1998 Fiscal Year Annual Research Report

自己免疫疾患「ランバート・イートン筋無力症候群」モデル動物の開発

Research Project

Project/Area Number	10557232
Research Institution	The University of Tokyo
Principal Investigator	桐野豊東京大学, 大学院・薬学系研究科, 教授 (10012668)
Co-Investigator(Kenkyū-buntansha)	川上善之エーザイ, 筑波探索研究所, 主幹研究員渡辺恵東京大学, 大学院・薬学系研究科, 助手 (80302610) 川原茂敬東京大学, 大学院・薬学系研究科, 助手 (10204752)
Keywords	自己免疫疾患 / ランバート・イートン症候群 / 神経筋接合部 / シビレエイ / 電気器官 / シナプトソーム / カルシウムチャネル / モノクローナル抗体
Research Abstract	1. シナプトソーム免疫によるモデル動物の作成:シビレエイ電気器官シナプトソームで免疫したマウスの神経-筋標本(横隔膜)を電気生理学的に調べた。自発的な微小終板電位(MEPP)の大きさに差は見られなかったが、運動神経を電気刺激して生じる終板電位(EPP)の大きさは減少していた。MEPPの結果はアセチルコリン受容体側に変化がないことを示している。EPPの量子解析からはquantal contentの減少が確認された。以上のことは、神経終末のCa^<2+>チャネルが阻害されていることを示しており、LEMS様症状を示すモデル動物を作成することができたと考えられた。 2. 受動免疫によるモデル動物の作成:上記と同様に電気器官シナプトソームを用いてウサギに能動免疫を行い、そのウサギの血清をマウスに注入することによりマウスへの受動免疫を行った。LEMS患者の血清を受動免疫した場合と同様に、quantalcontentの減少が認められた。 3. モノクローナル抗体産生の作成:電気器官シナプトソーム膜標品で免疫したマウスの脾臓より抗体産生ハイブリドーマを作成した。スクリーニングにはELISA法を用い、シナプトソーム膜標品に反応し、神経細胞体膜標品には反応しない抗体を産生するハイブリドーマを選択した。その結果、認識抗原が異なる7種類のハイブリドーマを得ることができた。次に、これらハイブリドーマがLEMS様症状を引き起こす抗体を産生するかどうかを調べるために、ハイブリドーマをマウス腹腔内で増殖させ、そのマウスの横隔膜標本の筋電位を細胞外記録により測定した。その結果、7種類のハイブリドーマのうちの1つにおいて、頻回刺激で筋電位の増強が生じる(facilitation)というLEMSに特徴的な現象を確認することができた。今後、さらに細胞内記録により量子解析を行ってLEMS様症状を確認する予定である。

Research Products
(20 results)

All Other

All Publications (20 results)

