The use of R-roscovitine to fit the 'time frame' on in vitro porcine embryo production by intracytoplasmic sperm injection
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Micromanipulation of oocytes is time consuming during ICSI experiments; however the 'time frame' to manipulate oocytes without a drop in efficiency is not very wide due to the use of not completely matured and/or aged MII oocytes. Therefore, the aim of this work was to study the effect of a short roscovitine pretreatment for 5 h and two different IVM periods (5R + 40IVM or 5R + 45IVM) and a prolonged IVM time from 45 h (45IVM) to 50 h (50IVM) on parthenogenetic and ICSI embryo development, in order to fit the time frame to manipulate pig oocytes to the whole labour day session. In the first experiment, oocytes, pretreated with roscovitine and IVM cultured for 5 h, showed a similar nuclear stage as non-cultured oocytes and a significantly higher percentage of GVI-GVII oocytes compared with non-roscovitine treated oocytes cultured for 5 h in IVM conditions. When COC were cultured tinder the 5R + 40IVM system, nuclear maturation and cleavage rates after electrical activation were significantly lower than when COC were cultured under the 45IVM, 50IVM and 5R + 45IVM culture systems (54.2% vs. 72.6-76.8% and 58.8% vs. 81.4-88.3%, respectively). However, this difference was not statistically significant for parthenogenote blastocyst rate. No differences were observed in MII and in parthenogenote and ICSI embryo development among 45IVM, 50IVM and 5R + 45IVM experimental groups. In conclusion, under our conditions and using parthenogenetic and ICSI embryos, we observed that it is feasible to prolong the pig oocyte manipulation 'time frame' by at least 5 h with no significant drop in blastocyst rate.