Advertisement

Histochemistry

, Volume 93, Issue 6, pp 607–615 | Cite as

Secretory glycoproteins of the rat subcommissural organ are N-linked complex-type glycoproteins. Demonstration by combined use of lectins and specific glycosidases, and by the administration of tunicamycin

  • H. Herrera
  • E. M. Rodríguez
Article

Summary

Two experimental protocols were used to investigate the secretory glycoproteins of the subcommissural organ (SCO).

Protocol I: Lectins, specific exoglycosidases and immunocytochemistry were sequentially applied to the same section or to adjacent semithin sections of the rat SCO fixed in Bouin's fluid and embedded in methacrylate. Lectins used: concanavalin A (con A), wheat germ agglutinin, Limulus polyphemus agglutinin, Ricinus communis agglutinin and Arachis hypogeae agglutinin. Glycosidases used: neuroaminidase, β-galactosidase, α-mannosidase, α-glucosidase and β-N-acetyl-glucosaminidase. For immunocytochemistry an antiserum against bovine Reissner's fiber (AFRU) was used. Lectins and glycosidases were used in sequences that allowed the cleaved sugar residue to be identified as well as that appearing exposed as a terminal residue. This approach led to the following conclusions: (1) the terminal sugar chain of the secreted glycoproteins has the sequence sialic acid-galactose-glucosamine-; (2) the con A-binding material present in the rough endoplasmic reticulum corresponds to mannose; (3) the apical secretory granules and Reissner's fibers displayed a strong con A affinity after removing sialic acid, thus indicating the presence of internal mannosyl residues in the secreted material; (4) after removing most of the sugar moieties the secretory material continued to be strongly immunoreactive with AFRU. Protocol II: Rats were injected into the lateral ventricle with Tunicamycin and killed 12, 24, 50 and 60 h after the injection. The SCO of rats from the last two groups showed a complete absence of con A binding sites. The results from the two experiments confirm that the secretory glycoproteins of the rat SCO are N-linked complex-type glycoproteins with the conformation previously suggested (Rodríguez et al. 1986).

