RATIO OF INNER CELL MASS AND TROPHOBLASTIC CELLS IN DEMI AND INTACT PIG EMBRYOS

Pig morulae, early blastocysts and blastocysts were microsurgically bi sected to produce zona-kee demi-embryos or remained nonbisected with o r without zona pellucida, and the presence of inner cell mass cells wa s determined using a differential fluorochrome staining technique. Aft er 24 h of in vitro culture, all demi-embryos were classified into thr ee categories, based on morphological criteria: 1, excellent; 2, fair; and 3, degenerated. The average number of total cells and inner cell mass cells in intact embryos cultured without zona pellucida for 24 h was higher (P < 0.05) than that for those with zona pellucida in morul ae and early blastocysts. The percentage of demi-embryos without inner cell mass cells in these different morphological categories was 18.7% , 22.2% and 29.8% for morulae, respectively; 3.8%, 16.7% and 30.8% for early blastocysts, respectively; and 3.7%, 32.0% and 36.4% for blasto cysts, respectively. The percentage of demi-embryos without inner cell mass cells was lower (P < 0.01) in demi-embryos classified in categor y 1 compared with category 3 in early blastocysts and in category 1 co mpared with categories 2 and 3 in blastocysts. Significant differences in the total number of cells and the number of inner cell mass cells were apparent among the three morphological categories of demi-embryos derived from morulae, early blastocysts and blastocysts. The ratio of total cells to inner cell mass cells was similar among intact pig emb ryos and the different morphological categories of demiembryos derived from morulae, early blastocysts and blastocysts, with the exception o f that between demi-blastocysts of category 1 and the other groups. Th e loss of cells attributed to bisection was 25-30% in category 1 demi- embryos, and increased to 65% in category 3 demi-embryos. Demi-embryos in category 1 derived from early blastocysts and blastocysts experien ced no loss of inner cell mass cells. It is concluded that (i) bisecti on of pig embryos can lead to a substantial proportion of demi-embryos lacking a functional inner cell mass, (ii) the proportion of total ce lls to inner cell mass cells is similar in demi-embryos and intact emb ryos, and (iii) the percentage of cell losses attributed to bisection increases with decreasing quality of demi-embryos. Collectively, these results indicate that the blastocyst is the optimal stage to obtain a maximum yield to viable pig demi-embryos.