title: Intrinsic and spontaneous neurogenesis in the postnatal slice culture of rat hippocampus.
abstract: Organotypic slice culture preserves the morphological and physiological features of the hippocampus of live animals for a certain time. The hippocampus is one of exceptional regions where neurons are generated intrinsically and spontaneously throughout postnatal life. We investigated the possibility that neurons are generated continuously at the dentate granule cell layer (GCL) in slice culture of the rat hippocampus. Using 5-bromodeoxyuridine (BrdU) labelling and retrovirus vector transduction methods, the phenotypes of the newly generated cells were identified immunohistochemically. At 4 weeks after BrdU exposure, BrdU-labelled cells were found in the GCL and were immunoreactive with a neuronal marker, anti-NeuN. There were fibrils immunoreactive with anti-glial fibrillary acidic protein (GFAP), an astrocyte marker, in the layer covering the GCL and occasionally encapsulated BrdU-labelled nuclei. When the newly divided cells were marked with the enhanced green fluorescent protein (EGFP) using a retrovirus vector, these cells had proliferative abilities throughout the following 4-week cultivation period. Four weeks after the inoculation, the EGFP-expressing cells consisted of various phenotypes of both early and late stages of differentiation; some were NeuN-positive cells with appearances of neurons in the GCL and some were immunoreactive with anti-Tuj1, a marker of immature neurons. Some EGFP-expressing cells were immunoreactive with anti-GFAP or anti-nestin, a marker of neural progenitors. The present study suggests that slice cultures intrinsically retain spontaneous neurogenic abilities for their cultivation period. The combination of slice culture and retrovirus transduction methods enable the newly divided cells to be followed up for a long period.
authors.affiliation: CREST, JST, Kawaguchi 332-0012, Japan.
authors.forename: Maki
authors.initials: M
authors.lastname: Kamada
authors.forename: Ren-Yong
authors.initials: RY
authors.lastname: Li
authors.forename: Mika
authors.initials: M
authors.lastname: Hashimoto
authors.forename: Masaaki
authors.initials: M
authors.lastname: Kakuda
authors.forename: Hiroshi
authors.initials: H
authors.lastname: Okada
authors.forename: Yoshio
authors.initials: Y
authors.lastname: Koyanagi
authors.forename: Toru
authors.initials: T
authors.lastname: Ishizuka
authors.forename: Hiromu
authors.initials: H
authors.lastname: Yawo
mesh_terms: D000818:Animals; D000831:Animals, Newborn; D001973:Bromodeoxyuridine / Q000378:metabolism; D064026:Calbindins; D002452:Cell Count / Q000379:methods; D002455:Cell Division / Q000502:physiology*; D049109:Cell Proliferation; D018891:Dentate Gyrus / Q000166:cytology* / Q000254:growth & development; D005822:Genetic Vectors / Q000502:physiology; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000378:metabolism; D021621:Imaging, Three-Dimensional / Q000379:methods; D007150:Immunohistochemistry / Q000379:methods; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D016571:Neural Networks, Computer; D009474:Neurons / Q000166:cytology* / Q000502:physiology* / Q000821:virology; D009924:Organ Culture Techniques; D010751:Phosphopyruvate Hydratase / Q000378:metabolism; D051381:Rats; D017208:Rats, Wistar; D012190:Retroviridae / Q000378:metabolism; D064030:S100 Calcium Binding Protein G / Q000378:metabolism; D013234:Stem Cells / Q000502:physiology / Q000821:virology; D013997:Time Factors; D014161:Transduction, Genetic / Q000379:methods; D014404:Tubulin / Q000378:metabolism; D015032:Zinc
chemical_list: D064026:Calbindins; D005904:Glial Fibrillary Acidic Protein; C489787:MAP2 protein, rat; D008869:Microtubule-Associated Proteins; D064030:S100 Calcium Binding Protein G; D014404:Tubulin; D049452:Green Fluorescent Proteins; D010751:Phosphopyruvate Hydratase; D001973:Bromodeoxyuridine; D015032:Zinc