Formation of spicules by sclerocytes from the freshwater spongeEphydatia muelleri in short-term cultures in vitro

  • Georg Imsiecke
  • Renate Steffen
  • Marcio Custodio
  • Radovan Borojevic
  • Werner E. G. Müller
Cellular Models


Cells from the freshwater spongeEphydatia muelleri were isolated by dissociating hatching gemmules. During the first 24 h the cells reaggregated, but the aggregates progressively disintegrated again to single cells, among which the spicule-forming sclerocytes were recognized. Such cultures were used to study spicule (megascleres) formation in vitro. The isolated sclerocytes formed the organic central axial filament onto which they deposited inorganic silicon. The size of the spicules (200 to 350µm in length) as well as the rate of spicule formation (1 to 10µm/h) under in vitro conditions were similar to the values measured in vivo. Immediately after completion of spicule formation, or even before, the sclerocyte could start formation of a new spicule; 5% of the cells were in the process of forming two spicules simultaneously. Cultivation of sclerocytes in the absence of silicon resulted in the formation of the axial filament only. We succeeded in maintaining the sclerocytes in a proliferating and spicule-forming state for up to 3 mo. These results demonstrate that the establishment of short-term cell cultures fromE. muelleri is possible; however, future studies must be undertaken to identify the growth factors required for a permanent culture of sponge cells.

