Establishment of in vitro model for the vomeronasal system and analysis of the synapse formation between the vomeronasal and accessory olfactory neurons

犁鼻系统体外模型的建立及犁鼻与副嗅神经元突触形成的分析

基本信息

批准号：
16500201
负责人：
MURAMOTO Kazuyo
金额：
$ 2.05万
依托单位：
Kochi University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-16500201/
关键词：
co-culture vomeronasal organ calcium imaging synapse formation 鋤鼻系 Calcium Imaging

项目摘要

Pheromones are detected by unique receptors in the vomeronasal organ (VNO), and then affect endocrinal status and behavior in various kinds of animals. Although it is well known that the VNO mediates sexually and developmentally different behavioral and neuroendocrine responses, the cellular mechanisms regulating these functions are not, much understood. The differences in pheromonal effects might be in part due to differential expression of pheromone receptors. In this project, I established the co-culture system of vomeronasal receptor neurons (VRN) and accessory olfactory bulb (AOB) neurons to assess roles of each neuronal species in the vomeronasal system on the pheromonal signal processing. Firstly, I demonstrated the synapse formation between cultured VRNs and AOB neurons using a calcium imaging technique combined with electrical stimulation and analyzing pharmacologically. Secondly, it was showed that the maturation of VRNs was induced by co-culture with dissociated AOB neurons. … More To characterize the receptor expression, I examined two representatives (VR1 and VR4) from different subfamilies of the V2R family of pheromone receptors in cultured VRNs. A western blotting analysis showed the expression of VR1 and VR4 in VRNs was induced and increased with days in co-culture, an effect which was not observed in the VRN-alone culture. Moreover, we applied charged compounds in mouse urine iontophoretically to the VRNs using a microelectrode and analyzed VRN response by a Ca^<2+> imaging method with or without cultured AOB cells. When urine compounds were ejected to the VRNs co-cultured with AOB cells with a current of -2μA, subpopulation of VRNs clearly showed long-lasting Ca^<2+> increases. Injections of a current below -5μA alone had no effect to the VRNs. Such Ca^<2+> increases were not observed without AOB cells. These results indicate that VRNs result in expressing pheromone receptors by interacting with AOB neurons, and then acquiring responsiveness to compounds in urine. Less

信息素由犁鼻器 (VNO) 中的独特受体检测，然后检测各种动物的内分泌状态和行为，尽管众所周知，VNO 介导性和发育上不同的行为和神经内分泌反应，但调节这些功能的细胞机制。信息素效应的差异可能部分归因于信息素受体的差异表达。在这个项目中，我建立了犁鼻的共培养系统。首先，我使用钙成像技术结合培养的 VRN 和 AOB 神经元来演示培养的 VRN 和 AOB 神经元之间的突触形成。其次，研究表明 VRN 的成熟是通过与分离的 AOB 神经元共培养来诱导的。为了表征受体表达，我检查了两个。来自培养的 VRN 中信息素受体 V2R 家族不同亚科的代表（VR1 和 VR4）。蛋白质印迹分析显示 VRN 中 VR1 和 VR4 的表达随着共培养天数的增加而被诱导并增加，但未观察到这种效应。此外，我们使用微电极将小鼠尿液中的带电化合物离子电渗至VRN，并通过Ca 2+ 分析VRN反应。使用或不使用培养的AOB细胞的成像方法，当以-2μA的电流将化合物尿液喷射到与AOB细胞共培养的VRN时，VRN的亚群清楚地显示电流的持久Ca 2+ 注射增加。在没有AOB细胞的情况下，单独低于-5μA对VRN没有影响。这些结果表明VRN通过与AOB相互作用而导致表达信息素受体。神经元，然后对尿液中的化合物产生反应。