Studies on steroidogenesis in the oocyte

Abstract

The steroid hormone has an important role in the early stages of reproduction. There has been abundant histochemical evidence that oocytes contain steroid hormones and are able to synthesize these hormones. But there have been few methods of analyzing one oocyte biochemically because it is too small and light. In order to study steroidogenesis in the oocyte, a microassay method sensitive enough to analyze the enzyme activities in one oocyte was developed using enzymatic cyling for amplifying the reaction product to 10,000-fold. An oil-well technique and a microtube method were applied in the assay for achieving the reaction in a medium as small as 1.0 to 5.0 microliters under a stereomicroscope. Immature Wistar rats were superovulated by PMS-hCG administration. Oocytes were collected by puncturing the follicle and flushing the tube. They were freeze-dried after washing to remove cumulus cells. The dry weight of one oocyte was 51.2 +/- 6.2 ng in a quartz fiber fishpole balance. The activity of 3 beta hydroxysteroid dehydrogenase (3 beta HSD) (picomol/oocyte/hr, substrate:pregnenolone) in the PMS-treated oocyte was 2.66 +/- 0.59, which corresponds to 3 times the activity of the ovarian homogenate as control, indicating the high capacity of oocytes to produce progesterone. The activity increased significantly (P less than 0.01) by hCG administration up to 4.17 +/- 0.29 after ovulation, suggesting that gonadotropin regulates steroidogenesis in the oocyte. The activities of G6PD and 6PGD were 8.41 +/- 1.09 picomol/oocyte/min and 3.85 +/- 2.02 picomol/oocyte/hr, respectively. The high activity of G6PD (more than 10 times that of the ovarian homogenate) suggests that the pentose phosphate shunt concerned with steroidogenesis is active in the oocyte. HCG decreased the activities of both G6PD and 6PGD. The present results show that steroidogenesis in the oocyte is very active under the control of gonadotropin, suggesting that steroid hormones may play an important role in oocyte maturation, ovulation and fertilization.