Key words

sponges Ephydatia muelleri spicules sclerocytes cell culture 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Bachmann, M.; Althoff, H.; Selenka, C., et al. Translocation of the nuclear autoantigen La to the cell surface of Herpes simplex virus type 1 infected cells. Autoimmunity 12:37–45; 1992.PubMedGoogle Scholar
  2. Belas, F. J.; Francis, J. C.; Poirrier, M. A. Significance of small organic chelators in laboratory cultures ofEphydatia fluviatilis (Porifera: Spongillidae) Trans. Am. Microsc. Soc. 111(3):169–179; 1992.CrossRefGoogle Scholar
  3. Bergquist, P. R. The sponges. London: Hutchinson Ltd.; 1978.Google Scholar
  4. Berthold, G. Untersuchungen über die Histoblastendifferenzierung in der Gemmula vonEphydatia fluviatilis. Z. Wiss. Mikr. Tech. 69:227–243; 1969.Google Scholar
  5. Coutinho, C.; Vissers, S.; Van de Vyver, H. Evidence of homeobox genes in the freshwater spongeEphydatia fluviatilis. In: Soest, R. W. M. v.; Kempen T. M. G. v.; Braekman, J. C., eds. Sponges in time and space. Rotterdam: Balkema Press; 1994:47–54.Google Scholar
  6. Evans, R. The structure and metamorphosis of larvae ofSpongilla lacustris. Q. J. Microsc. Sci. 43:363–477; 1899.Google Scholar
  7. Garrone, R. Collagène, spongine et squelette mineral chez l’épongeHaliclona rosea (O.S.). J. Micro. 8:581–598; 1969.Google Scholar
  8. Harrison, F. W.; Cowden, R. R. Dormancy release and development from gemmules of the fresh-water sponge,Spongilla lacustris: a supravital study with acridine orange. Trans. Am. Microsc. Soc. 102:309–318; 1983.CrossRefGoogle Scholar
  9. Holvoet, S.; van de Vyver, G. Skeletogenesis inEphydatia fluviatilis grown in the presence of puromycin and hydroxyurea. In: Rützler, K., ed. New perspectives in sponge biology. Washington, DC: Smithsonian Institution Press; 1985a:200–205.Google Scholar
  10. Holvoet, S.; van de Vyver, G. Effects of 2,2′-bipyridine on skeletogenesis ofEphydatia fluviatilis. In: Rützler, K., ed. New perspectives in sponge biology. Washington, DC: Smithsonian Institution Press; 1985b:206–210.Google Scholar
  11. Imsiecke, G. Ingestion, digestion, and egestion inSpongilla lacustris (Porifera, Spongillidae) after pulse feeding withChlamydomonas reinhardtii (Volvocales). Zoomorphology (Berl) 113:233–244; 1993.CrossRefGoogle Scholar
  12. Imsiecke, G.; Borojevic, R.; Müller, W. E. G. Retinoic acid acts as a morphogen in freshwater sponges. Invertebr. Reproduc. & Dev. 26:89–98; 1994.Google Scholar
  13. Minchin, E. A. Sponge spicules. A summary of the present knowledge. Ergeb. Fortschr. Zool. 2:171–274; 1909.Google Scholar
  14. Müller, W. E. G.; Zahn, R. K.; Maidhof, A.Spongilla gutenbergiana n.sp., ein Süßwasserschwamm aus dem Mittel-Eozän von Messel. Senckenb. Lethaea 63:465–472; 1982.Google Scholar
  15. Müller, W. E. G.; Rottmann, M.; Diehl-Seifert, B., et al. Role of the aggregation factor in the regulation of phosphoinositide metabolism in sponges. Possible consequences on calcium efflux and on mitogenesis. J. Biol. Chem. 262:9850–9858; 1987.PubMedGoogle Scholar
  16. Müller, W. E. G.; Müller, I. M.; Gamulin, V. On the monophyletic evolution of the metazoa. Brasil. J. Med. Biol. Res. 27:2083–2096; 1994.Google Scholar
  17. Orlov, Y. A. Fundamentals of palaeontology, vol II. Jerusalem: Israel program for scientific translations; 1971:11.Google Scholar
  18. Ott, E.; Volkheimer, W. Palaeospongilla chubutensis n.g. et n.sp., ein Süßwasserschwamm aus der Kreide Patagoniens. N. Jb. Geol. Paläont. Abh. 140:49–63; 1972.Google Scholar
  19. Pé; J. Étude quantitative de la régulation du squelette chez une éponge d’eau douce. Arch. Biol. (Bruxelles) 84:147–173; 1973.Google Scholar
  20. Penney, J. T.; Racek, A. A. Comprehensive revision of a worldwide collection of freshwater sponges (Porifera: Spongillidae). US Nat. Mus. Bull. 272:1–184; 1968.Google Scholar
  21. Poirrier, M. A.; Trabanino, S. Freshwater sponges (Porifera, Spongillidae) from lake Ilopango, El Salvador, with observations on spicule malformation in Spongilla alba. Trans. Am. Microsc. Soc. 108:211–214; 1989.CrossRefGoogle Scholar
  22. Rasmont, R. Une technique de culture des éponges d’eau douce en milieu controlé. Ann. Soc. R. Zool. Belg. 91:147–155; 1961.Google Scholar
  23. Russell, W. C.; Newman, C.; Williamson, D. H. A simple cytochemical technique for demonstration of DNA in cells infected with mycoplasma viruses. Nature 153:461–462; 1975.CrossRefGoogle Scholar
  24. Rozenfeld, F. Effects of puromycin on the differentiation of the freshwater sponge:Ephydatia fluviatilis. Differentiation 17:193–198; 1980.PubMedCrossRefGoogle Scholar
  25. Schäcke, H.; Schröder, H. C.; Gamulin, V., et al. Molecular cloning of a tyrosine kinase gene from the marine spongeGeodia cydonium: a new member belonging to the receptor tyrosine kinase class II family. Mol. Memb. Biol. 11:101–107; 1994.CrossRefGoogle Scholar
  26. Schmidt, I. Étude préliminaire de la différenciation des thésocytes d’Ephydatia fluviatilis L. Extraits méchaniquement de la gemmules. C. R. Acad. Sci. Paris Sér. D. 271:924–927; 1970.Google Scholar
  27. Schulze, P. Beiträge zur Kenntnis der Kieselnadelbildung besonders bei den Spongilliden. Arch. Zellforsch. 17:105–130; 1923.Google Scholar
  28. Shore, R. E. Axial filament of siliceous sponge spicules, its organic components and synthesis. Biol. Bull. 143:689–698; 1972.CrossRefGoogle Scholar
  29. Simon, L. Über die Spezifität der Nadeln und die Variabilität der Arten bei den Spongilliden. Zool. Jahrb. 64:97–266; 1953.Google Scholar
  30. Simpson, T. L. The biology of sponges. New York: Springer-Verlag; 1984.Google Scholar
  31. Simpson, T. L. Effects of germanium on silica deposition in sponges. In: Simpson, T. L.; Volcani, B. E., eds. Silicon and siliceous structures in biological systems. New York: Springer-Verlag; 1981:527–550.Google Scholar
  32. Simpson, T. L.; Vaccaro, C. A. An ultrastructural study of silica deposition in the freshwater spongeSpongilla lacustris. J. Ultrastruct. Res. 47:296–309; 1974.PubMedCrossRefGoogle Scholar
  33. Weissenfels, N. Biologie und Mikroskopische Anatomie der Süßwasserschwämme (Spongillidae). Stuttgart, New York: Fischer; 1989.Google Scholar
  34. Weissenfels, N.; Landschoff, H. W. Bau und Funktion des SüsswasserschwammsEphydatia fluviatilis L. (Porifera). IV. Die Entwicklung der monaxialen SiO2-Nadeln in Sandwich-Kulturen. Zool. Jb. Anat. 98:355–371; 1977.Google Scholar
  35. Yourassowsky, C.; Rasmont, R. The differentiation of sclerocytes in freshwater sponges grown in a silica-poor medium. Differentiation 25:5–9; 1983.CrossRefGoogle Scholar
  36. Zeuthen, E. On the hibernation ofSpongilla lacustris (L.). Z. Vgl. Physiol. 26:537–547; 1939.Google Scholar

Copyright information

© Society for In Vitro Biology 1995

Authors and Affiliations

  • Georg Imsiecke
    • 1
  • Renate Steffen
    • 1
  • Marcio Custodio
    • 2
  • Radovan Borojevic
    • 2
  • Werner E. G. Müller
    • 1
  1. 1.Institut für Physiologische Chemie, UniversitätMainzGermany
  2. 2.Departamento de Bioquímica, Instituto de QuímicaUniversidade Federal do Rio de JaneiroRio de JaneiroBrazil

Personalised recommendations