[Publications] Ko-ichi Iijima: "Phosphorylated Form of Alzheimer's Amyloid Precursor Protein Exists in Cholinergic Nerve T ermini from Electric Ray Electric Organ" Biochem.J.330. 29-33 (1998)
[Publications] Susumu Tomita: "Cleavege of Alzheimer's Amyloid Precursor Protein by Secretases Occurs after O-glycosyla-tion of APP in Protein Secretory Pathway" J.Biol.Chem.273. 6277-6284 (1998)
[Publications] Hirotasu Kojima: "Development of a Fluorescent Indicator for Nitric Oxide Based on the Fluorescein Chromophore" Chem.Pharm.bull.46. 373-375 (1998)
[Publications] Yasushi Satoh: "Lambert-Eaton Syndrome Antibodies Inhibit Acetylcholine Release and P/Q-type Ca^<2+> Channels in Electric Ray Nerve Endings" J.Physiol.508. 427-438 (1998)
[Publications] Tetsuya Kimura: "Behavioral Modulation Induced by Food-Odor Aversive Conditioning,and Its Influence on the Olfactory Responses of an Oscillatory Brain Network in the Slug, Limax marginatus" Learning and Memory. 4. 365-375 (1998)
[Publications] Tetsuya Kimura: "Optical Recording Analysis of Olfactory Response of the PC Lobe in the Slug Brain" Learning and Memory. 4. 389-400 (1998)
[Publications] Satoshi Watanabe: "Morphological Characterization of the Bursting and Nonbursting Neurons in the Olfactory Cent er of the Terrestrial Slug Limax marginatus" J.Exp.Biol.201. 925-930 (1998)
[Publications] Yong I.Kim: "An Autoimmune Animal Model of the Lambert-Eaton Syndrom" Ann.N.Y.Acad.Sci. 841. 670-676 (1998)
[Publications] Shigeo Watanabe: "Differential Roles of Two Types of Voltage-gated Ca^<2+> Channels in the Dendrites of Rat Cerebellar Purkinje Neurons" Brain Res.791. 43-55 (1998)
[Publications] Susumu Tomita: "A Basic Amino Acid in the Cytopolasmic Domain of Altzheimer's β-Amyloid Precursor Protein(APP)Is Essential for Cleavage of APP at the α-site" J.Biol.Chem.273. 19304-19310 (1998)
[Publications] Hirotatsu Kojima: "Detection and Imaging of Nitric Oxide with Novel Fluorescent Indicators: Diaminofluoresceins" Anal.Chem.70. 2446-2453 (1998)
[Publications] 桐野豊: "学習記憶研究の戦略" 放射線医学. 38. 174-177 (1998)
[Publications] Dong-Seok Lee: "Phospholipid Translocation from Outer to Inner Leaflet of Synaptic Vesicle membranes Isolated from the Electric Organ of Japanese Electric Ray Narke japonica" J.Biochem.124. 798-803 (1998)
[Publications] Tsuyoshi Inoue: "Selective Optical Recording of the Neural Activity in the Olfactory Center of Land Slug Using a Calcium Indicator Dye" Bioimages. 6. 59-67 (1998)
[Publications] Azusa Nakagawa-Inoue: "Odorant-evoked Increase in Cytosolic Free Calcium in Cultured Anntenal Neurons of Blowflies" Zool.Sci.15. 661-666 (1998)
[Publications] Yutaka Hirakura: "Membrane Perturbation by the Alzheimer Amyloid Fragment β25-35 Requires Aggregation" Biochem.Mol.Biol.Intnl.46. 787-794 (1998)
[Publications] Susumu Tomita: "Interaction of a Neuron-specific Protein Containing PDZ Domains with AlzHeimer's Amyloid Precursor Protein" J.Biol.Chem.274. 2243-2254 (1999)
[Publications] Satoshi Watanabe: "Glutamate Induces Cl^- and K^+ Currents in the Olfacotry Interneurons of a Terrestrial Slug" J.Comp.Physiol.A. (in press). (1999)
[Publications] Kanae Ando: "Role of Phsphorylation of Alzheimer's Amyloid Precursor Protein during Neuronal Differentiation" J.Neurosci.(in press). (1999)
[Publications] 桐野豊: "基礎薬学物理化学(改訂版)" 廣川書店(in press), (1999)

1998 Fiscal Year Annual Research Report

自己免疫疾患「ランバート・イートン筋無力症候群」モデル動物の開発

Principal Investigator

桐野 豊 東京大学, 大学院・薬学系研究科, 教授 (10012668)

Research Products

[Publications] Ko-ichi Iijima: "Phosphorylated Form of Alzheimer's Amyloid Precursor Protein Exists in Cholinergic Nerve T ermini from Electric Ray Electric Organ" Biochem.J.330. 29-33 (1998)

[Publications] Susumu Tomita: "Cleavege of Alzheimer's Amyloid Precursor Protein by Secretases Occurs after O-glycosyla-tion of APP in Protein Secretory Pathway" J.Biol.Chem.273. 6277-6284 (1998)

[Publications] Hirotasu Kojima: "Development of a Fluorescent Indicator for Nitric Oxide Based on the Fluorescein Chromophore" Chem.Pharm.bull.46. 373-375 (1998)

