权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Studies on signal transduction mechanisms from nicotinic acetylcholine receptors to nucleus

烟碱型乙酰胆碱受体至细胞核信号转导机制的研究

基本信息

批准号：
11680761
负责人：
NAKAYAMA Hitoshi
金额：
$ 1.98万
依托单位：
Nara Medical University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2000
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11680761/
关键词：
acetylcholine receptor protein phosphorylation nicotine nicotinic cAMP NGF MAP kinase CREB PC12細胞

项目摘要

Considering the crosstalk between NGF/cAMP signaling and nicotinic acetylcholine receptor (nAChR)-mediating signaling pathways, we first investigated regulation of nAChR expression by NGF and cAMP in three sublines of PC12 cells, PC12h cells, PC12 cells expressing dominant negative Ras and the parential PC12 cells (PC12-wild). cAMP analogue down-regulated level of nAChR α3 subunit mRNA (α3mRNA) in PC12h and PC12-wild cells through protein kinase A.NGF up-regulated the α3mRNA level in PC12h cells and down-regulated the α3mRNA in PC12-wild cells, which is mediated by Ras-MAP kinase cascade. Membrane depolarization with high K^+ had no effect on the α3mRNA level in PC12h cells. Based on these results, we proposed that at least two unknown downstream factors of MAP kinase (ERK) and CREB regulate α3mRNAs in PC12 cells.Next we investigated mechnisms of nicotine-induced MAP kinase (ERK) and CREB phosphorylation in PC12h cells. Nicotine-induced ERK phosphorylation was transient and its level was lower than that of ERK phosphorylation induced by high K^+ depolarization and NGF.α7 nAChR subunit-selective antagonists had no effect on nicotine-induced ERK phosphorylation. L-type voltage-sensitive calcium channel antagonists inhibited nicotine-induced ERK phosphorylation. These results suggest that no α7-containing nAChRs was involved in nicotine-induced ERK phosphorylation. Inhibition of nicotine-induced ERK phosphorylation by a calmodulin antagonist, CaM kinase inhibitor, MAP kinase kinase inhibitor and expression of dominant inhibitory Ras shows that CaM kinase and Ras-MAP kinase cascade are involved in nicotine-induced ERK phosphorylation. Furthermore, it has been suggested that most of nicotine-induced CREB phosphorylation is mediated by nicotine-induced ERK phosphorylation in PC12h cells.

考虑到NGF/cAMP信号通路与烟碱乙酰胆碱受体（nAChR）介导的信号通路之间的相互干扰，我们首先在PC 12细胞、PC 12 h细胞、表达显性负性Ras的PC 12细胞和亲代PC 12细胞（PC 12-wild）中研究了NGF和cAMP对nAChR表达的调控。cAMP类似物通过蛋白激酶A下调PC 12 h和PC 12-wild细胞nAChR α3亚基mRNA（α 3 mRNA）水平。NGF通过Ras-MAP激酶级联反应上调PC 12 h细胞α 3 mRNA水平，下调PC 12-wild细胞α 3 mRNA水平。高K^+去极化对PC 12 h细胞α 3 mRNA水平无影响。在此基础上，我们进一步研究了尼古丁诱导PC 12 h细胞中MAP激酶（ERK）和CREB磷酸化的机制。尼古丁诱导的ERK磷酸化是短暂的，其水平低于高K^+去极化和NGF诱导的ERK磷酸化。α7 nAChR亚基选择性拮抗剂对尼古丁诱导的ERK磷酸化没有影响。L型电压敏感性钙通道拮抗剂抑制尼古丁诱导的ERK磷酸化。这些结果表明，没有含α7的nAChR参与尼古丁诱导的ERK磷酸化。通过钙调蛋白拮抗剂、CaM激酶抑制剂、MAP激酶抑制剂和显性抑制性Ras的表达抑制烟碱诱导的ERK磷酸化，表明CaM激酶和Ras-MAP激酶级联参与烟碱诱导的ERK磷酸化。此外，已经表明，在PC 12 h细胞中，尼古丁诱导的CREB磷酸化的大部分由尼古丁诱导的ERK磷酸化介导。