Skip to main content
SpringerLink
Log in

Ultrastructural and cytochemical identification of apoptotic cell death accompanying development of the fetal rat olfactory nerve layer

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

Abstract

It has been previously shown that the embryonic olfactory nerve contains, in addition to glial ensheathing cells, a large population of differentiated neurons that migrate from the developing olfactory epithelium, in close association with the olfactory axon fascicles. The purpose of our study was to verify the hypothesis according to which a process of physiological cell death might be involved in the progressive disappearance of these migrating neurons that has been reported during late embryonic stages in several immunocytochemical studies. To do so, we have investigated the development of the olfactory nerve layer in rat embryos by using light and electron microscopy, with special reference to the presence of cell death processes within this structure. We have also applied the histochemical TUNEL method allowing in situ visualization of cells degenerating by apoptosis. In order to determine if neurons were present among dying cells, a procedure of double-labeling was performed by combining the DNA-specific bisbenzimide with two neuronal markers, the protein B-50/GAP-43 and the lectin Ulex europaeus I. Results brought out the precise temporal and spatial patterns of programmed cell death accompanying the morphogenesis of the olfactory nerve layer. A cell death process was observed within the olfactory nerve layer from its onset at embryonic day 13 (E13). While only few pycnotic cells were observed in E13 and E14 embryos, their number increased from E15 to reach a maximum at E16 and then diminished. Few dying cells were also observed along the olfactory axon fascicles when they penetrated the olfactory nerve layer. Degenerating cells appeared strongly TUNEL-labeled and exhibited morphological features of cell death by apoptosis. Double-labeling experiments revealed that some of the apoptotic cells were neurons. These observations indicate that apoptosis may account for the progressive decrease in the number of migrating neurons present within the embryonic olfactory nerve layer. Otherwise, a zone of massive cell death by apoptosis was observed at E14 within the nasal mesenchyme located ventrally and caudally to the olfactory nerve layer. Double-labeling experiments showed that apoptotic cells present within this zone were not neurons. Our findings strongly suggest that apoptotic cell death of migrating neurons may allow the elimination of non-functional cells whereas that of mesenchymal cells may facilitate outgrowth of the newly formed olfactory axon fascicles by pathway formation.

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

  • Baker H, Farbman AI (1993) Olfactory afferent regulation of the dopamine phenotype in the fetal rat olfactory system. Neuroscience 52:115–134

    Google Scholar 

  • Carney PR, Silver J (1983) Studies on cell migration and axon guidance in the developing distal auditory system of the mouse. J Comp Neurol 215:359–369

    Google Scholar 

  • Chuah MI, Au C (1991) Olfactory Schwann cells are derived from precursor cells in the olfactory epithelium. J Neurosci Res 29:172–180

    Google Scholar 

  • Clarke PGH (1990) Developmental cell death: morphological diversity and multiple mechanisms. Anat Embryol 181:195–213

    CAS  PubMed  Google Scholar 

  • Cuadros MA, Rios A (1988) Spatial and temporal correlation between early nerve fiber growth and neuroepithelial cell death in the chick embryo retina. Anat Embryol 178:543–551

    Google Scholar 

  • Cunningham TJ (1982) Naturally occurring neuron death and its regulation by developing neural pathways. Int Rev Cytol 74:163–186

    Google Scholar 

  • Cuschieri A, Bannister LH (1975) The development of the olfactory mucosa in the mouse: light microscopy. J Anat 119:277–286

    Google Scholar 

  • Doucette JR (1984) The glial cells in the nerve fiber layer of the rat olfactory bulb. Anat Rec 210:385–391

    Google Scholar 

  • Doucette R (1989) Development of the nerve fiber layer in the olfactory bulb of mouse embryos. J Comp Neurol 285:514–527

    Google Scholar 

  • Doucette R (1990) Glial influences on axonal growth in the primary olfactory system. Glia 3:433–449

    Google Scholar 

  • Doucette R (1993) Glial cells in the nerve fiber layer of the main olfactory bulb of embryonic and adult mammals. Microsc Res Tech 24:113–130

    Google Scholar 

  • Doucette JR, Kiernan JA, Flumerfelt BA (1983) Two different patterns of retrograde degeneration in the olfactory epithelium following transection of primary olfactory axons. J Anat 136:673–689

    Google Scholar 

  • Ellis RE, Yuan J, Horvitz HR (1991) Mechanisms and function of cell death. Annu Rev Cell Biol 7:663–698

    Google Scholar 

  • Farbman AI (1986) Prenatal development of mammalian olfactory receptor cells. Chem Senses 11:3–18

    Google Scholar 

  • Farbman AI, Menco BPhM (1986) Development of olfactory epithelium in the rat. In: Breipohl W (ed) Ontogeny of olfaction. Springer, Berlin Heidelberg New York, pp 45–54

    Google Scholar 

  • Farbman AI, Squinto LM (1985) Early development of olfactory receptor cell axons. Brain Res Dev Brain Res 19:205–213