[Publications] Yasushi Satoh: "Lambert-Eaton Syndrome Antibodies Inhibit Acetylcholine Release and P/Q-type Ca^<2+> Channels in Electric Ray Nerve Endings" J.Physiol.508. 427-438 (1998)

[Publications] Tetsuya Kimura: "Behavioral Modulation Induced by Food-Odor Aversive Conditioning,and Its Influence on the Olfactory Responses of an Oscillatory Brain Network in the Slug, Limax marginatus" Learning and Memory. 4. 365-375 (1998)

[Publications] Tetsuya Kimura: "Optical Recording Analysis of Olfactory Response of the PC Lobe in the Slug Brain" Learning and Memory. 4. 389-400 (1998)

[Publications] Satoshi Watanabe: "Morphological Characterization of the Bursting and Nonbursting Neurons in the Olfactory Cent er of the Terrestrial Slug Limax marginatus" J.Exp.Biol.201. 925-930 (1998)

[Publications] Yong I.Kim: "An Autoimmune Animal Model of the Lambert-Eaton Syndrom" Ann.N.Y.Acad.Sci. 841. 670-676 (1998)

[Publications] Shigeo Watanabe: "Differential Roles of Two Types of Voltage-gated Ca^<2+> Channels in the Dendrites of Rat Cerebellar Purkinje Neurons" Brain Res.791. 43-55 (1998)

[Publications] Susumu Tomita: "A Basic Amino Acid in the Cytopolasmic Domain of Altzheimer's β-Amyloid Precursor Protein(APP)Is Essential for Cleavage of APP at the α-site" J.Biol.Chem.273. 19304-19310 (1998)

[Publications] Hirotatsu Kojima: "Detection and Imaging of Nitric Oxide with Novel Fluorescent Indicators: Diaminofluoresceins" Anal.Chem.70. 2446-2453 (1998)

[Publications] 桐野 豊: "学習記憶研究の戦略" 放射線医学. 38. 174-177 (1998)

[Publications] Dong-Seok Lee: "Phospholipid Translocation from Outer to Inner Leaflet of Synaptic Vesicle membranes Isolated from the Electric Organ of Japanese Electric Ray Narke japonica" J.Biochem.124. 798-803 (1998)

[Publications] Tsuyoshi Inoue: "Selective Optical Recording of the Neural Activity in the Olfactory Center of Land Slug Using a Calcium Indicator Dye" Bioimages. 6. 59-67 (1998)

[Publications] Azusa Nakagawa-Inoue: "Odorant-evoked Increase in Cytosolic Free Calcium in Cultured Anntenal Neurons of Blowflies" Zool.Sci.15. 661-666 (1998)

[Publications] Yutaka Hirakura: "Membrane Perturbation by the Alzheimer Amyloid Fragment β25-35 Requires Aggregation" Biochem.Mol.Biol.Intnl.46. 787-794 (1998)

[Publications] Susumu Tomita: "Interaction of a Neuron-specific Protein Containing PDZ Domains with AlzHeimer's Amyloid Precursor Protein" J.Biol.Chem.274. 2243-2254 (1999)

[Publications] Satoshi Watanabe: "Glutamate Induces Cl^- and K^+ Currents in the Olfacotry Interneurons of a Terrestrial Slug" J.Comp.Physiol.A. (in press). (1999)

[Publications] Kanae Ando: "Role of Phsphorylation of Alzheimer's Amyloid Precursor Protein during Neuronal Differentiation" J.Neurosci.(in press). (1999)

[Publications] 桐野 豊: "基礎薬学 物理化学(改訂版)" 廣川書店(in press), (1999)

桐野豊東京大学, 大学院・薬学系研究科, 教授 (10012668)

[Publications] 桐野豊: "学習記憶研究の戦略" 放射線医学. 38. 174-177 (1998)

[Publications] 桐野豊: "基礎薬学物理化学(改訂版)" 廣川書店(in press), (1999)