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 configuration of oocytes isolated from late preantral and early antral bovine ovarian follicles immediately after recovery and after the in vitro culture. Individual follicles were isolated by microdissection from slices of the ovarian cortex. Follicles were sorted by diameter into 175 to 224, 225 to 274 and 275 to 325 microm-size classes. The follicles selected for in vitro culture were placed singly into 40 microL droplets of medium (TCM 199 enriched with FCS, insulin, transferrin, sodium selenite, sodium pyruvate, 1-glutamine, hypoxanthine, FSH and estradiol-17beta) and cultured for 6, 8, 11, 14 or 17 d. The sizes of follicles and oocytes were related to the duration of culture and gradually increased as culture duration was prolonged. The analysis of the relationship between mean diameters of oocytes at the time of recovery and after the in vitro culture, has shown significant differences after culture lasting 8 d (76.9+/-9.9 vs. 86.1+/-11.1 microm; P < 0.05), 11 d (77.0+/-9.9 vs. 91.9+/-17.5 microm; P < 0.01), 14 d (80.0+/-9.5 vs. 97.9+/-16.5 microm; P < 0.01) and 17 d (82.6+/-6.6 vs. 97.2+/-11.5 microm; P< 0.01). No statistical differences were shown among oocytes in the 5 pre-culture groups (79.5+/-8.8; 76.9+/-9.9; 77.1+/-9.9; 80.1+/-9.5 and 82.6+/-6.6 microm). Meiotic arrest was preserved in 71.9% of oocytes in our culture system up to 14 d. Frequency of the germinal vesicle (GV) stage did not significantly differ among oocytes evaluated "fresh" or cultured for 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.
Katska
L
L
Department of Animal Reproduction, National Research Institute of Animal Production, Balice/Kraków, Poland. lkatska@izoo.krakow.pl
Alm
H
H
Ryńska
B
B
D000818:Animals; D002417:Cattle / Q000502:physiology*; D002843:Chromatin / Q000502:physiology; D004396:Coloring Agents / Q000737:chemistry; D005260:Female; D008540:Meiosis / Q000502:physiology; D008858:Microscopy, Phase-Contrast / Q000662:veterinary; D009865:Oocytes / Q000166:cytology / Q000502:physiology*; D009924:Organ Culture Techniques / Q000379:methods / Q000662:veterinary*; D006080:Ovarian Follicle / Q000166:cytology / Q000502:physiology*; D010078:Oxazines / Q000737:chemistry; D015044:Zona Pellucida / Q000502:physiology
D002843:Chromatin; D004396:Coloring Agents; D010078:Oxazines; C018437:aceto-orcein