1993 Fiscal Year Annual Research Report

NMDA受容体チャネル欠損マウスの作成

Research Project

Project/Area Number	05404085
Research Institution	Niigata University
Principal Investigator	三品昌美新潟大学, 脳研究所, 教授 (80144351)
Co-Investigator(Kenkyū-buntansha)	崎村建司新潟大学, 脳研究所, 助教授 (40162325)
Keywords	シナプス可塑性 / 記憶 / 学習 / シナプス形成 / NMDA受容体チャネル / 遺伝子ノックアウト / ES細胞
Research Abstract	本研究は、ジーンターゲッティング法によりNMDA受容体チャネルの各サブユニットを欠損した変異マウスを作成し、NMDA受容体チャネルのシナプス可塑性、シナプス伝達、記憶・学習、シナプス形成および脳の病態時における神経細胞死に果たす役割を個体レベルで解明することを目的とする。研究計画が順調に進展し、本年度は下記の成果を挙げた。 1.クローン化したcDNAをプローブとしてマウスNMDA受容体チャネルの多様性を決定している4種類のεサブユニット(ε1、ε2、ε3およびε4サブユニット)の遺伝子断片を単離した。制限酵素地図を作成し、エキソン部分の塩基配列を決定することにより、単離した断片の遺伝子構造を明らかにした。2.NMDA受容体チャネルの各εサブユニット遺伝子のエキソン部分にネオマイシン耐性遺伝子を挿入することにより、NMDA受容体チャネルのサブユニット遺伝子を不活化した。さらにネガティブセレクションのためにジフテリアトキシン遺伝子を有するベクターに不活化したNMDA受容体チャネルのεサブユニット遺伝子を連結し、標的遺伝子組換えに用いるターゲッティングベクターを構築した。3.不活化したNMDA受容体チャネルの各εサブユニット遺伝子を連結したターゲッティングベクターをマウスES細胞にエレクトロポレーション法により導入した。G418耐性株を単離し、さらにPCR法およびサザンブロット法によりε1、ε2およびε4サブユニット遺伝子をノックアウトした細胞株を得ることに成功した。ε3サブユニット遺伝子については現在スクリーニングを続けている。

Research Products

(16 results)

All Other

All Publications (16 results)

[Publications] Nakano,R.et al.: "A novel mutation in Cu/Zn superoxide dismutase gene in Japanese familial amyotrophic lateral sclerosis" Biochem.Biophys.Res.Commun.(in press). (1994)
[Publications] Watanabe,M.et al.: "Distinct spatio-temporal expressions of five NMDA receptor channel subunit mRNAs in the cerebellum" J.Comparative Neurology. (in press). (1994)
[Publications] Watanabe,M.et al.: "Distinct distributions of five NMDA receptor channel subunit mRNAs in the brainstem" J.Comparative Neurology. (in press). (1994)
[Publications] Watanabe,M.et al.: "Distinct gene expression of the N-methyl-_D-aspartate receptor channel subunit in peripheral neurons of the mouse sensory ganglia and adrenal gland" Neuroscience Letters. (in press). (1994)
[Publications] Watanabe,M.et al.: "Distinct spatio-temporal distributions of the NMDA receptor channel subunit mRNAs in the mouse cervical cord" J.Comparative Neurology. (in press). (1994)
[Publications] Ishida,N.et al.: "Circadian expression of NMDA recptor mRNAs,ε3 and ζ1,in the suprachiasmatic nucleus of rat brain" Neurosci.Lett.(in press). (1994)
[Publications] Watanabe,M.et al.: "Differential distributions of the NMDA receptor channel subunit mRNAs in the mouse retina." Brain Res.634. 328-332 (1994)
[Publications] Araki,K.et al.: "Selective expression of the glutamate receptor channel δ2 subunit in cerebellar Purkinje cells." Biochem.Biophtys.Res.Commun.197. 1267-1276 (1993)
[Publications] Takano,H.et al.: "Chromosomal localization of the ε1,ε3 and ζ1 subunit genes of the human NMDA receptor channel." Biochem.Biophtys.Res.Commun.197. 922-926 (1993)
[Publications] Yamakura,T.et al.: "Phosphorylation of the carboxyl-terminal domain of the ζ1 subunit is not responsible for potentiation by TPA of the NMDA receptor channel." Biochem.Biophtys.Res.Commun.196. 1537-1544 (1993)
[Publications] Watanabe,M.et al.: "Distinct distributions of five N-methyl-_D-aspartate receptor channel subunit mRNAs in the forebrain." J.Comp.Neurol.338. 377-390 (1993)
[Publications] Watanabe,M.et al.: "Distinct spatio-temporal distributions of the NMDA receptor channel subunit mRNAs in the brain." Ann.NY Acad.Sci.707. 463-466 (1993)
[Publications] Kawamoto,S.et al.: "Expression of the α1 and α2 subunits of the AMPA-selective glutamate receptor channel in insect cells using a Baculovirus vector" Ann.NY Acad.Sci.707. 460-462 (1993)
[Publications] Mishina,M.et al.: "Molecular and functional diversity of the NMDA receptor channel" Ann.NY Acad.Sci.707. 136-152 (1993)
[Publications] Yamakura,T.et al.: "Different sensitivities of NMDA receptor channel subtypes to noncompetitive antagonists" NeuroReport. 4. 687-690 (1993)
[Publications] Mori,H.et al.: "Involvement of the carboxyl-terminal region in modulation by TPA of the NMDA receptor channel" NeuroReport. 4. 519-522 (1993)

