Heat stress is a major contributing factor to low fertility among dairy cattle, as reflected by the dramatic reduction in conception rate during the hot months. The effects of thermal stress on oocyte competence and embryonic development have been well documented. However, timing of embryonic cleavage, which may be considered a parameter for the identification of good-quality embryos, and its association with elevated temperatures have not been studied. Two experiments were performed to examine and characterize seasonal effects (i.e. thermal stress) on cleavage timing of bovine parthenogenetic embryos. Oocytes were aspirated from ovaries collected at the local abattoir in 2 seasons: cold (Dec–Apr) and hot (May–Nov). Matured oocytes were chemically activated (ionomycin followed by 6-DMAP) and cultured in vitro; cleavage timing to the 2- and 4-cell stages was observed and documented. The one-way ANOVA procedure was used for statistical analysis. In the first experiment (n = 5416 oocytes), cleavage was documented at specific time points during development post-activation. The peak in embryonic development to the 2-cell stage was earlier (22 to 27 vs. 27 to 40 h after activation) and the cleavage rate higher (39 vs. 21%; P < 0.0001) during the cold season relative to the hot season, respectively. Similarly, the peak in 4-cell-stage development was also observed earlier (46–52 vs. 52–70 h after activation) and corresponded with a higher proportion of developing embryos (33 vs. 21%; P < 0.0001) during the cold season as compared to the hot season, respectively. These results indicate that embryonic development is delayed and a lower proportion of embryos cleaved during the hot season. To better understand the delay in cleavage timing, a second experiment (n = 308 oocytes) was performed through two consecutive hot seasons. A time-lapse system (EmbryoGuard; IMT, Ltd., Ness-Ziona, Israel) was employed to collect accurate data on the first cleavage division, known to be indicative of embryo quality. The time-lapse system was pre-programmed to take photos at 1-h intervals such that culture dishes did not need to be removed from the incubator. Similar to the pattern noted for the hot season in the first experiment, a wide distribution of cleavage timing (18-40 h after activation) was observed. Further analysis revealed that embryos cleaved in 2 distinct waves: cleavage timing of the first wave (18 to 25 h after activation) was characterized by a time frame similar to that in the cold season, suggesting good-quality embryos; however, the second wave, from 27 to 40 h after activation, presented a delay in cleavage timing, suggesting that these late-cleaving embryos are of inferior quality. Taken together, the results of the 2 experiments lead to the assumption that oocytes harvested from lactating cows during the hot season are of reduced developmental potential, which may be explained, in part, by the pattern of 2 cleavage waves. Furthermore, cleavage timing appears to be a good indicator of embryo potential and may increase the chances of selecting better in vitro-derived embryos during the hot season for embryo transfer.
Effects of heat stress during in vitro maturation on cytoplasmic versus nuclear components of mouse oocytes
