In vitro embryo production via ovum pick-up (OPU) and intracytoplasmic sperm injection (ICSI) in pure and crossbred Japanese Hokkaido native ponies.

The research aims to assess the effectiveness of in vitro embryo production using ovum pick-up (OPU) and intracytoplasmic sperm injection (ICSI) techniques in breeding Japanese Hokkaido native ponies.

Methodology

• The study involved 9 pure and crossbred Hokkaido native ponies.
• Oocytes, or female egg cells, were collected via the transvaginal ultrasound-guided follicle aspiration technique, also known as ovum pick-up (OPU).
• These collected oocytes were then matured in vitro, that is, outside the living organism in a controlled lab setting.
• Intracytoplasmic sperm injection (ICSI) was then performed on these matured oocytes. In ICSI, a single sperm cell is injected directly into the cytoplasm of the egg using a microneedle. The sperm used for the study was frozen or cryopreserved.

Embryo Culture and Transfer

• The resulting embryo cultures were monitored using time-lapse cinematography, which provides detailed and continuous visual tracking of embryo development.
• The developed blastocysts, an early stage embryo, were then cryopreserved for future use. Cryopreservation is the process of preserving cells or whole tissues by cooling them to sub-zero temperatures.
• Post-thawing, these blastocysts were non-surgically transferred into recipient mares.

Results

• The research showed that on average, 23.9 follicles could be aspirated in each of the nine OPU sessions, with the number ranging from 13 to 49.
• The oocyte recovery rate was 35.3% (76 out of 215), indicating that about a third of the oocytes could be successfully retrieved from the aspirated follicles.
• The oocyte maturation rate was 61.5% (40 out of 65), showing that about two-thirds of the retrieved oocytes could be matured successfully in vitro.
• The cleavage rate, or the rate at which the fertilized oocytes started dividing into multiple cells, was 57.5% (23 out of 40).
• Of these cleaved embryos, about 56.5% (13 out of 23) were arrested at the 4-cell to 8-cell stage, that is, their development ceased at this stage.
• However, five embryos still managed to develop into early blastocysts.
• Three of these resulting embryos were successfully transferred leading to a successful pregnancy.

Conclusion

The study concluded that the use of OPU-ICSI is a viable assisted reproductive technology for boosting the population of Japanese native horses. The method demonstrated a fair success rate in terms of oocyte recovery, in vitro maturation, and resulting pregnancies.