1993 Fiscal Year Annual Research Report

NMDA受容体チャネル欠損マウスの作成

Principal Investigator

三品 昌美 新潟大学, 脳研究所, 教授 (80144351)

Research Products

[Publications] Nakano,R.et al.: "A novel mutation in Cu/Zn superoxide dismutase gene in Japanese familial amyotrophic lateral sclerosis" Biochem.Biophys.Res.Commun.(in press). (1994)

[Publications] Watanabe,M.et al.: "Distinct spatio-temporal expressions of five NMDA receptor channel subunit mRNAs in the cerebellum" J.Comparative Neurology. (in press). (1994)

[Publications] Watanabe,M.et al.: "Distinct distributions of five NMDA receptor channel subunit mRNAs in the brainstem" J.Comparative Neurology. (in press). (1994)

[Publications] Watanabe,M.et al.: "Distinct gene expression of the N-methyl-_D-aspartate receptor channel subunit in peripheral neurons of the mouse sensory ganglia and adrenal gland" Neuroscience Letters. (in press). (1994)

[Publications] Watanabe,M.et al.: "Distinct spatio-temporal distributions of the NMDA receptor channel subunit mRNAs in the mouse cervical cord" J.Comparative Neurology. (in press). (1994)

[Publications] Ishida,N.et al.: "Circadian expression of NMDA recptor mRNAs,ε3 and ζ1,in the suprachiasmatic nucleus of rat brain" Neurosci.Lett.(in press). (1994)

[Publications] Watanabe,M.et al.: "Differential distributions of the NMDA receptor channel subunit mRNAs in the mouse retina." Brain Res.634. 328-332 (1994)

[Publications] Araki,K.et al.: "Selective expression of the glutamate receptor channel δ2 subunit in cerebellar Purkinje cells." Biochem.Biophtys.Res.Commun.197. 1267-1276 (1993)

[Publications] Takano,H.et al.: "Chromosomal localization of the ε1,ε3 and ζ1 subunit genes of the human NMDA receptor channel." Biochem.Biophtys.Res.Commun.197. 922-926 (1993)

[Publications] Yamakura,T.et al.: "Phosphorylation of the carboxyl-terminal domain of the ζ1 subunit is not responsible for potentiation by TPA of the NMDA receptor channel." Biochem.Biophtys.Res.Commun.196. 1537-1544 (1993)

[Publications] Watanabe,M.et al.: "Distinct distributions of five N-methyl-_D-aspartate receptor channel subunit mRNAs in the forebrain." J.Comp.Neurol.338. 377-390 (1993)

[Publications] Watanabe,M.et al.: "Distinct spatio-temporal distributions of the NMDA receptor channel subunit mRNAs in the brain." Ann.NY Acad.Sci.707. 463-466 (1993)

[Publications] Kawamoto,S.et al.: "Expression of the α1 and α2 subunits of the AMPA-selective glutamate receptor channel in insect cells using a Baculovirus vector" Ann.NY Acad.Sci.707. 460-462 (1993)

[Publications] Mishina,M.et al.: "Molecular and functional diversity of the NMDA receptor channel" Ann.NY Acad.Sci.707. 136-152 (1993)

[Publications] Yamakura,T.et al.: "Different sensitivities of NMDA receptor channel subtypes to noncompetitive antagonists" NeuroReport. 4. 687-690 (1993)

[Publications] Mori,H.et al.: "Involvement of the carboxyl-terminal region in modulation by TPA of the NMDA receptor channel" NeuroReport. 4. 519-522 (1993)

三品昌美新潟大学, 脳研究所, 教授 (80144351)