Nuclear configuration of bovine oocytes derived from fresh and in vitro-cultured preantral and early antral ovarian follicles

The aim of this experiment was to characterize the growth and nuclear confi guration of oocytes isolated from late preantral and early antral bovine ov arian follicles immediately after recovery and after the in vitro culture. Individual follicles were isolated by microdissection from slices of the ov arian cortex. Follicles were sorted by diameter into 175 to 224, 225 to 274 and 275 to 325 mu m-size classes. The follicles selected for in vitro cult ure were placed singly into 40 mu L droplets of medium (TCM 199 enriched wi th FCS, insulin, transferrin, sodium selenite,,odium pyruvate, 1-glutamine, hypoxanthine, FSH and estradiol-17 beta) and cultured for 6, 8, 11, 14 or 17 d. The sizes of follicles and oocytes were related to the duration of cu lture and gradually increased as culture duration was prolonged. The analys is of the relationship between mean diameters of oocytes at the rime of rec overy and after the in vitro culture, has shown significant differences aft er culture lasting 8 d (76.9 +/- 9.9 vs. 86.1 +/- 11.1 mu m; P < 0.05), 11 d (77.0 +/- 9.4 vs. 91.9 +/- 17.5 mu m; P < 0.01), 14 d (80.0 +/- 9.5 vs. 9 7.9 +/- 16.5 mu m; P < 0.01) and 17 d (82.6 +/- 6.6 vs. 97.2 +/- 11.5 mu m; P < 0.01). No statistical differences were shown among oocytes in the 5 pr e-culture groups (79.5 +/- 8.8; 76.9 +/- 9.9; 77.1 +/- 9.9; 80.1 +/- 9.5 an d 82.6 +/- 6.6 mu m). Meiotic arrest was preserved in 71.9% of oocytes in o ur culture system up to 14 d. Frequency of the germinal vesicle (GV) stage did not significantly differ among oocytes evaluated "fresh" or cultured fo r 6, 8, 11 or 14 d. No relationship was observed between the size class of follicles and the frequency of the GV-stage. Prolonging the culture period to 17 d drastically decreased the percentage of oocytes in the GV-stage (18 .7%) and increased the percentage of oocytes having premature initiation of meiosis (GVBD; 46.3%) and degeneration (25.0%). These results suggest that out of all culture periods used in our experiment, Day 14 was found to be the longest culture time allowing for both oocyte growth and maintenance of nuclear configuration at the GV-stage. (C) 2000 by Elsevier Science inc.