Distribution of hydrolytic enzymes in early rat and mouse embryos — A reappraisal

  • Davor Solter
  • Ivan Damjanov
  • Nikola Škreb


The time of appearance and the distribution of alkaline and acid phosphatase and nonspecific esterase was investigated in cleavage and early postimplantation stages of mouse and rat embryos.

Alkaline and acid phosphatase appeared for the first time in 8-cell embryos. Activity of both enzymes grew progressively stronger to blastocyst stage. Acid phosphatase activity was revealed in the form of fine and coarse granules distributed evenly in the cytoplasm. Alkaline phosphatase was predominantly localized in plasma membranes. There was no difference in intensity of reaction between trophoblastic cells and the inner cell mass.

After implantation acid phosphatase was localized in coarse granules in the apical portion of entodermal cells. With the appearance of mesoderm, the cells of embryonal entoderm became flattened and devoid of acid phosphatase activity which was restricted to cells of extraembryonic entoderm. The activity of nonspecific esterase was not detected in preimplantation stages. In postimplantation embryos it roughly corresponded to the activity of acid phosphatase. Alkaline phosphatase was localized in cell membranes of ectodermal cells. The mesodermal cells of mouse embryo displayed a somewhat weaker activity than ectodermal cells, while in the rat embryo the same layer remained completely nonreactive.

Our findings on the distribution of the enzymes mentioned did not reveal any kind of polarity or bilateral symmetry in preimplantation stages. In postimplantation stages acid phosphatase and nonspecific esterase are probably bound to lysosomes and play an important role in embryonic nutrition. The absence of alkaline phosphatase from entodermal cells is somewhat puzzling and suggests that the process of molecular transport in those cells is most probably restricted to endocytosis. Our results suggest that all blastomeres are identical with respect to enzyme distribution and that the first signs of differentiation of enzyme content appear with the formation of germ layers.

Key words

Cleavage stages Egg-cylinder Mouse Rat Hydrolytic enzymes distribution Differentiation 


