The maturation in vitro of oocytes (MIV) is an important reproduction technique that allows the use of oocytes collected from follicles in different stage of development that, probably, desapeared in vivo due to subordination or atresia. However, in herd, when oocyte-cumulus complex (COCs) are removed from follicles and in vitro cultures and when in good conditions, they re-start meiosis spontaneously and stop citoplasmatic maturation necessary to afford further embryo development. One strategy to increase the potential development of oocytes in vitro is to temporarelly block the nuclear development with the use of pharmacological meiotic inhibitors. This research aim to evaluate and compare the efficiency of Roscovitine and Cycloheximide in synchronize nuclear and citoplasmatic maturation. To reach this objective, COCs from ovaries of adult ovines, obtained from slaughterhouse, will be cultivated in an specific medium (TCM 199 supplemented with 0,1UI/mL FSH, 0,1 UI/mL hCG, 1µg/mL estradiol, 0,3 mM sodium pyruvate, 75 µg/mL de streptomycin, 10 % SFB e 100 µM/mL cysteamine) without drugs added (check treatment) or with 50 ¼M of Roscovitine (treatment 2) added; or 2 ¼g/mL of Cycloheximide (treatment 3) added for 22-24 hours at 39ºC in 5% CO2 atmosphere. The reversable action of these drugs will be evaluated after COCs cultive in a maturation medium free of inhibitors and in the same conditions (as described before). After in vitro culture, COCs will be analyzed according to cells cumulus capacity of increasing under stereomicroscope observation, to nuclear maturation under fluorescence microscope and to citoplasmatic maturation, based on the distribution patterns of cytoplasmic organelles, under electronic transmission microscope.
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