Analysis of Fertilization Phenomenon Using the Patch Clamp Method.

本研究旨在利用电生理领域新发展起来的膜片钳法对施肥现象进行分析。(1)金仓鼠卵的研究我们在金仓鼠皮下组织注射PMS和hCG。显微镜下收集高刺激卵，在含胰蛋白酶和杂硝基肼溶液中孵育。将收集的无带卵子与膜片钳电极连接，记录并分析钙离子通道的开关现象。通过对无精子溶液中钙离子通道的研究，将金仓鼠精子加入溶液中。在相差显微镜下，精子进入卵子后，钙离子通道的开关频率增加。这些数据已经用其他方法推测出来了，但重要的是直接得到这些数据。(2)人类卵子的研究由于目前日本妇产科协会对人类卵子受精的研究有限制，因此无法对人类卵子进行精确的研究。我们在患者同意的情况下检查了在妇科手术中收集的人类卵子。用膜片钳电极吸附经胰蛋白酶和杂环肼处理后的无带卵。卵泡期钙离子通道开关频率低，黄体期频率高。这些数据是新发现的。

This research aimed at the analysis of fertilization phenomenon using the patch clamp method which is newly developed method in electro-physiology area.(1) Study of golden hamster ovumWe injected PMS and hCG in subcutaneous tissue of golden hamster. Hyperstimulated ova were collected using microscope and incubated in trypsin and hyaronidaze contained solution. Collected zona free ovum was attached with patch clamp electrode, and on-off phenomena of calcium ion channel were recorded and analyzed. After study of calcium ion channel in sperm free solution, sperm of golden hamster were added into solution. Frequency of on-off phenomena of calcium ion channel were increased after penetration of sperm into ovum under phase contrast microscope. These data are already speculated in other methods, but it is important to get these data in direct method.(2) Study of human ovumBecause studies of fertilization of human ovum are restricted under Japanese Obstetrics & gynecology Association now, the precise studies of human ovum cannot be done. We examined human ova which were collected in gynecological operations under permission of patients. Zona free ovum which was obtained after treatment of trypsin and hyaronidaze was attached with patch clamp electrode. Frequency of on-off phenomena of calcium ion channel was low in follicular phase, but in luteal phase it was high frequency. These data are newly found out.