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  1. Barka, T., Anderson, P. J.: Histochemistry, Theory, Practice, and Bibliography. New York, Evanston, and London: Harper and Row 1965.Google Scholar
  2. Barlowe, P., Owen, D. A. J., Graham, C.: DNA synthesis in the preimplantation mouse embryo. J. Embryol. Exp. Morph. 27, 431–455 (1972).Google Scholar
  3. Beck, F., Lloyd, J. B., Griffiths, A.: A histochemical and biochemical study of some aspects of placental function in the rat using maternal injection of horseradish peroxidase. J. Anat. (Lond.) 101, 461–478 (1967a).Google Scholar
  4. Beck, F., Lloyd, J. B., Griffiths, A.: Lysosomal enzyme inhibition by trypan blue: a theory of teratogenesis. Science 157, 1180–1182 (1967b).Google Scholar
  5. Borghese, E.: Recent histochemical results of studies on embryos of some birds and mammals. Int. Rev. Cytol. 6, 289–341 (1957).Google Scholar
  6. Brachet, J.: The biochemistry of development. London: Pergamon 1960.Google Scholar
  7. Davidson, E. H.: Gene activity in early development. New York and London: Academic Press 1968.Google Scholar
  8. Deuchar, E. M.: Biochemical pattern in early developmental stages of vertebrates. In: The biochemistry of animal development. vol. 1, p. 245–304 (R. Weber ed.) New York and London: Academic Press 1965.Google Scholar
  9. Enders, A. C., Schlafke, S. J.: The fine structure of the blastocyst: some comparative studies. In: Preimplantation stages of pregnancy. Ciba Fdn. Symp. p. 29–59 (G. E. W. Wolstenholme and M. O'Connor, eds.). London: Churchill 1965.Google Scholar
  10. Gardner, R. L.: Manipulation on the blastocyst. Advanc. Biosci. 6, 279–301 (1971).Google Scholar
  11. Goldfischer, S., Essner, E., Novikoff, A. B.: The localization of phosphatase activities at the level of ultrastructure. J. Histochem. Cytochem. 12, 72–95 (1964).Google Scholar
  12. Graham, C. F.: The design of the mouse blastocyst. In: Control mechanisms of growth and differentiation, p. 371–378 (D. Davies and M. Balls, eds.). Cambridge: University Press 1971.Google Scholar
  13. Holt, S. J.: Factors governing the validity of staining methods for enzymes, and their bearing upon the Gomori acid phosphatase technique. Exp. Cell Res. Suppl. 7, p. 1–27 (1959).Google Scholar
  14. Holt, S. J.: Some observations on the occurence and nature of esterases in lysosomes. In: Lysosomes. Ciba Fdn. Symp. p. 114–125 (A. V. S. de Reuck and M. P. Cameron eds.). London: Churchill 1965.Google Scholar
  15. Holt, S. J., Hobbiger, E. E., Pawan, G. L. S.: Preservation of integrity of rat tissues for cytochemical staining purposes. J. biophys. biochem. Cytol. 7, 383–386 (1960).Google Scholar
  16. Levak-Švajger, B., Švajger, A., Škreb, N.: Separation of germ layers in presomite rat embryos. Experientia (Basel) 25, 1311–1312 (1969).Google Scholar
  17. McLaren, A.: Recent studies on developmental regulation in vertebrates. In: Handbook of molecular cytology, p. 639–655 (A. Lima-de-Faria-, ed.). Amsterdam and London: North-Holland 1969.Google Scholar
  18. Merker, H.-J., Villegas, H.: Elektronenmikroskopische Untersuchungen zum Problem des Stoffaustausches zwischen Mutter und Keim bei Rattenembryonen des Tages 7–10. Z. Anat. Entwickl.-Gesch. 131, 325–346 (1970).Google Scholar
  19. Mintz, B.: Formation of genetically mosaic mouse embryos, and early development of “lethal (t12/t12)-normal” mosaics. J. exp. Zool. 157, 273–292 (1964).Google Scholar
  20. Moore, N. W., Adams, C. E., Rowson, L. E. A.: Developmental potential of single blastomeres of the rabbit egg. J. Reprod. Fertil. 17, 527–531 (1968).Google Scholar
  21. Mulnard, J. G.: Contribution à la connaissance des enzymes dans l'ontogénèse. Les phosphomonoésterases acide et alcaline dans le développement du rat et de la souris. Arch. Biol. (Liège) 66, 527–688 (1955).Google Scholar
  22. Mulnard, J. G.: Studies of regulation of mouse ova in vitro. In: Preimplantation stages of pregnancy. Ciba Fdn Symp. p. 123–144 (G. E. W. Wolstenholme and M. O'Conner, eds.). London: Churchill 1965.Google Scholar
  23. Pearse, A. G.: Histochemistry theoretical and applied, vol. 1. London: Churchill 1968.Google Scholar
  24. Reale, E.: Electron microscopic localization of alkaline phosphatase from material prepared with the cryostat microtome. Exp. Cell Res. 26, 210–211 (1962).Google Scholar
  25. Rodé, B., Damjanov, I., Škreb, N.: Distribution of acid and alkaline phosphatases activity in early stages of rat embryos. Bull. Sci. Yougosl. 13, 304 (1968).Google Scholar
  26. Rossi, F.: Histochemie der Enzyme bei der Entwicklung. In: Handbuch der Histochemie VII/4, S. 109–298 (W. Graumann and K. Neumann, Hrsg.). Stuttgart: G. Fischer 1964.Google Scholar
  27. Seidel, F.: Die Entwicklungsfähigkeiten isolierter Furchungszellen aus dem Ei des Kaninchens Oryctolagus cuniculus. Arch. Entwickl.-Mech. Org. 152, 43–130 (1960).Google Scholar
  28. Smith, M. S. R., Wilson, I. B.: Histochemical observations on early implantation in the mouse. J. Embryol. exp. Morph. 25, 165–174 (1971).Google Scholar
  29. Solter, D., Damjanov, I., Škreb, N.: Ultrastructure of mouse egg-cylinder. Z. Anat. Entwickl.-Gesch. 132, 291–298 (1970).Google Scholar
  30. Solter, D., Škreb, N., Damjanov, I.: Cell cycle analysis in the mouse egg-cylinder. Exp. Cell Res. 64, 331–334 (1971).Google Scholar
  31. Spors, S.: Elektronenmikroskopische Untersuchungen der lysosomalen sauren Phosphatase in den Keimblättern des Ratten-Embryo von Tag 8–10. Histochemie 25, 143–151 (1971).Google Scholar
  32. Tarkowski, A. K., Wroblewska, J.: Development of blastomeres of mouse eggs isolated at the 4- and 8-cell stage. J. Embryol. exp. Morph. 18, 155–180 (1967).Google Scholar
  33. Wilson, I. B., Bolton, E., Cuttler, R. H.: Preimplantation differentiation in the mouse egg as revealed by microinjection of vital markers. J. Embryol. exp. Morph. 27, 467–479 (1972).Google Scholar
  34. Zugibe, F. T., Kopaczyk, K. C., Cape, W. E., Last, J. H.: A new carbowax method for routinely performing lipid, hematoxylin and eosin and elastin staining techniques on adjacent freeze-dried or formalin-fixed section. J. Histochem. Cytochem. 6, 133–138 (1958).Google Scholar

Copyright information

© Springer-Verlag 1973

Authors and Affiliations

  • Davor Solter
    • 1
  • Ivan Damjanov
    • 1
  • Nikola Škreb
    • 1
  1. 1.Department of Biology and Department of Pathology, Medical FacultyUniversity of ZagrebZagrebYugoslavia

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