Synchronization of the ovulation and copulation timings increased the number of in vivo fertilized oocytes in superovulated female mice

Abstract

The number of sperm that reaches the oocytes in mammalian species is limited. In mice, 8-10 oocytes are ovulated, a similar number of sperm reaches the oocytes, and nearly all oocytes are fertilized via natural mating. Meanwhile, our improved superovulation technique (ultrasuperovulation: administration of inhibin antiserum and equine chorionic gonadotropin [IASe]) produced 100 oocytes from a single female C57BL/6 mouse but resulted in only approximately 20 fertilized oocytes via mating. We hypothesized that sperm shortage in the ampulla might cause this low fertilization rate. Mice were mated in the proestrus stage or after hormone injection, but ovulation timing was not considered. In clinical application, the rhythm method supports fertilization by testing the ovulation period and synchronizing the ovulation and copulation timings. Therefore, this study examined the effects of ovulation and copulation timings on in vivo fertilization in female mice with IASe. Synchronization of the ovulation and copulation timings increased fertilization efficiency in female mice with ultrasuperovulation. The number of embryos obtained post ovulation was three times higher than that obtained pre ovulation. This study suggests that synchronized ovulation and copulation timings improve the efficiency of in vivo fertilization in IASe-treated female mice. This technique can be used to produce genetically modified mice and develop technologies for infertility treatment.

Fig 1. Effect of synchronizing the timings of ovulation and copulation on the fertilization rate in vivo.

Female mice (4 weeks old) treated with IASe were divided into three groups: pre ovulation (0–10 h after hCG injection), during ovulation (10–15 h after hCG injection), and post ovulation (15–19 h after hCG injection). These female mice were then copulated with male mice in each period. (A) The successful copulation rate was calculated using the following equation: Successful copulation rate (%) = total number of plugged female mice / total number of female mice used for copulation × 100. (B) The fertilization rate was calculated using the following equation: fertilization rate (%) = total number of two-cell embryos / total number of collected oocytes and two-cell embryos × 100. (C) The average number of fertilized oocytes obtained from each female mouse with a plug. (D) The efficiency was the average number of fertilized oocytes obtained from each female mouse used for copulation. Values are presented as the mean ± SD (n = 15–21). *p < 0.05 compared with pre ovulation.