P-108: Effect of Phosphodiesterase Type 3 Inhibitor on Nuclear Maturation and In Vitro Development of Ovine Oocytes

Background: Present study aims to investigate if PMC of ovine oocytes in the presence of cilostamide a phosphodiesterase type 3 (PDE3) inhibitor can improve the shortcomings of conventional IVM system. Materials and Methods: Immature ovine oocytes were retrieved from pre antral follicles of abattoir ovaries by aspiration and after washing in Htcm + %10 FBS were immediately classified in four groups: three groups were cultured in PMC medium in the presence of 1, 10 and 20 μm cilostamide for 22 hours to arrest meiotic maturation and then they were washed and transferred to MM to resume meiotic maturation. In control group, oocytes were cultured in MM in the absence of cilostamide. The effect of cilostamide on gap junction communications and nuclear status was studied. The variables assessed were chromosome organization, spindle pattern, polar body extrusion, and embryonic development. Results: According to the results, inhibition of PDE3 could not permanently block nuclear maturation in ovine oocytes but it delayed the process of nuclear maturation. Elevation of intra-oocyte cAMP concentration could inhibit cumulus cells expansion and maintain gap junction communications between oocyte and cumulus cells. Deletion of cilostamide and refreshing maturation medium after 22 hours culture revealed that cumulus cells were completely expanded. The inhibitory effect induced by 1 μM cilostamide was reversible, and it increased the number of mature oocytes with aligned chromosomes and normal spindle. However, the inhibitory effects of 10 and 20 μM cilostamide was not fully reversible and was associated with deleterious effects on chromosome organization and spindle pattern. Investigation of embryonic development via parthenogenetic activation and in vitro fertilization revealed that the blastocyst rate of oocytes that were prematured with 1 μM cilostamide was not significantly different from oocytes that underwent conventional IVM but it was significantly reduced in oocytes that were prematured with 10 and 20 μM cilostamide. Conclusion: Our results provide the evidence that reduced cAMP via PDE3 is not the only mechanism that controls the progress of nuclear maturation in sheep oocytes, and that alternative or additional mechanisms may also exist.