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Observation of the motion of a single molecule of neuronal nitric oxide synthase in the nucleus tractus solitarius

孤束核神经元一氧化氮合酶单分子运动观察

基本信息

批准号：
12670071
负责人：
MAEDA Masanobu
金额：
$ 2.5万
依托单位：
Wakayama Medical Universtiy
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-12670071/
关键词：
nucleus tractus solitarii nitric oxide nitric oxide synthase green fluorescent protein (GFP)fluorescence brain circulation gene 孤束核

项目摘要

In order to investigate the motion of a single molecule of neuronal nitric oxide synthase (nNOS) in the nucleus tractus solitarius (NTS), we tried gene (green fluorescent protein and nNOS) transduction into the NTS.1. The simple methods for gene transduction in vivo into the brain using plasmid vector have not been developed. We microinjected green fluorescent protein (GFP) Fusion TOPO Cloning Vectors with cationic lipids (LipofextAMINE 2000) into the NTS and the hypoglossal nucleus, and removed the brain three, five, seven and ten days after the microinjection, and observed the fluorescence.2. We could observe the fluorescence four brains microinjected out of five rats five days after the injection of plasmid vectors. The intensity of the fluorescence was the strongest five days after the injection. It is considered that the gene expression was the strongest five days after the transduction.3. Next, we microinjected LacZ Vectors with cationic lipids into the brain. However, we could not observe beta-gal. It may be considered that the efficiency of gene transduction into the brain using a little large vector may be low.4. Several new transfection drugs have been developed rapidly. We tried some new transfection drugs. We could observe the beta-gal using a certain new transfection drug.

为了研究神经元一氧化氮合酶（nNOS）单分子在孤束核（NTS）中的运动，我们尝试了基因（绿色荧光蛋白和nNOS）在孤束核（NTS）中的转导。利用质粒载体在体内将基因转导到脑内的简单方法尚未开发。我们将含有阳离子脂质（lipofexamine 2000）的绿色荧光蛋白（GFP）融合TOPO克隆载体微注射到NTS和舌下核中，并在微注射后3、5、7和10天取出脑，观察荧光。注射质粒载体后5天，我们可以观察到5只大鼠的4个脑的荧光。注射后第5天荧光强度最强。在转导后的第5天，基因表达最强。接下来，我们将带阳离子脂质的LacZ载体微注射到大脑中。然而，我们无法观察到β -gal。可以认为，使用稍大的载体将基因转导到大脑的效率可能较低。一些新的转染药物被迅速开发出来。我们尝试了一些新的转染药物。我们可以用一种新的转染药物来观察β -半乳糖。