Studies on brain functions by gene replacement method.

通过基因替代方法研究脑功能。

基本信息

批准号：
05454673
负责人：
KATSUKI Motoya
金额：
$ 4.22万
依托单位：
Kyushu University
依托单位国家：
日本
项目类别：
Grant-in-Aid for General Scientific Research (B)
财政年份：
1993
资助国家：
日本
起止时间：
1993 至 1995
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-05454673/
关键词：
Gene Targeting Knockout Mouse Gene Replacement Glutamate receptor Dopamine receptor mGluR6 alpha-MSH Retina 光刺激 NMDAR2A NMDAR2C

项目摘要

Recent advances in embryo manipulating technologies made it possible to generate the desired mutaion for elucidation of brain functions. We have generated knockout mice lacking neurotransmitter receptors. First, taking advantage of the restricted expression of metabotropic glutamate receptor subtype6 (mGluR6) in retinal ON bipolar cells, we generated knockout mice lacking mGluR6 expression. The homozygous mutnat mice showed a loss of ON responses but unchanged OFF responses to light. The mutant mice displayd no obvious changes in retinal cell organization nor in the projection of optic fibers to the brain. Furthemore, the mGluR6-deficient mice showed visual behavioral responses to light stimulation as examined by shuttle box avoidance behavior experiments using light exposure as a conditioned stimulus. The results demonstrate that mGluR6 is essential in synaptic transmission to the ON bipolar cell and that the OFF response provides an important means for transmitting visual inforamation.Secondly, to elucidate the specific role of the dopamine D2 receptor (D2R) in the pituitary and brain, we generated D2R deficient (D2R mutant) mice using the gene targeting method. Electrophysiological studies as well as ligand binding assays show no functional D2R in mutant mice. The D2R mutant mice display a hypoacitivity and a slow movement with creeping manner. The expression of enkephalin mRNA in the striatum is increased in the mutant mice, but not that of dynorphin and substanceP.D2R mutant mice have significant darker coat color than wild type littermates and show an elevation of plasma alpha-melanocyte stimulating hormone levels. We found corresponding hyperplastic changes of intermediate lobe of the pituitary and the increased expression of proopiomelanocortin in the pituitarv in D2R mutant mice. These findings suggest that D2R plays a critical role in the dopaminergic system in the pituitary and the striatum.

胚胎操纵技术的最新进展使得产生阐明大脑功能所需的突变成为可能。我们已经培育出缺乏神经递质受体的基因敲除小鼠。首先，利用视网膜 ON 双极细胞中代谢型谷氨酸受体亚型 6 (mGluR6) 的限制性表达，我们产生了缺乏 mGluR6 表达的基因敲除小鼠。纯合突变小鼠表现出对光的ON反应丧失，但OFF反应未改变。突变小鼠的视网膜细胞组织和光纤向大脑的投射没有表现出明显的变化。此外，mGluR6 缺陷小鼠表现出对光刺激的视觉行为反应，通过使用光暴露作为条件刺激的穿梭箱回避行为实验进行了检查。结果表明，mGluR6对于向ON双极细胞的突触传递至关重要，而OFF反应为传递视觉信息提供了重要手段。其次，阐明多巴胺D2受体（D2R）在垂体和大脑中的具体作用，我们使用基因打靶方法生成了 D2R 缺陷（D2R 突变体）小鼠。电生理学研究以及配体结合测定显示突变小鼠中没有功能性 D2R。 D2R突变小鼠表现出活动能力减退和爬行方式缓慢运动。突变小鼠纹状体中脑啡肽 mRNA 的表达增加，但强啡肽和 P 物质的表达没有增加。D2R 突变小鼠的毛色比野生型同窝小鼠明显更深，并且血浆 α-黑素细胞刺激激素水平升高。我们发现D2R突变小鼠的垂体中叶有相应的增生性变化，并且垂体中阿黑皮素原的表达增加。这些发现表明 D2R 在垂体和纹状体的多巴胺能系统中发挥着关键作用。