    Google Scholar 

  • Gavrieli Y, Sherman Y, Ben-Sasson SA (1992) Identification of programmed cell death in situ via specific labeling of nuclear DNA fragmentation. J Cell Biol 119:493–501

    Article  CAS  PubMed  Google Scholar 

  • Gesteland RC, Yancey RA, Farbman AI (1982) Development of olfactory receptor neuron selectivity in the rat fetus. Neuroscience 7:3127–3136

    Google Scholar 

  • Gispen WH, Nielander HB, DeGraan PNE, Oestreicher AB, Schrama LH, Schotman P (1991) Role of the growth-associated protein B-50/GAP-43 in neuronal plasticity. Mol Neurobiol 5:61–85

    Google Scholar 

  • Glücksmann A (1951) Cell deaths in normal vertebrate ontogeny. Biol Rev Comb Philos Soc 26:59–86

    Google Scholar 

  • Graziadei PPC, Monti Graziadei GA (1980) Neurogenesis and neuron regeneration in the olfactory system of mammals. III. Deafferentation and reinnervation of the olfactory bulb following section in the fila olfactoria in rat. J Neurocytol 9:145–162

    Google Scholar 

  • Holcomb JD, Guevara J, Calof AL (1994) Dynamics of apoptosis in murine olfactory epithelium following removal of target-derived trophic support: studies in vitro and in vivo. Soc Neurosci Abst 20:188.10

    Google Scholar 

  • Hopkinson-Woolley J, Hughes D, Gordon S, Martin P (1994) Macrophage recruitment during limb development and wound healing in the embryonic and foetal mouse. J Cell Sci 107:1159–1167

    Google Scholar 

  • Horsburgh GM, Sefton AJ (1986) The early development of the optic nerve and chiasm in embryonic rat. J Comp Neurol 243:547–560

    Google Scholar 

  • Hurle JM (1988) Cell death in developing systems. Methods Achiev Exp Pathol 13:55–86

    Google Scholar 

  • Jessen KR, Brennan A, Morgan L, Mirsky R, Kent A, Hashimoto Y, Gavrilovic J (1994) The Schwann cell precursor and is fate: a study of cell death and differentiation during gliogenesis in rat embryonic nerves. Neuron 12:509–527

    Google Scholar 

  • Källén B (1965) Degeneration and regeneration in the vertebrate central nervous system during embryogenesis. Prog Brain Res 14:153–161

    Google Scholar 

  • Kerr JFR, Wyllie AH, Currie AR (1972) Apoptosis: a basic biological phenomenon with wide-ranging implications in tissue kinetics. Br J Cancer 26:239–257

    CAS  PubMed  Google Scholar 

  • Lejour-Jeanty M (1965) Etude morphologique et cytochimique du dévelopment du palais primaire chez le rat. Arch Biol (Liège) 76:97–168

    Google Scholar 

  • Livne I, Gibson MJ, Silverman AJ (1993) Biochemical differentiation and intercellular interactions of migratory gonadotropinreleasing hormone (GnRH) cells in the mouse. Dev Biol 159:643–656

    Google Scholar 

  • Marin-Padilla M, Amieva MR (1989) Early neurogenesis of the mouse olfactory nerve: Golgi and electron microscopic studies. J Comp Neurol 288:339–352

    Google Scholar 

  • Michel D, Moyse E, Brun G, Jourdan F (1994) Induction of apoptosis in rat olfactory neuroepithelium by synaptic target ablation. Neuroreport 5:1329–1332

    Google Scholar 

  • Norgren RB, Ratner N, Brackenbury R (1992) Development of olfactory nerve glia defined by a monoclonal antibody specific for Schwann cells. Dev Dyn 194:231–238

    Google Scholar 

  • Oestreicher AB, VanDogen CJ, Zwiers H, Gispen WH (1983) Affinity-purified anti-B-50 protein antibody: interference with the function of the phosphoprotein B-50 in synaptic plasma membranes. J Neurochem 41:331–340

    Google Scholar 

  • Oppenheim RW (1991) Cell death during development of the nervous system. Annu Rev Neurosci 14:453–501

    Article  CAS  PubMed  Google Scholar 

  • Pellier V, Astic L (1994a) Histochemical and immunocytochemical study of the migration of neurons from the rat olfactory placode. Cell Tissue Res 275:587–598

    Google Scholar 

  • Pellier V, Astic L (1994b) Cell death in the developing olfactory epithelium of rat embryos. Dev Brain Res 79:307–315

    Google Scholar 

  • Pellier V, Astic L (1994c) Detection of apoptosis by electron microscopy and in situ labelling in the rat olfactory pit. Neuroreport 5:1429–1432

    Google Scholar 

  • Pellier V, Astic L, Oestreicher AB, Saucier D (1994) B-50/GAP43 expression by the olfactory receptor cells and the neurons migrating from the olfactory placode in embryonic rats. Dev Brain Res 80:63–72

    Google Scholar 

  • Raff MC (1992) Social controls on cell survival and cell death. Nature 356:397–400

