title: Primary culture of cortical neurons, type-1 astrocytes, and microglial cells from cynomolgus monkey (Macaca fascicularis) fetuses.
abstract: We established selective primary cultures of neurons, astrocytes, and microglial cells from cryopreserved fetal cerebral cortex of cynomolgus monkeys (Macaca fascicularis). At 14 days in serum-containing medium, the cell cultures of the fetal cerebral cortex consisted primarily of neurons, astrocytes, and floating microglial cells. At 21 days, we observed a small number of myelin basic protein (MBP)-positive oligodendrocytes. The addition of cytosine arabinoside (a selective DNA synthesis inhibitor) at 2 days in culture eliminated proliferative glial cells, allowing adequate numbers of neurons to survive selectively. A chemically defined serum-free medium successfully supported neuronal survival at a level equivalent to that supported by the serum-containing medium. Brain-derived neurotrophic factor (BDNF) significantly affected the survival of primate neurons. Glutamate induced a significant degree of neuronal cell death against primate neurons and MK-801, a selective N-methyl-D-aspartate receptor (NMDAR) antagonist, blocked cell death, which suggests that primate cortical neurons have NMDAR and the glutamate-induced cell toxicity is mediated by NMDAR. In the serum-free medium, type-1 astrocytes responded to dibutyryl cyclic AMP and showed a process-bearing morphology. The growth of type-1 astrocytes in the serum-free medium was stimulated by epidermal growth factor (EGF), basic fibroblast growth factor (bFGF), and hydrocortisone, which are known growth factors in rat type-1 astrocytes. Cultured microglial cells expressed CD68, a monocyte marker. Macrophage-colony stimulating factor (M-CSF) stimulated microglial cell growth in the serum-free medium. These selective primary culture systems of primate cerebral cortical cells will be useful in issues involving species specificity in neuroscience.
authors.affiliation: Department of Biomedical Science, Graduate School of Agricultural and Life Sciences, The University of Tokyo, 1-1-1 Yayoi, Bunkyo-ku, Tokyo 113-8657, Japan. taka-u@yayoi.club.ne.jp
authors.forename: Takayuki
authors.initials: T
authors.lastname: Negishi
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Ishii
authors.forename: Shigeru
authors.initials: S
authors.lastname: Kyuwa
authors.forename: Yoichiro
authors.initials: Y
authors.lastname: Kuroda
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Yoshikawa
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D015214:Antigens, Differentiation, Myelomonocytic / Q000378:metabolism; D001253:Astrocytes / Q000187:drug effects / Q000502:physiology*; D018929:Cell Culture Techniques / Q000379:methods*; D016923:Cell Death / Q000502:physiology; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D016895:Culture Media, Serum-Free / Q000494:pharmacology; D003563:Cyclic CMP / Q000031:analogs & derivatives* / Q000494:pharmacology; D003561:Cytarabine / Q000494:pharmacology; D016291:Dizocilpine Maleate / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004815:Epidermal Growth Factor / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D005333:Fetus / Q000166:cytology*; D005346:Fibroblast Growth Factors / Q000494:pharmacology; D005904:Glial Fibrillary Acidic Protein / Q000378:metabolism; D018698:Glutamic Acid / Q000494:pharmacology; D007150:Immunohistochemistry; D007166:Immunosuppressive Agents / Q000494:pharmacology; D008252:Macaca fascicularis; D017628:Microglia / Q000187:drug effects / Q000502:physiology*; D008869:Microtubule-Associated Proteins / Q000378:metabolism; D004676:Myelin Basic Protein / Q000378:metabolism; D009474:Neurons / Q000187:drug effects / Q000502:physiology*; D000097382:CD68 Molecule
chemical_list: D015703:Antigens, CD; D015214:Antigens, Differentiation, Myelomonocytic; D016895:Culture Media, Serum-Free; D003563:Cyclic CMP; D003561:Cytarabine; D016291:Dizocilpine Maleate; D004815:Epidermal Growth Factor; D018691:Excitatory Amino Acid Antagonists; D005346:Fibroblast Growth Factors; D005904:Glial Fibrillary Acidic Protein; D018698:Glutamic Acid; D007166:Immunosuppressive Agents; D008869:Microtubule-Associated Proteins; D004676:Myelin Basic Protein; C067980:CD68 antigen, human; D000097382:CD68 Molecule; C053441:dibutyryl cyclic-3',5'-cytidine monophosphate