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Functional clarification of the bombesin-like peptide system.

铃蟾肽样肽系统的功能澄清。

基本信息

批准号：
12680769
负责人：
OHKI-HAMAZAKI Hiroko
金额：
$ 2.56万
依托单位：
Tokyo Medical and Dental University
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

项目摘要

1. Generation and analysis of mice lacking NMB-R and GRP-R : We obtained male double-knocked out mice as well as male wild-type mice by mating female NMB-R +/-, GRP-R +/- mice and male NMB-R +/- mice. The mutant mice did not show great difference as compared with wild-type mice. The possible alterations observed were the points already noticed in the GRP-R or NMB-R single mutant mice.2. Development of the chick embryos over-expressing NMB-R, GRP-R or BRS-3 : We constructed the retrovirus vectors containing cDNA coding for each subtype of mouse receptpr with EGFP cDNA fused at its C-terminal end. Infection of virus particles prepared using these vectors to the brain region of the chick embryos, resulted in the strong and wide-spread expression of EGFP. We are currently analyzing the phenotypes of these embryos by histological methods.3. Analysis of bombesin systems in birds : In order to elucidate the bombesin like peptides system in birds, we successfully cloned two bombesin-like peptide receptors from chick brain by using RT-PCR with degenerate primers. We named these clones as chGRP-R which showed high similarity with GRP-R of other species, and chBRS-3.5 which showed similarity with BRS-3 and BB4. In situ hybridization analysis showed that chGRP-R was expressed in the hypothalamus and in discrete regions of forebrain, and chBRS-3.5 was widely expressed in the forebrain including striatum. None of these two receptors was expressed in the cerebellum. These results demonstrated that the bombesin-like peptides system exists also in birds and this suggests that this system plays an important role in birds like in mammals.

1.缺乏NMB-R和GRP-R的小鼠的产生和分析：我们通过使雌性NMB-R +/-、GRP-R +/-小鼠和雄性NMB-R +/-小鼠交配来获得雄性双敲除小鼠以及雄性野生型小鼠。突变小鼠与野生型小鼠相比没有显示出很大的差异。观察到的可能的改变是在GRP-R或NMB-R单突变小鼠中已经注意到的点。过表达NMB-R、GRP-R和BRS-3的鸡胚发育：构建了编码小鼠受体各亚型cDNA的逆转录病毒载体，并在其C末端融合了EGFP cDNA。用这些载体制备的病毒颗粒感染鸡胚脑区，可使EGFP得到强而广泛的表达。我们目前正在通过组织学方法分析这些胚胎的表型。鸟类蛙皮素系统的分析：为了阐明鸟类蛙皮素样肽系统，我们利用简并引物RT-PCR技术从鸡脑中成功克隆了两个蛙皮素样肽受体。我们将这些克隆命名为chGRP-R和chBRS-3.5，其中chGRP-R与其他物种的GRP-R具有高度的相似性，chBRS-3.5与BRS-3和BB 4具有相似性。原位杂交结果显示，chGRP-R在下丘脑和前脑的不同区域表达，chBRS-3.5在前脑包括纹状体广泛表达。这两种受体在小脑中均不表达。这些结果表明，蛙皮素样肽系统也存在于鸟类中，这表明该系统在鸟类中起着与哺乳动物一样的重要作用。