Inositol 1,4,5,-triphosphate activated Calcium Channels in oocytes.

肌醇 1,4,5,-三磷酸激活卵母细胞中的钙通道。

• 批准号: 01570043
• 负责人: KURODA Hideyo
• 金额: $ 1.34万
• 依托单位: Nagoya University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1989
• 资助国家: 日本
• 起止时间: 1989 至 1990
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-01570043/
• 关键词: IP_3; IP_3 receptor; calcium release; calcium channel; sea urchin egg; fertilization; イノシト-ルミりん酸; イノシト-ル三りん酸

A transient increase in intracellular calcium concentration occurs in sea urchin eggs during fertilization. Inositol 1,4,5,-triphosphate(IP_3) is thought to be the second messenger which activates calcium channels of intracellular store and to cause the Ca-transient. However, there is no direct evidence for such a IP_3-activated Ca channel in sea urchin eggs. These experiments aimed to show the evidences.1. IP_3 caused a transient release of Ca^<2+> from microsomal fraction of unfertililized eggs. The release was inhibited by the application of heparin (1 mg/ml). We found in addition, that caffein also caused a transient release of Ca^<2+> from the microsomal fraction in dose dependent manner. Heparin did not block the release. This means that caffeine causes a Ca release via a different mechanism from the IP_3-induced Ca release.2. We tried to measured the current passing a IP_3-induced Ca releasing channel. Single channel recordings of the microsomal membrane which fused with a planar bilayer membrane were examined. Several kinds of Ca^<2+>, K^+ and Cl^- channels were found but no IP_3-induced Ca channel was observed.3. Another approach to show the existence of IP_3- induced Ca releasing channel must be the purification of "IP_3-receptors" from the microsomal fraction. IP_3 binding proteins of about 200 KD were obtained. The reconstitution of the protein into an artificial membrane is now examining.

在受精过程中，海胆卵细胞内钙离子浓度会短暂增加。肌醇1，4，5-三磷酸（Inositol 1，4，5，-triphosphate，IP_3）被认为是激活细胞内钙通道并引起钙瞬变的第二信使。然而，没有直接证据表明，这种IP_3激活的Ca通道在海胆卵。这些实验旨在展示证据。IP_3引起未受精卵微粒体部分的Ca^<2+>瞬时释放。通过应用肝素（1 mg/ml）抑制释放。此外，我们还发现咖啡因还能引起微粒体组分中Ca^<2+>的瞬时释放，并呈剂量依赖性。肝素未阻断释放。这意味着咖啡因通过与IP_3诱导的Ca释放不同的机制引起Ca释放。我们尝试测量IP_3诱导的钙释放通道的电流。对与平面双层膜融合的微粒体膜进行了单通道记录。结果表明，IP_3诱导的Ca ~（2+）、K ~+和Cl ~-通道均存在，但未发现IP_3诱导的Ca ~（2+）通道.另一个证明IP_3诱导的Ca释放通道存在的方法是从微粒体组分中纯化IP_3受体。获得了分子量约为200 KD的IP_3结合蛋白。将这种蛋白质重组成人工膜的方法正在研究中。

项目成果

R.Kuroda,H.Kuroda,T.Murai and Y.Imae: "Purification ofinositol 1,4,5,-triphosphate receptors from sea urchin eggs." Dev.Biol.

R.Kuroda、H.Kuroda、T.Murai 和 Y.Imae：“从海胆卵中纯化肌醇 1,4,5,-三磷酸受体”。

H.Soga,S.Takeda,T.Soga,R.Kuroda and H.Kuroda: "Inositol triphosphateーstimulated Ca^<2+> release from microsome fractions of sea urchin eggs." Biochim.Biophys.Acta.

H.Soga、S.Takeda、T.Soga、R.Kuroda 和 H.Kuroda：“三磷酸肌醇刺激海胆卵微粒体部分释放 Ca^<2+>”。

Obata,S.and Kuroda,H.: "Voltage-independent component of the fertilization current in sea urchin eggs." Submitted to Dev.Biol.

Obata,S. 和 Kuroda,H.：“海胆卵受精电流的电压无关部分。”

Obata, S. and Kuroda, H.: "Membrane potential change in a sea urchin egg induced by calcium ionophore A23187 has the same component that is involved in the fertilization potential." Cell Struct. Funct.14. 697-706 (1989)

Obata, S. 和 Kuroda, H.：“钙离子载体 A23187 诱导的海胆卵膜电位变化与受精电位中涉及的成分相同。”

R.Kuroda, H.Kuroda, T.Murai and Y.Imae.: "Purification of inositol 1,4,5,-triphosphate receptors from sea urchin eggs." Dev. Biol.

R.Kuroda、H.Kuroda、T.Murai 和 Y.Imae.：“从海胆卵中纯化肌醇 1,4,5,-三磷酸受体。”