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信号传导与烟碱乙酰胆碱受体（NACHR）的串扰，降低了信号通路，我们首先研究了NGF和cAMP在PC12细胞的三个subline，PC12H细胞，PC12细胞，PC12细胞中对NACHR表达的调节，PC12细胞，PC12细胞表达主导RAS和PC12细胞PC12细胞（PC12细胞）。 cAMP模拟在PC12H和PC12-wild细胞中通过蛋白激酶A.NGF上调的PC12H细胞中的α3MRNA水平下调的NACHRα3亚基mRNA（α3MRNA）的水平下调水平，并下调了PC12-wild细胞中α3MRNA的α3MRNA水平，这些细胞中α3MRNA被PC12-Wild细胞中介导了Ras-Map kescace cascace cascace cascace cascace ras kinases casccade。高K^+的膜沉积对PC12H细胞中的α3MRNA水平没有影响。基于这些结果，我们提出，MAP激酶（ERK）和CREB的至少两个未知的下游因子在PC12细胞中调节α3MRNA。尼古丁诱导的ERK磷酸化是瞬时的，其水平低于高k^+ depolariation诱导的ERK磷酸化和NGF.α7NACHR亚基选择性拮抗剂对尼古丁诱导的ERK磷酸化没有影响。 L型电压敏感的钙通道拮抗剂抑制了尼古丁诱导的ERK磷酸化。这些结果表明，尼古丁诱导的ERK磷酸化无含α7的NACHR。通过钙调蛋白拮抗剂，CAM激酶抑制剂，MAP激酶激酶抑制剂对尼古丁诱导的ERK磷酸化的抑制作用，并且显性抑制性RA的表达表明CAM激酶和RAS-MAP激酶级联反应参与烟碱诱导的ERK磷酸化。此外，已经提出，大多数尼古丁诱导的CREB磷酸化是由尼古丁诱导的PC12H细胞中尼古丁诱导的ERK磷酸化介导的。