TOTAL PROTEIN-CONTENT AND PROTEIN-SYNTHESIS WITHIN PRE-ELONGATION STAGE BOVINE EMBRYOS

Protein content was measured in zona-free bovine oocytes and pre-elong ation stage embryos, following in vitro maturation, fertilisation, and then culture in Synthetic Oviduct Fluid medium supplemented with amin o acids and 8 mg ml(-1) bovine serum albumin (BSA). Values (ng embryo( -1)) of 122 +/- 7.8, 137 +/- 8.6, 111 +/- 8.8, 115 +/- 10.4, 139 +/- 9 .0 and 152 +/- 10.1 were obtained for zona-free mature oocytes, 2-cell (day 2), 8-cell (day 3), compact morula (day 6), blastocyst (day 7), and expanded blastocyst (day 8) stage embryos, respectively The protei n content of day 7 zona-enclosed blastocysts was 337 +/- 58.0 ng embry o(-1). These values suggest that prior to compaction and blastulation, the early cleavage stage bovine embryo has a higher rate of protein d egradation than that of synthesis. Net growth is observed only after i nitiation of compaction. The protein content of day 7 blastocysts was measured in embryos following in vitro production and culture in the s ame media supplemented with either 0.5% w/v polyvinyl alcohol (PVA), 8 mg ml(-1) BSA, 8 mg ml(-1) BSA and further supplemented with 10% feta l calf serum (FCS) from the beginning of culture (FCS-D1), 8 mg ml(-1) BSA and 10% FCS from the fourth day of culture (day 5 of development) or from in vivo-derived day 7 blastocysts. Protein content was signif icantly (P < 0.05) lower in PVA-cultured embryos than other treatments . To determine if this difference in PVA-cultured embryos was due to a difference in the rate of protein synthesis, comparisons were made be tween day 7 embryos derived from BSA-culture and either PVA-culture, F CS-D1 culture or in vivo-derived embryos. Despite differences in diame ter, no significant difference was observed in the incorporation of L- [2,3,4,5,6-H-3]-phenylalanine into the TCA-precipitable fraction in an y of the three comparisons made. However, incubation in the presence o f FITC-labelled BSA or beta-casein and examination under either fluore scence or confocal microscopy revealed that protein in the extraembryo nic environment was actively taken up by the trophectoderm of day 7 bl astocysts, most likely by endocytosis. These results suggest that exog enous protein is an important nutritive source, probably maintaining i ntracellular amino acid pools. Results obtained from the production of embryos in protein-free medium should be viewed with the knowledge th at such embryos differ metabolically from those embryos grown in the p resence of protein, including in vivo-derived embryos. (C) 1998 Wiley- Liss, Inc.