P-64: The Effect of Ovine Oocyte Diameter on Nuclear Maturation

Background: The study was carried out to investigate the relationship between the development potential and the diameter of ovine oocytes. Materials and Methods: Ovaries collected from a local abattoir were transported to the laboratory within 1-3 h of slaughter. The oocytes from follicles (2-6 mm in diameter) were recovered by aspiration and stored in a preincubated (at 38.6 C, 5% CO2 and maximum humidity) hepesmodified TCM 199 solution. Good quality oocytes (evenly granulated cytoplasm with at least two layers of surrounding cumulus cells) were selected and subjected to culture in TCM 199 supplemented with 0.05 IU/ml rhFSH, 1 IU/ ml hCG and 1 μg/ml E2 (OCM; oocyte culture medium). Before culturing, the selected oocytes (n= 1495) were divided into three categories based on diameter: i. <110 μm, ii. 110-150 μm and iii. >150 μm. In each category half of the oocytes were denuded of the cumulus cells and the rest remained intact (cumulus enclosed oocytes). All of the oocytes, denuded and cumulus enclosed oocytes in each category were cultured in OCM for a period of 26- 27 hours. After the incubation period, the nuclear status of the oocytes in each experimental group was assessed using a 2% orcein staining technique. Results: The percentage of oocytes that reached the MII stage was 81, 82, and 84% for the cumulus enclosed oocytes with diameters of <110, 110-150, >150 μm, respectively. The corresponding values for the denuded oocytes were 4, 5, and 6%, respectively. The oocytes displayed no size-related ability to undergo meiotic maturation in both cumulus enclosed and denuded oocytes. However, as expected the rate of nuclear maturation in cumulus enclosed oocytes was higher than that of denuded oocytes in all the size categories. Conclusion: The results suggest that in sheep the antral follicles ranging from 2 to 6 mm contain fully grown oocytes which, despite their variability in diameter, show good competence for in vitro nuclear maturation.