Effect of different parthenogenetic activation methods on the developmental competence of in vitro matured porcine oocytes

The aim of this study was to investigate the effect of electrical pulse, ethanol, and ionomycin combined with cycloheximide (CHX), cytochalasin B (CB), and 6-dimethylaminopurine (6-DMAP) on parthenogenetic developmental competence of in vitro matured porcine oocytes. In experiment 1, oocytes were treated with direct current electrical pulse (DC pulse) and then incubated in the NCSU- 23 medium supplemented with CHX, 6- DMAP, CB+CHX, and CB+6- DMAP for 6 h, respectively. The rate of blastocyst development in DC pulse+CB+6- DMAP group was significantly higher than those in other groups (42.4% vs 23.9% similar to 35.8%; P < 0.05); however, there were no differences in both of the cleavage rate and the cell number of blastocysts among four groups. In experiment 2, oocytes were treated with NCSU-23 medium containing 20 mu M ionomycin for 40 min and then incubated in the NCSU- 23 medium supplemented with CHX, 6-DMAP, CB+CHX and CB+6- DMAP for 6 h, respectively. The rates of cleavage and blastocyst development in ionomycin+6- DMAP group were higher than those obtained in other groups ( 66.2% vs 46.3% similar to 57.3%; 22.3% vs 7.4% similar to 16.1%; P< 0.05). In experiment 3, the activation effects of ethanol combined with 6- DMAP, CHX, CB+6- DMAP and CB+CHX were investigated. The rates of cleavage and blastocyst development in ethanol+CB+6-DMAP group were significantly higher than those in other groups ( 55.5% vs 42% similar to 46.2%; 18.0% vs 7.1% similar to 11.9%; P < 0.05). In experiment 4, the optimal activation protocols in each group plus DC pulse+ionomycin+6-DMAP were compared. The results showed the rates of cleavage in DC pulse+CB+6-DMAP group and ionomycin+6- DMAP were higher than those in ethanol+CB+6-DMAP and DC pulse+ionomycin+6-DMAP ( 73.8 - 74.4% vs 56.5 - 57.5%; P < 0.05), but the blastocyst development only in DC pulse+CB+6-DMAP group was significantly higher than that in other groups ( 34.1% vs 13.4% similar to 22.3%; P < 0.05). Total cell number of blastocysts in the group of DC pulse + ionomycin+6- DMAP was higher than that in other groups ( 34.1 vs 25.3 - 27.2; P < 0.05). In conclusion, DC pulse, ethanol, CB, and 6-DMAP all affected the parthenogenesis of porcine oocytes matured in vitro, but their combination of DC pulse+CB+6-DMAP showed the best result in both of cleavage and blastocyst development.