Nucleolar ultrastructure and RNA synthesis in cow preimplantation embryos

Early preimplantation embryos (4-8 blastomeres) collected from superovulated cows at slaughter by flushing of oviducts and uterine horns were treated for fine-structural morphological and autoradiographic analysis of nucleologenesis and RNA synthesis. Early cow embryos were cultured for 30 min in Dulbeco PSB culture medium (SERVA, Heidelberg, FRG) with addition of 100 micro Ci/ml of (5-sup(3)H) uridine, specific activity 740 Bg/mM (VVVR, Prague, Czech Republic), pH=7.2-7.4. Following this, the embryos were fixed by 3 % glutaraldehyde and postfixed by 1 % OsO4, dehydrated by alcohol and set in EPON 812 resin. Semi-thin sections were coated with liquid nuclear emulsion Ilford K.5 and exposed in dark boxes at 5 deg C for 1 month. After development in Kodak D.19 and Methylene Blue staining the slides were observed by light-microscopy. RNA synthetic activity was assessed by Ag-grains labeling of nuclei and nucleoli, respectively, of 4-8 cell embryos. It was possible to conclude, based on fine-structure morphological and autoradiolographic studies, that early preimplantation cow embryos (4-8 blastomeres) at the begining of compactization exhibit dynamic changes in nucleolus structure. Nucleologenesis in this species may be divided into four stages starting with the compact rounded Nucleolus Precursors Body (NPB) detected in 4-cell embryos (stage 1); in the next stage (2) a primary vacuole appears in NLB of 8-cell embryos. This processes by formation of secondary vacuoles in NLB (stage 3) in late 8-cell embryos. At the end of the 8-cell stage a functional fibrillo-granular nucleolus is differentiated (stage 4). RNA synthetic activity was localized to the stages 3 and 4 based on (5-sup(3)H) uridine incorporation. Extrapolating from recent research on the goat embryo (Pivko et al 1995) a similar type of nucleologenesis in all ruminants is suggested