    Google Scholar 

  • Raff MC, Barres BA, Burne JF, Coles HS, Ishizaki Y, Jacobson MD (1993) Programmed cell death and the control of cell survival: lessons from the nervous system. Science 262:695–700

    Google Scholar 

  • Robecchi MG (1972) Ultrastructure of differentiating sensory neurons in the olfactory placode of the chick embryo. Minerva Otorinolaryngol 22:196–204

    Google Scholar 

  • Saunders JW (1966) Death in embryonic systems. Science 154:604–612

    PubMed  Google Scholar 

  • Schwanzel-Fukuda M, Pfaff DW (1989) Origin of luteinizing hormone-releasing hormone neurons. Nature 338:161–164

    Google Scholar 

  • Schwartzman RA, Cidlowsky JA (1993) Apoptosis: the biochemistry and molecular biology of programmed cell death. Endocr Rev 14:133–151

    Google Scholar 

  • Schweichel JU, Merker HJ (1973) The morphology of various types of cell death in prenatal tissues. Teratology 7:253–266

    Google Scholar 

  • Schwob JE, Mieleszko KE, Stasky AA (1992) Olfactory sensory neurons are trophically dependent on the olfactory bulb for their prolonged survival. J Neurosci 12:3896–3919

    Google Scholar 

  • Skene JHP (1989) Axonal growth-associated proteins. Annu Rev Neurosci 12:127–156

    Google Scholar 

  • Soriano E, DelRio JA, Auladell C (1993) Characterization of the phenotype and birthdates of pyknotic dead cells in the nervous system by a combination of DNA staining and immunohistochemistry for 5′-bromodeoxyuridine and neural antigens. J Histochem Cytochem 41:819–827

    Google Scholar 

  • Tosney KW, Landmesser LT (1985) Development of the major pathways for neurite outgrowth in the chick hindlimb. Dev Biol 109:193–214

    Google Scholar 

  • Tosney KW, Schroeter Pokrzywinski JA (1988) Cell death delineates axon pathways in the hindlimb and does so independently of neurite outgrowth. Dev Biol 130:558–572

    Google Scholar 

  • Valverde F, Lopez-Mascaraque L (1991) Neuroglial arrangements in the olfactory glomeruli of the hedgehog. J Comp Neurol 307:658–674

    Google Scholar 

  • Valverde F, Santacana M, Heredia M (1992) Formation of an olfactory glomerulus: morphological aspects of development and organization. Neuroscience 49:255–275

    Google Scholar 

  • Valverde F, Heredia M, Santacana M (1993) Characterization of neuronal cell varieties migrating from the olfactory epithelium during prenatal development in the rat Immunocytochemical study using antibodies against olfactory marker protein (OMP) and luteinizing hormone-releasing hormone (LH-RH). Dev Brain Res 71:209–220

    Google Scholar 

  • VanCampenhout E (1956) Le développement embryonnaire comparé des nerfs olfactif et auditif. Acta Otorhinolaryngol Belg 11:279–287

    Google Scholar 

  • Verhaagen J, Oestreicher AB, Grillo M, Khew-Goodall YS, Gispen WH, Margolis FL (1990) Neuroplasticity in the olfactory system: differential effects of central and peripheral lesions of the primary olfactory pathway on the expression of B-50/GAP43 and the olfactory marker protein. J Neurosci Res 26:31–44

    Google Scholar 

  • Wray S, Grant P, Gainer H (1989a) Evidence that cells expressing luteinizing hormone-releasing hormone mRNA in the mouse are derived from progenitor cells in the olfactory placode. Proc Natl Acad Sci USA 86:8132–8136

    Google Scholar 

  • Wray S, Nieburgs A, Elkabes S (1989b) Spatiotemporal cell expression of luteinizing hormone-releasing hormone in the prenatal mouse: evidence for an embryonic origin in the olfactory placode. Dev Brain Res 46:309–318

    Google Scholar 

  • Wyllie AH (1987) Cell death. Int Rev Cytol [Suppl] 17:755–785

    Google Scholar 

  • Zakeri Z, Quaglino D, Ahuja HS (1994) Apoptotic cell death in the mouse limb and its suppression in the hammertoe mutant. Dev Biol 165:294–297

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Laboratoire de Physiologie Neurosensorielle, Université Claude Bernard/Lyon I, F-69622, Villeurbanne Cedex, France

    Véronique Pellier & Liliane Astic

  2. Université de Moncton, Centre Universitaire de Shippagan, Shippagan, NB, Canada

    Diane Saucier

  3. Rudolf Magnus Institute for Neurosciences, Utrecht University, P.O. Box 80040, NL-3508, TA Utrecht, The Netherlands

    A. Beate Oestreicher

Authors
  1. Véronique Pellier
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Diane Saucier
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. A. Beate Oestreicher
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Liliane Astic
    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

Pellier, V., Saucier, D., Oestreicher, A.B. et al. Ultrastructural and cytochemical identification of apoptotic cell death accompanying development of the fetal rat olfactory nerve layer. Anat Embryol 194, 99–109 (1996). https://doi.org/10.1007/BF00196319

Download citation

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00196319

Key words

Search

Navigation