Analysis of molecular mechanisms for neurogenesis in the mammalian olfactory sensory epithelium

哺乳动物嗅觉上皮神经发生的分子机制分析

• 批准号: 13680863
• 负责人: YAMAGUCHI Masahiro
• 金额: $ 2.3万
• 依托单位: The University of Tokyo
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13680863/
• 关键词: neurogenesis; vomeronasal organ; nestin; GFP

Neurons are continuously generated in the mammalian olfactory sensory epithelium even in the adult period. To examine the cellular and molecular mechanism of adult neurogensis, neurogenesis in the olfactory epithelium was enhanced by the unilateral removal of the olfactory bulb (bulbeclomy) in the adult mice, and the newly-generated neurons were followed by bromodeoxyuridine (BrdU) labeling. Because the enhancement of neurogenesis was more prominent in the vomeronasal organ (VNO) specified for pheromone sensation rather than in the main olfactory epithelium for odorant detection, VNO was intensively examined in this study. In the VNO of control side, BrdU labeled ceils were sparsely located at the basal layer. In contrast, in the operated side, many BrdU labeled cells appeared throughout the basal layer, which indicated that most stem cells are quiescent but can be stimulated to proliferate with bulbectomy. Within several days BrdU labeled cells migrated to superficial layers and expressed neuronal marker proteins. There are two subtypes in the sensory neurons in VNO, the Go positive and the Gi positive neurons. BrdU labeling experiments showed that new neurons were at first Go positive immature cells and then differentiated into Go positive and Gi positive neurons, indicating that two types of neurons in the VNO originate from common Go positive progenitors. To analyze the molecular mechanism of olfactory neurogenesis in detail, nestin promoter-EGFP transgenic mice were utilized, where newly-generated cells can be visualized by GFP fluorescence. GFP expressing cells in VNO were found to be immature progenitors from their marker protein expression. The sensory epithelium of the transgenic mice was stripped, the cells were dispersed, and the GFP expressing cells were successfully recovered under fluorescent microscopy. This system was considered to be useful to further examine the molecular mechanism of olfactory neurogenesis in vitro.

即使在成年时期，神经元在哺乳动物嗅觉感觉上皮也会不断产生。为了检查成年神经血集的细胞和分子机制，通过单方面去除嗅球（在成年小鼠中）的单方面去除，从而增强了嗅觉上皮的神经发生，并在成年小鼠中的单方面去除，新生成的神经元之后是溴脱氧尿苷（BRDU）的lib。由于神经发生的增强在针对信息素感觉而不是用于气味检测的主要嗅觉上皮中的呕吐器官（VNO）中更为突出，因此在这项研究中对VNO进行了深入研究。在控制侧的VNO中，BRDU标记的天花板位于基础层。相比之下，在操作的一侧，许多BRDU标记的细胞出现在整个基础层，这表明大多数干细胞是静止的，但可以刺激以通过囊状切除术刺激。几天之内，BRDU标记的细胞迁移到表面层和表达的神经元标记蛋白。 VNO的感觉神经元中有两个亚型，GO阳性和GI阳性神经元。 BRDU标记实验表明，新的神经元起初是阳性未成熟细胞的，然后分化为阳性和胃肠道阳性神经元，表明VNO中的两种神经元来自普通的GO阳性祖细胞。为了详细分析嗅觉神经发生的分子机制，利用了Nestin启动子-EGFP转基因小鼠，其中可以通过GFP荧光可视化新生成的细胞。发现VNO中表达细胞的GFP是从其标记蛋白表达中的不成熟祖细胞。剥去了转基因小鼠的感觉上皮，分散细胞，并在荧光显微镜下成功回收了GFP表达细胞。该系统被认为可用于进一步检查体外嗅觉神经发生的分子机制。