Synchronization protocols of oocyte population for OPU/IVM purpose in cattle

Oocytes isolated post mortem from bovine antral follicles are heterogeneous in chromatin configuration, gap- junction functionality between oocyte and cumulus cells, transcriptional activity and developmental competence. This heterogeneity could account for the low efficiency of current OPU/IVM strategies. We aimed to characterize the oocyte population obtained by OPU at a random day and to develop a protocol to homogenize the oocyte population destined to IVM/IVF. In experiment 1, 10 lactating Holstein cows were used in a crossover design. Treatments were: 1) OPU at a random day (Control); 2) aspiration of all visible follicles at a random day (D0), two IM injections of FSH (Folltropin; 56mg) 12h apart on D2, OPU from follicles >2mm on D4 (ASP-FSH/D4). In Experiment 2, 4 lactating Holstein/Girolanda cows were subjected to ASP-FSH with OPU on D5 (ASP-FSH/D5) and oocytes obtained on D0 were used as Control. Oocytes were fixed in 60% methanol/PBS, stained with Hoechst 33342 and examined by fluorescence microscopy to be classified according with the chromatin configuration in the germinal vesicle as GV0, GV1, GV2, GV3 or as GVBD when resuming meiosis. Data were arcsine transformed and groups compared by paired T test. Control oocytes from experiment 1 (n=90); were in GV0 (8.87%), GV1 (21.94%), GV2 (38.8%), GV3 (19.77%), GVBD (1.25%) and degenerated (9.75%). The protocol ASP-FSH/D4 (n=69 oocytes) abolished GV0, GVBD and degenerated oocytes, increased GV1 to 46.55% (P=0.03), tended to decrease GV3 (13.36%; P<0.1), but did not alter GV2 (40.09%). The number of oocytes recovered was not significantly different but recovery rate was lower in ASP-FSH/D4 (34.5% vs. 54.55%; P=0.01). The protocol ASP- FSH/D5 (n=57 oocytes) increased GV2 (80.76% vs. 47.17%) and decreased GV3 (9.72% vs. 34.5%; P<0.05) in comparison with the Control (n=83). In conclusion, oocytes recovered by OPU at a random day are heterogeneous and strategies combining follicle aspiration with FSH treatment can promote a more homogeneous population of oocytes for OPU/IVM purpose. Acknowledgements: Prof. José Luiz Moraes Vasconcelos, TECNOPEC-União Química.