Skip to main content
SpringerLink
Log in

Ultrastructure, protein synthesis and secretion of day-6 rabbit blastocysts cultured in a chemically defined, protein-free medium

  • Original Articles
  • Published:
Anatomy and Embryology Aims and scope Submit manuscript

Summary

Day-6 rabbit blastocysts were cultured in Ham's F10 medium supplemented with polyvinylpyrrolidone as a macromolecular component, for 4 to 12 h. The integrity of the blastocyst cells was demonstrated by electron microscopy. Expansion and biosynthesis of proteins and of DNA were studied after culturing in the presence of 35S-methionine and 3H-thymidine. Polyvinylpyrrolidone did not interfere with the subsequent protein analysis, which was performed by two dimensional gel electrophoresis followed by silver staining and fluorography. More than 600 labelled proteins were found in the blastocyst tissue, many of them were also present in the blastocyst fluid and in the blastocyst coverings. Several proteins seemed to be produced for incorporation into the blastocyst coverings; others, only detected in the culture medium, might have been synthesized for secretion into the environment.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Similar content being viewed by others

References

  • Bartol FF, Roberts RM, Bazer FW, Lewis GS, Godkin JD, Thatcher WW (1985) Characterization of proteins produced in vitro by peri-attachment bovine conceptuses. Biol Reprod 32:681–693

    Google Scholar 

  • Blum H, Beier H, Gross HJ (1987) Improved silver staining of plant proteins, RNA and DNA in polyacrylamide gels. Electrophoresis 8:93–99

    CAS  Google Scholar 

  • Cholewa JA, Whitten WK (1970) Development of two-cell mouse embryos in the absence of fixed-nitrogen source. J Reprod Fertil 22:553–555

    Google Scholar 

  • Daniel JC Jr (1964) Some effects of steroids on cleavage of rabbit eggs in vitro. Endocrinology 75:706–710

    Google Scholar 

  • Daniel JC Jr, Levy JD (1964) Action of progesterone as a cleavage inhibitor of rabbit ova in vitro. J Reprod Fertil 7:323–329

    Google Scholar 

  • Dannhorn DR, Henkel R, Kirchner C (1988) Synthese von Uteroglobin in der Blastozyste des Kaninchens? Autoradiographische und immunocytochemische Untersuchungen. Fertilität 4:223–226

    Google Scholar 

  • Denker HW, Gerdes HJ (1979) The dynamic structure of rabbit blastocyst coverings. Anat Embryol 157:15–34

    Google Scholar 

  • El-Shershaby AM, Hinchliffe JR (1974) Cell redundancy in the zona-intact preimplantation mouse blastocyst: A light and electron microscope study of dead cells and their fate. J Embryol Exp Morphol 31:643–654

    Google Scholar 

  • Enders AC (1971) The fine structure of the blastocyst. In: Blandau RJ (ed) The Biology of the Blastocyst, University of Chicago Press, Chicago London, pp 71–94

    Google Scholar 

  • Fuchs AR, Beling C (1974) Evidence for early ovarian recognition of blastocysts in rabbits. Endocrinology 95:1054–1058

    Google Scholar 

  • Fujimoto S, Euker JS, Riegle GD, Dukelow WR (1975) On a substance crossreacting with luteinizing hormone in the pre-implantation blastocyst fluid of the rabbit. Proc Jpn Acad 51:123–125

    Google Scholar 

  • Godkin JD, Bazer FW, Moffatt J, Sessions F, Roberts RM (1982a) Purification and properties of a major, low molecular weight protein released by the trophoblast of sheep blastocysts at day 13–21. J Reprod Fertil 65:141–150

    Google Scholar 

  • Godkin JD, Bazer FW, Lewis GS, Geisert RD, Roberts RM (1982b) Synthesis and release of polypeptides by pig conceptuses during the period of blastocyst elongation and attachment. Biol Reprod 27:977–987

    Google Scholar 

  • Godkin JD, Bazer FW, Thatcher WW, Roberts RM (1984) Proteins released by cultured day 15–16 conceptuses prolong luteal maintenance when introduced into the uterine lumen of cyclic ewes. J Reprod Fertil 71:57–64

    Google Scholar 

  • Godkin JD, Bazer FW, Roberts RM (1985) Protein production by cultures established from day 14–16 sheep and pig conceptuses. J Reprod Fertil 74:377–382

    Google Scholar 

  • Görg A, Postel W, Weser J, Günther S, Hanash SM, Somerlot L (1987) Elimination of point streaking on silver stained two-dimensional gels by addition of iodoacetamide to the equillibration buffer. Electrophoresis 8:122–124

    Google Scholar 

  • Ham RG (1963) An improved nutrient solution for diploid chinese hamster and human cell lines. Exp Cell Res 29:515–526

    Google Scholar 

  • Imakawa K, Anthony RV, Kazemi M, Marotti KR, Polites HG, Roberts RM (1987) Interferon-like sequence of ovine trophoblast protein secreted by embryonic trophectoderm. Nature 330:377–379

    Google Scholar 

  • Jung T, Fischer B, Beier HM (1987) Quantitative aspects of protein synthesis in non-cultured and cultured rabbit blastocysts. Hum Reprod 2:23–27

    Google Scholar 

  • Kane MT (1983) Variability in different lots of commercial bovine serum albumin affects cell multiplication and hatching of rabbit blastocysts in culture. J Reprod Fertil 69:555–558

    Google Scholar 

  • Kane MT (1985) A low molecular weight extract of bovine serum albumin stimulates rabbit blastocyst cell division and expansion in vitro. J Reprod Fertil 73:147–150

    Google Scholar 

  • Kane MT (1987) Minimal nutrient requirements for culture of one-cell rabbit embryos. Biol Reprod 37:775–778

