Functional analysis of neuronal Ca^<2+> channel domains as targets of therapeutics

作为治疗靶标的神经元 Ca^2 通道域的功能分析

• 批准号: 13672278
• 负责人: KANEKO Shuji
• 金额: $ 2.3万
• 依托单位: KYOTO UNIVERSITY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2001
• 资助国家: 日本
• 起止时间: 2001 至 2002
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-13672278/
• 关键词: calcium channels; voltage-dependent; TRP; Alzheimer's disease; alternative splicing; Xenopus oocytes; neuropathic pain; neuronal death; TRPチャネル; アンチセンスDNA; 免疫染色; RT-PCR; 初代培養ニューロン; 不活性化; アフリカツメガエル卵母細胞; スプライシングバリアント; GTP結合タンパク; FK960; PMP

What we found were (1) The interaction of N terminal region of Ga_O and C-terminal domain of α_<1A> subunit causes the voltage-resistant inhibition of P/Q-type channel current. (2) Cytosolic Ca^<2+> is required for the opening of store-operated TRP4 channels, whereas the activation of inositol-1,4,5-trisphosphate receptor (IP_3-R) is required and sufficient for the opening of receptor-activated TRP5 channels. (3) Two cDNAs encoding RNA splicing variants of human Ca^<2+> channel α1B subunit were isolated from human brain cDNA libralies. These variants were RNA splicing variants by genomic analysis. One of the variants lacking the II III linker region formed Ca^<2+>-permeable channel having lowωconotoxin sensitivity, as revealed by exogenous expression in human embryonic kidney (HER) cells. (4) An insect peptide PMP-D2 selectively inhibited R-type Cav2.3 channels. (5) A novel antiamneasic drug, FK960, selectively potentiated N-type Cav2.2 channel current in a protein kinase C-dependent manner. (6) A novel analgesic, ONO-2921, selectively inhibited N- and R- type channel currents due to its specific action on inactivated channels. (7) The store-operated and receptoractivated Ca^<2+> entry in rat cerebral cortical neurons in culture by Fura-2 fluorometry. By RT-PCR analysis, we found the presence of rat TRP1, 3, 5 and 6 mRNAs in neurons. Immunostaninig results also suggest that TRP1 and 5 proteins are expressed in the cultured neurons.

我们发现的是（1）α_<1A_>亚基的Ga_O和C末端结构域的N末端区域的相互作用会导致抗压抑制P/Q-Type通道电流。 （2）开放储存的TRP4通道需要胞质Ca^<2+>，而肌醇1,4,4,5-三磷酸受体（IP_3-R）的激活是需要受体激活的TRP5通道的开放的。 （3）从人脑cDNA文库中分离出人Ca^<2+>α1b亚基的两个编码RNA剪接变体的cDNA。这些变体通过基因组分析是RNA剪接变体。缺乏III III连接器区域的一种变体形成了Ca^<2+> - 具有低晶毒素敏感性的渗透通道，如人类胚胎肾脏（她的）细胞中的外源表达所揭示的那样。 （4）抑制肽PMP-D2选择性抑制R型CAV2.3通道。 （5）一种新型的抗neasic药物FK960，以蛋白激酶C依赖性方式有选择性化的N型CAV2.2通道电流。 （6）一种新型的镇痛作用，ONO-2921，由于其对灭活通道的特定作用，选择性抑制了N型通道电流。 （7）通过Fura-2荧光测定法进入储存和识别的CA^<2+>在培养中的大鼠脑皮质神经元中的进入。通过RT-PCR分析，我们发现神经元中存在大鼠TRP1、3、5和6 mRNA。免疫稳态的结果还表明，TRP1和5蛋白在培养的神经元中表达。

项目成果

金子周司: "Ca2+チャネル遺伝子の選択的スプライシングとその機能的意義"日本薬理学雑誌. 121. 233-240 (2003)

Shuji Kaneko：“Ca2+通道基因的选择性剪接及其功能意义”日本药理学杂志 121. 233-240 (2003)。

Hiroyuki Tabata, Satoshi Tanaka, Yukihiko Sugimoto, Hideaki Kanki, Shuji Kaneko and Atsushi Ichikawa: "Possible coupling of prostaglandin E receptor EP_1 to TRP5 expressed in Xenopus oocytes"Biochem. Biophys. Res. Commun.. 298. 398-402 (2002)

Hiroyuki Tabata、Satoshi Tanaka、Yukihiko Sugimoto、Hideaki Kanki、Shuji Kaneko 和 Atsushi Ichikawa：“前列腺素 E 受体 EP_1 与爪蟾卵母细胞中表达的 TRP5 的可能偶联”Biochem。

Kinoshita, M. et al.: "Binding of G α o is responsible for the voltage-resistant inhibition of α_<1A>1A(P/Q-type, Cav2.1 ) Ca^<2+> channels"The Journal of Biological Chemistry. 276. 28731-28738 (2001)

Kinoshita, M. 等人：“G α o 的结合负责 α_<1A>1A（P/Q 型，Cav2.1 ）Ca^<2+> 通道的耐电压抑制”The Journal of生物化学。276。28731-28738（2001）

Kume T.et al.: "Isolation of a diprenoid substance with potent neuroprotective activity from fetal calf serum"Proceedings of the National Academy of Sciences USA. 99. 3288-3293 (2002)

Kume T.等人：“从胎牛血清中分离出具有有效神经保护活性的二烯类物质”美国国家科学院院刊。

Tabata, H. et al.: "Possible coupling of prostaglandin E receptor EP1 to TRP5 expressed in Xenopus laevis oocytes"Biochemical and Biophysical Research Communications. 298. 398-402 (2002)

Tabata, H. 等人：“前列腺素 E 受体 EP1 与非洲爪蟾卵母细胞中表达的 TRP5 的可能偶联”生物化学和生物物理研究通讯。