纹状体和黑质神经元线粒体α-Syn的相对高水平可能具有特殊的病理生理意义，值得进一步研究。
α-Synuclein (α-Syn) is a brain-enriched protein of 140 amino acids. Despite of strong evidence showing the implication of the protein in the pathogenesis of several neurodegenerative diseases, its physiological function remains poorly understood. To study the physiological function of α-Syn, a depiction of its precise subcellular localization is necessary. Although α-Syn expression in the brain has been extensively investigated using several different antibodies, its precise subcellular localization in neurons remains elusive. In this study, immunogold electron microscopy with a newly produced 3D5 monoclonal antibody recognizing the C-terminal 115–121 amino acids of α-Syn was used to examine its subcellular localization in rat brain neurons. In addition, the relative amount of the protein in different subcellular pools of the neurons in several brain regions was evaluated and compared. The results showed that α-Syn-positive gold particles were unevenly distributed in axons, presynaptic terminals, cytoplasm and nucleus in the neuron, with the density of gold particles being greater in presynaptic terminals and nucleus than in other subcellular pools. In the cytoplasmic region, relatively dense gold particles were seen in some mitochondria. In the same subcellular pools, the density of gold particles was varied among the neurons from different brain regions. Although the cortical neurons showed much higher density of gold particles in the presynaptic terminals and nuclei than in striatal, hippocampal and substantia nigral neurons, the density of gold particles in their mitochondria was much lower compared with the mitochondria of striatal, hippocampal and substantia nigral neurons. The relative high level of mitochondrial α-Syn in hippocampus, striatum and substantia nigral neurons may have special pathophysiological significance, which deserves further investigation.