Keywords

Sialic Acid Wheat Germ Agglutinin Tunicamycin Sugar Chain Secretory Glycoprotein 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. Alroy J, Orgad U, Ucci AA, Pereira MEA (1984) Identification of glycoprotein storage disease by lectins: a new diagnostic method. J Histochem Cytochem 32:1280–1284Google Scholar
  2. Bargmann W, Schiebler TH (1952) Histologische und cytochemische Untersuchungen am Subcommissuralorgan von Säugern. Z Zellforsch 37:583–596Google Scholar
  3. Diederen JHB (1970) The subcommissural organ of Rana temporaria (L.). A cytologic, cytochemical, cytoenzymological and electron microscopical study. Z Zellforsch 111:379–403Google Scholar
  4. Gibbons RA (1963) The sensitivity of neuroaminosidic linkage in mucosubstances towards acid and towards neuroaminidase. Biochem J 89:380Google Scholar
  5. Hickmann S, Theodorakis JL, Greco JM, Brown PH (1984) Processing of MOPC 315 immunoglobulin A oligosaccharides: evidence for endoplasmic reticulum and trans Golgi α 1–2-mannosidase activity. J Cell Biol 98:407–416Google Scholar
  6. Hubbard SC, Ivatt RJ (1981) Synthesis and processing of asparagine-linked oligosaccharides. Ann Rev Biochem 50:555–583Google Scholar
  7. Hunt LA (1979) Biosynthesis and maturation of cellular membrane glycoproteins. J Supramol Struct 12:209–226Google Scholar
  8. Mahoney WC, Duksin D (1979) Biological activities of the two major components of Tunicamycin. J Biol Chem 254:6572–6575Google Scholar
  9. Meiniel R, Meiniel A (1985) Analysis of the secretions of the subcommissural organ of several vertebrates species by use of fluorescent lectins. Cell Tissue Res 239:359–364Google Scholar
  10. Meiniel A, Molat JL, Meiniel R (1988) Complex-type glycoproteins synthesized in the subcommissural organ of mammals. Light-and electron-microscopic investigations by use of lectins. Cell Tissue Res 253:383–395Google Scholar
  11. Muresan V, Iwanij V, Smith ZDJ, Jamieson JD (1982) Purification and use of limulin: a sialic acid specific lectin. J Histochem Cytochem 30:938Google Scholar
  12. Naumann W (1968) Histochemische Untersuchungen am Subcommissuralorgan und am Reissnerschen Faden von Lampetra planeri (Bloch). Z Zellforsch 87:571–591Google Scholar
  13. Oksche A (1962) Histologische, histochemische und experimentelle Studien am Subkommissuralorgan von Anuren (mit Hinweisen auf den Epiphysenkomplex). Z Zellforsch 57:240–326Google Scholar
  14. Olsson R (1958) Studies on the subcommissural organ. Acta Zool 39:71–102Google Scholar
  15. Peruzzo B, Rodríguez EM (1989) Light and electron microscopical demonstration of concanavalin A and wheat-germ agglutinin binding sites by use of antibodies against the lectin or its label (peroxidase). Histochemistry 92:505–513Google Scholar
  16. Podell SB, Vacquer VD (1984) Wheat germ agglutinin blocks the acrosome reaction in Strongylocentrotus purpuratus sperm by binding a 210,000-mol-wt membrane protein. J Cell Biol 99:1598–1604Google Scholar
  17. Rodríguez EM, Oksche A, Hein S, Rodríguez S, Yulis CR (1984a) Comparative immunocytochemical study of the subcommissural organ. Cell Tissue Res 237:427–441Google Scholar
  18. Rodríguez EM, Yulis C, Peruzzo B, Alvial G, Andrade R (1984b) Standardization of various applications of methacrylate embedding and silver methenamine for light and electron microscopy immunocytochemistry. Histochemistry 81:253–263Google Scholar
  19. Rodríguez EM, Herrera H, Peruzzo B, Rodríguez S, Hein S, Oksche A (1986) Light and electron-microscopic immunocytochemistry and lectin histochemistry of the subcommissural organ. Evidence for processing of the secretory material. Cell Tissue Res 243:545–559Google Scholar
  20. Rodríguez EM, Hein S, Rodríguez S, Herrera H, Peruzzo B, Nualart F, Oksche A (1987) Analysis of the secretory products of the subcomissural organ. In: Scharrer B, Korf H-W, Hartwig H-G (eds) Functional morphology of neuroendocrine systems. Springer, Heidelberg Berlin New York, pp 189–202Google Scholar
  21. Rodríguez EM, Peruzzo B, Alfaro L, Herrera H (1988) Combined used of lectin histochemistry and immunocytochemistry for the study of neurosecretion. In: Pickering BT, Wakerley JB, Summerlee AJS (eds) Neurosecretion. Cellular aspects of the production and release of neuropeptides. Plenum Press, New York London, pp 71–80Google Scholar
  22. Rodríguez S, Rodríguez EM, Jara P, Peruzzo B, Oksche A (1990) Single injection into the cerebrospinal fluid of antibodies against the secretory material of the subcommissural organ reversibly blocks formation of Reissner's fiber: immunocytochemical investigation in the rat. Exp Brain Res (in press)Google Scholar
  23. Roth J (1978) The lectins: molecular probes in cell biology and membrane research. Exp Pathol 3[Suppl]:1–180Google Scholar
  24. Spicer SS, Warren L (1960) The histochemistry of sialic acid containing mucoproteins. J Histochem Cytochem 8:135Google Scholar
  25. Staneloni RJ, Leloir LF (1982) The biosynthetic pathway of the asparagine-linked oligosaccharides of glycoproteins. CRC Crit Rev Biochem 12:289–326Google Scholar
  26. Sterba G, Wolf G (1969) Vorkommen und Funktion der Sialinsäure im Reissnerschen Faden. Histochemie 17:57–63Google Scholar
  27. Sternberger LA, Hardy PH, Jr Cuculis JJ, Meyer HG (1970) The unlabeled antibody enzyme method of immunohistochemistry; preparation and properties of soluble antigen-antibody complex (horseradish peroxidase-antiperoxidase) and its use in identification of spirochetes. J Histochem Cytochem 18:315–333Google Scholar
  28. Stutinsky F (1950) Colloïde, corps de Herring et substance Gomori-positive de la neurohypophyse. CR Soc Bid (Paris) 144:1357–1360Google Scholar
  29. Wislocki GB, Leduc E (1952) The cytology and histochemistry of the subcommissural organ and Reissner's fiber in rodents. J Comp Neurol 97:515–544Google Scholar

Copyright information

© Springer-Verlag 1990

Authors and Affiliations

  • H. Herrera
    • 1
  • E. M. Rodríguez
    • 1
  1. 1.Instituto de Histología y Patología, Facultad de MedicinaUniversidad Austral de ChileValdiviaChile

Personalised recommendations