Development and Aging of Neuronal Synapse

神经元突触的发育和衰老

基本信息

批准号：
09480219
负责人：
SHIRAO Tomoaki
金额：
$ 8.38万
依托单位：
Gunma University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1997
资助国家：
日本
起止时间：
1997 至 1999
项目状态：
已结题

项目摘要

(A) Analysis of spine cytoskeletonsWe prepare the cytoskeletal protein complex in the dendritic spine using the bead bound to anti-drebrin monoclonal antibody M2F6. The prepared sample contained actin, myosin I, myosin II, myosin V and gelsolin in addition to drebrin itself. Further, there are many other unidentified proteins in that sample ; therefore, we raised monoclonal antibodies using this sample as an immunogen. As a consequence we raised anti-myosin antibody and anti-CaMKII antibody. The anti-myosin antibody specifically recognized non-muscle myosin II heavy chan, which is present in cell soma and dendritic shaft in addition to dendritic spine. This indicates that spine cytoskelton is characterized by actin binding proteins but not by myosins.(B) The change of spine cytoskeletons during development and aging.In the developing brain, drebrin E is observed in the migrating neurons and within the growing axons and dendrite. In the adult brain, drebrin A is localized in the dendrit … More ic spines. In this study we investigated when drebrin disappeared from cell soma and enriched in dendritic spines during development. Immunohistochemical staining of the cerebrum and cerebellum using anti-drebrin monoclonal antibody (MBL, Japan) demonstrated that drebrin immunoreactivity was observed in the cell soma and dendrites from postnatal 0 day to 10 day, although drebrin expression as a whole was decreased according to the postnatal days. However, in 14-day-old rat, drebrin immunoreactivity was only observed as dot-like pattern in neuropil. Drebrin immunoreactivity was hardly observed within the cell soma and dendrites. These data indicates that drebrin subcellular localization was changed synchronously around postnatal 12 days, indicating that the final maturation of neuronal cytoskeleton may occurred at postnatal 12 days. Application of 100 μM glutamate weakened drebrin immunoreactivity at dendritic spines. Transfection experiments demonstrated that overexpression of drebrin A in neuron resulted the elongation of the spine length. These data indicate that developmental changes in the distribution of drebrin from dendritic shafts into dendritic spines may be related to the maturation of synaptic function. Less

（a）分析脊柱细胞骨骼，使用与抗蛋白抗蛋白单克隆抗体M2F6结合的珠子在树突状脊柱中制备细胞骨架蛋白复合物。准备好的样品还含有肌动蛋白，肌球蛋白I，肌球蛋白II，肌球蛋白V和凝胶蛋白，除了Drebrin本身。此外，该样品中还有许多其他未识别的蛋白质。因此，我们使用该样品作为免疫原提高了单克隆抗体。结果，我们提出了抗肌球蛋白抗体和抗CAMKII抗体。抗肌球蛋白抗体特异性识别非肌肉肌球蛋白II重木，除树突状脊柱外，还存在于细胞体和树突状轴中。这表明脊柱细胞骨架的特征是肌动蛋白结合蛋白，而不是肌动物。（b）在发育和衰老过程中脊柱细胞骨架的变化。在发育中的大脑中，在迁移的神经元以及生长的轴突中观察到drebrin e。在成年大脑中，Drebrin A位于树突中……更多的IC刺。在这项研究中，我们调查了Drebrin从细胞体内消失并在发育过程中富集在树突状刺中的研究。使用抗蛋白酶单克隆抗体（MBL，日本）对脑和小脑进行免疫组织化学染色表明，在drebrin表达的日期至10天的细胞体内和树突中观察到了Drebrin免疫反应性，但根据POSTNATAL的研究，DREBRIN表现为整个天。然而，在14天大的大鼠中，仅在神经皮质中观察到德雷布林免疫反应性。在细胞体和树突中，几乎没有观察到Drebrin免疫反应性。这些数据表明，在产后12天左右，Drebrin亚细胞定位是同步更改的，这表明神经元细胞骨架的最终成熟可能在产后12天发生。 100μM谷氨酸的应用削弱了树突状棘的DREBRIN免疫反应性。转染实验表明，神经元中Drebrin A的过表达导致脊柱长度的伸长。这些数据表明，从树突轴向树突状棘中Drebrin的分布的发展变化可能与突触功能的成熟有关。较少的