title: Regulation of neuronal differentiation by N-methyl-D-aspartate receptors expressed in neural progenitor cells isolated from adult mouse hippocampus.
abstract: In vitro culture of neural progenitor cells isolated from adult murine hippocampus according to the Percoll density gradient method resulted in formation of round spheres not immunoreactive to microtubule-associated protein-2 (MAP-2) or glial fibrillary acidic protein in the presence of basic fibroblast growth factor within 12 days in vitro (DIV). Reverse-transcription PCR analysis revealed constitutive expression in these neurospheres of different subunits required for assembly of functional heteromeric N-methyl-D-aspartate (NMDA) receptor channels. Immunocytochemical analysis confirmed expression of NR1, NR2A, and NR2B subunits in neurospheres cultured for 4-12 DIV. Brief (5 min) exposure to NMDA induced marked expression of c-Fos, Fos-B, Fra-2, and c-Jun proteins in neurospheres cultured for 12 DIV 2 hr later. The NMDA receptor antagonist dizocilpine markedly inhibited expression of both c-Jun and c-Fos proteins in NMDA-exposed neurospheres. Sustained exposure to NMDA not only markedly inhibited neurosphere formation by 12 DIV when exposed from 4-12 DIV, but also resulted in facilitation of subsequent differentiation of neurospheres exposed to all-trans retinoic acid to cells immunoreactive to both neuron-specific enolase and neuronal nuclei, in addition to MAP-2, as revealed by Western blot and immunocytochemistry analyses. These results suggest that functional heteromeric NMDA receptors may be expressed constitutively in neural progenitor cells before differentiation to play a crucial role in commitment and differentiation to neurons in adult murine hippocampus.
authors.affiliation: Laboratory of Molecular Pharmacology, Kanazawa University Graduate School of Natural Science and Technology, Kanazawa, Japan.
authors.forename: Tomoya
authors.initials: T
authors.lastname: Kitayama
authors.forename: Masanori
authors.initials: M
authors.lastname: Yoneyama
authors.forename: Keisuke
authors.initials: K
authors.lastname: Tamaki
authors.forename: Yukio
authors.initials: Y
authors.lastname: Yoneda
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D015153:Blotting, Western / Q000379:methods; D002452:Cell Count / Q000379:methods; D002454:Cell Differentiation / Q000502:physiology*; D020934:Ciliary Neurotrophic Factor / Q000494:pharmacology; D016291:Dizocilpine Maleate / Q000494:pharmacology; D004334:Drug Administration Schedule; D004347:Drug Interactions; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D015870:Gene Expression; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D006624:Hippocampus / Q000166:cytology*; D007150:Immunohistochemistry / Q000379:methods; D007381:Intermediate Filament Proteins / Q000378:metabolism; D008297:Male; D051379:Mice; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D016202:N-Methylaspartate / Q000494:pharmacology; D009419:Nerve Tissue Proteins / Q000378:metabolism; D064231:Nestin; D009474:Neurons / Q000166:cytology* / Q000378:metabolism; D016759:Oncogene Protein p65(gag-jun) / Q000378:metabolism; D016761:Oncogene Proteins v-fos / Q000378:metabolism; D010751:Phosphopyruvate Hydratase / Q000378:metabolism; D018809:Proliferating Cell Nuclear Antigen / Q000378:metabolism; D012333:RNA, Messenger / Q000096:biosynthesis; D016194:Receptors, N-Methyl-D-Aspartate / Q000502:physiology*; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000379:methods; D013234:Stem Cells / Q000166:cytology / Q000378:metabolism*; D013997:Time Factors; D014212:Tretinoin / Q000494:pharmacology
chemical_list: D020934:Ciliary Neurotrophic Factor; D018690:Excitatory Amino Acid Agonists; D018691:Excitatory Amino Acid Antagonists; D005904:Glial Fibrillary Acidic Protein; D007381:Intermediate Filament Proteins; D008869:Microtubule-Associated Proteins; D009419:Nerve Tissue Proteins; C577485:Nes protein, mouse; D064231:Nestin; D016759:Oncogene Protein p65(gag-jun); D016761:Oncogene Proteins v-fos; D018809:Proliferating Cell Nuclear Antigen; D012333:RNA, Messenger; D016194:Receptors, N-Methyl-D-Aspartate; D014212:Tretinoin; D016202:N-Methylaspartate; D016291:Dizocilpine Maleate; D010751:Phosphopyruvate Hydratase