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

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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.