    Google Scholar 

  • Kane MT, Foote RH (1970a) Factors affecting blastocyst expansion of rabbit zygotes and young embryos in defined media. Biol Reprod 4:41–47

    Google Scholar 

  • Kane MT, Foote RH (1970b) Culture of two- and four-cell rabbit embryos to the expanding blastocyst stage in synthetic media. Proc Soc Exp Biol Med 133:921–925

    Google Scholar 

  • Kane MT, Headon DR (1980) The role of commercial bovine serum albumin preparations in the culture of one-cell rabbit embryos to blastocysts. J Reprod Fertil 60:469–475

    Google Scholar 

  • Karp G, Manes C, Hahn WE (1973) RNA-synthesis in the preimplantation rabbit embryo: radioautographic analysis. Dev Biol 31:404–408

    Google Scholar 

  • Kirchner C, Seitz KA (1972) Elektronenmikroskopische Untersuchungen über die Blastocyste des Kaninchens vor der Implantation in bezug auf ihre Wechselbeziehung zur uterinen Umgebung. Wilhelm Roux' Arch Entwicklungsmech Org 170:221–233

    Google Scholar 

  • Kuzan FB, Pomeroy KO, Seidel GE Jr (1982) Polyvinyl alcohol as a macromolecular substitute for bovine serum albumin in mouse embryo culture medium. Biol Reprod [Suppl] 26:65A

    Google Scholar 

  • Leiser R, Denker HW (1988) The dynamic structure of rabbit blastocyst coverings. II. Ultrastructural evidence for a role of the trophoblast in neozona formation. Anat Embryol 179:129–134

    Google Scholar 

  • Marshall T (1983) Artefacts associated with Nonidet P-40 upon sodium dodecyl sulphate polyacrylamide gel electrophoresis of protein. Electrophoresis 4:436–438

    Google Scholar 

  • Maurer RR, Beier HM (1976) Uterine proteins and development in vitro of rabbit preimplantation embryos. J Reprod Fertil 48:33–41

    Google Scholar 

  • Maurer RR, Onuma H, Foote RH (1970) Viability of cultured and transferred rabbit embryos. J Reprod Fertil 21:417–422

    Google Scholar 

  • Morrissey JH (1981) Silver-stain for proteins in polyacrylamide gels: A modified procedure with enhanced uniform sensitivity. Anal Biochem 117:307–310

    Google Scholar 

  • Mothes-Wagner U, Wagner G, Reitze HK, Seitz KA (1984) A standardized technique for the in toto epoxy resin embedding and precipitate-free staining of small specimens covered by strong protective outer surfaces. Microscopy 134:307–313

    Google Scholar 

  • Mukherjee AB, Ulane RE, Agrawal AK (1982) Role of uteroglobin and transglutaminase in masking the antigenicity of implanting rabbit embryos. Am J Reprod Immunol 2:135–141

    Google Scholar 

  • Nieder GL, Weitlauf HM, Suda-Hartman M (1987) Synthesis and secretion of stage-specific proteins by peri-implantation mouse embryos. Biol Reprod 36:687–699

    Google Scholar 

  • O'Farrell PH (1975) High resolution two-dimensional electrophoresis of proteins. J Biol Chem 250:4007–4021

    Google Scholar 

  • Reynolds ES (1963) The use of lead citrate at high pH as an electron opaque stain in electron microscopy. J Cell Biol 17:208–212

    Article  CAS  PubMed  Google Scholar 

  • Roberts RM, Imakawa K, Niwano Y, Kazemi M, Malathy PV, Hansen TR, Glass AA, Kronenberg LH (1989) Interferon production by the preimplantation sheep embryo. J Interferon Res 9:175–187

    Google Scholar 

  • Salamonsen LA, Manikhot J, Findlay JK (1988) Studies on the inhibitory action of interferons on prostaglandin release by ovine endometrial cells in vitro. J Reprod Fertil Abstract Series No. 2, 70:37

    Google Scholar 

  • Schultz GA, Tucker EB (1977) Protein synthesis and gene expression in preimplantation rabbit embryos. In: Johnson MH (ed) Development in Mammals, Vol 1. Elsevier North-Holland Biomedical Press, Amsterdam, pp 69–97

    Google Scholar 

  • Spurr AR (1969) A low-viscosity epoxy resin embedding medium for electron microscopy. J Ultrastruct Res 26:31–43

    CAS  PubMed  Google Scholar 

  • Tucker EB, Schultz GA (1978) Two-dimensional electrophoretic analysis of proteins synthesized during the early cleavage period in rabbit embryos. Exp Cell Res 114:438–443

    Google Scholar 

  • Van Blerkom J, Manes C (1974) Development of preimplantation rabbit embryos in vivo and in vitro. II. A comparison of qualitative aspects of protein synthesis. Dev Biol 40:40–51

    Google Scholar 

  • Van Blerkom J, Manes C, Daniel JC Jr (1973) Development of preimplantation rabbit embryos in vivo and in vitro. I. An ultrastructural comparison. Dev Biol 35:262–282

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Biology, Philipps-University, D-3550, Marburg, Federal Republic of Germany

    R. Henkel, D. R. Dannhorn, U. Petzoldt & C. Kirchner

Authors
  1. R. Henkel
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. D. R. Dannhorn
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. U. Petzoldt
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. C. Kirchner
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Henkel, R., Dannhorn, D.R., Petzoldt, U. et al. Ultrastructure, protein synthesis and secretion of day-6 rabbit blastocysts cultured in a chemically defined, protein-free medium. Anat Embryol 182, 465–472 (1990). https://doi.org/10.1007/BF00178911

Download citation

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00178911

Key words

Search

Navigation