CONSTRUCTION OF A MONITERING SYSTEM OF ENDOCRINE DISRUPTING SUBSTANCES USING MEDAKA

实施本项目的目的是开发一套可靠的系统，利用青竹测试环境内分泌干扰物的影响。宽叶大米。我们可以重申青竹作为雌激素物质存在的测试有机体的有效性，不仅是作为实验动物，也是作为野生动物。取得了以下研究成果：1.利用Y染色体上的遗传标记，可以准确区分给定个体的遗传性别。利用该标记对野生青竹进行筛选，共鉴定出3个性别分化突变。由内分泌干扰物引起的性逆转也是可以识别的。阐明了野生青竹血清中卵黄蛋白原含量的年变化。在雌性中，卵黄蛋白原在春夏季较高，秋冬季较低。在男性中，我们全年都检测不到卵黄蛋白原。这些结果表明，野生青竹的卵黄蛋白原含量是雌激素类物质对环境污染的良好指标。用性变化率和卵黄蛋白原含量两个不同的指标，评价了青竹近交系对雌激素的敏感性。性激素诱导的性逆转比率因品系不同而不同。同基因菌株和重组菌株的使用表明，性别反转率的差异可以归因于与Y染色体上的性别决定基因密切相关的遗传因素。我们还研究了雌二醇处理鱼血清中卵黄蛋白原含量的剂量-反应关系和时程变化。雄鱼在用药后7天内出现卵黄蛋白原，并存在一定的剂量依赖关系。不同菌株间卵黄蛋白原含量无明显差异。

The present project was performed in order to develop a reliable system for testing the effects of environmental endocrine disruptors using medaka. Oryzias latipes. We could reaffirm the validity of the medaka as a test organism for the presence of estrogenic substances, not only as a laboratory animal but also as a wild life. The following results were obtained.1. Using a genetic marker on Y chromosome, the genetic sex of a given individual become able to be distinguished exactly. From the screening of wild medaka by this marker, we identified 3 mutations as to the sex differentiation. The sex reversal induced by endocrine-disruptors can also be identified.2. Annual change of vitellogenin content in the serum of wild medaka was clarified. In females, vitellogenin was high in spring and summer, and low in autumn and winter. In males, we could not detected vitellogenin throughout the year. These results indicate that the vitellogenin content of wild medaka is a good indicator of the contamination of estrogenic substances to environment.3. Using two different indicators, the rate of sex reversal and the vitellogenin content, the sensitivity to estrogens of inbred medaka strains was evaluated. The rates of the sex-reversal induced by sex steroids varied among strains. The use of congenic strains as well as recombinant strains clarified that the difference in the rate of sex-reversal can be attributed to a genetic factor that is strongly linked to the sex-determining gene on Y chromosome. We also examined the dose-response relationship and a time-course change of the vitellogenin content in the serum of fish treated with estradiol. Vitellogenin appeared in male fish within 7 days of treatment and a definite dose-dependent relationship was identified. There was no obvious difference in the content of vitellogenin among strains used.