CONSTRUCTION OF A MONITERING SYSTEM OF ENDOCRINE DISRUPTING SUBSTANCES USING MEDAKA

本项目是为了开发一个可靠的系统来测试环境内分泌干扰物对medaka的影响。Oryzias latipes。我们可以重申medaka作为雌激素物质存在的测试生物的有效性，不仅作为实验动物，而且作为野生动物。得到了以下结果：利用Y染色体上的遗传标记，可以准确地区分给定个体的遗传性别。利用该标记对野生medaka进行筛选，鉴定出3个性别分化突变。由内分泌干扰物引起的性别逆转也可以被识别。阐明了野生medaka血清卵黄蛋白原含量的年变化。雌性卵黄原蛋白在春夏季较高，秋冬季较低。在雄性中，我们全年未检测到卵黄蛋白原。这些结果表明，野生medaka的卵黄原蛋白含量是衡量雌激素对环境污染的一个很好的指标。采用性别逆转率和卵黄原蛋白含量两种不同指标，评价了近交系medaka菌株对雌激素的敏感性。性激素诱导的性逆转率在不同品系之间存在差异。同源菌株和重组菌株的使用表明，性别逆转率的差异可归因于与Y染色体上的性别决定基因密切相关的遗传因素。我们还研究了经雌二醇处理的鱼血清中卵黄蛋白原含量的量效关系和时间变化。卵黄原蛋白在7天内出现在雄鱼体内，并确定了一定的剂量依赖关系。不同菌株的卵黄原蛋白含量无明显差异。

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.