228 IN VITRO EMBRYO PRODUCTION AFTER EXPOSURE OF BOVINE OOCYTES TO DIFFERENT TRANSPORTATION MEDIA AND PERIODS: PRELIMINARY RESULTS

Abstract

This study was designed to evaluate the effect of different oocyte transportation media and time. Immature oocytes were recovered from slaughterhouse bovine ovaries. Oocytes (n = 492) of quality grades I to III were randomly allotted to one of the following transportation media based on TCM-199, either buffered with HEPES (control) or buffered with NaHCO₃ and added with FSH/LH (maturing). Both media were supplemented with pyruvate, penicillin G (10 000 IU), streptomycin (0.05 mg mL^–1), and 10% fetal bovine serum (FBS). In control medium, the oocytes were kept for 1 or 8 h at 37°C, and thereafter were transferred to maturing medium until the maturation period (24 h) was completed, under controlled atmosphere (5% CO₂) and temperature, in an incubator. The maturing oocytes were distributed in two types of equipment developed for oocyte and embryo transportation, with and without 5% CO₂, and kept for 1, 8, or 24 h before maturation or fecundation (IVF) procedures. The oocytes kept for 24 h in the transportation device were placed directly for IVF. All procedures used for in vitro maturation, IVF, and in vitro culture were the same as those adopted for commercial in vitro embryo production at Biotran LTDA (Alfenas, Minas Gerais, Brazil). The cleavage rate was evaluated on Day 3 post-insemination, and the blastocyst production was evaluated on Day 7. The statistical model included the main effects of treatment (control and maturing with or without 5% CO₂), time, replicate, and the interaction of media × time. Data (3 replicates) were analysed by ANOVA and differences were identified by Tukey’s test. The time before in vitro maturation at the incubator negatively (P < 0.007) affected cleavage rates (76.4 ± 16.9 v. 58.8 ± 13.2 and 52.2 ± 18.5%, respectively, for 1, 8, and 24 h). However, treatment had no effects (P = 0.3) on cleavage (66.9 ± 15.0, 57.8 ± 19.7, and 67.7 ± 19.8% for the control and maturation with and without 5% CO₂, respectively). Similarly, blastocyst production rates differed (P < 0.04) between 1 h (33.4 ± 14.7) and 24 h (19.3 ± 17.0%), whereas blastocyst production at 8 h did not show significant effects (19.3 ± 17.0). Although it was not significant (P > 0.05), only 14.1% of the oocytes kept in medium 2 with a controlled atmosphere for 24 h became embryos. This difference was probably related to the trend in treatment effect (P = 0.06) and the lower rate of embryo production (22.2 ± 13.6%) in this treatment compared with the control (33.8 ± 18.1%) and maturation without 5% CO₂ (29.8 ± 17.3%). These results showed that 24 h of transportation is detrimental for the oocyte development potential and that the buffered medium used in this study with 5% CO₂ did not efficiently maintain embryo production.

Supported by CNPq, CAPES, FAPEMIG, and Biotran LTDA.