title: Regulated appearance of NMDA receptor subunits and channel functions during in vitro neuronal differentiation.
abstract: The schedule of NMDA receptor subunit expression and the appearance of functional NMDA-gated ion channels were investigated during the retinoic acid (RA) induced neuronal differentiation of NE-4C, a p53-deficient mouse neuroectodermal progenitor cell line. NR2A, NR2B, and NR2D subunit transcripts were present in both nondifferentiated and neuronally differentiated cultures, while NR2C subunits were expressed only transiently, during the early period of neural differentiation. Several splice variants of NR1 were detected in noninduced progenitors and in RA-induced cells, except the N1 exon containing transcripts that appeared after the fourth day of induction, when neuronal processes were already formed. NR1 and NR2A subunit proteins were detected both in nondifferentiated progenitor cells and in neurons, while the mature form of NR2B subunit protein appeared only at the time of neuronal process elongation. Despite the early presence of NR1 and NR2A subunits, NMDA-evoked responses could be detected in NE-4C neurons only after the sixth day of induction, coinciding in time with the expression of the mature NR2B subunit. The formation of functional NMDA receptors also coincided with the appearance of synapsin I and synaptophysin. The lag period between the production of the subunits and the onset of channel function suggests that subunits capable of channel formation cannot form functional NMDA receptors until a certain stage of neuronal commitment. Thus, the in vitro neurogenesis by NE-4C cells provides a suitable tool to investigate some inherent regulatory processes involved in the initial maturation of NMDA receptor complexes.
authors.affiliation: Department of Neural Cell Biology, Institute of Experimental Medicine Hungarian Academy of Sciences, Budapest, Hungary.
authors.forename: Márta
authors.initials: M
authors.lastname: Jelitai
authors.forename: Katalin
authors.initials: K
authors.lastname: Schlett
authors.forename: Patrícia
authors.initials: P
authors.lastname: Varju
authors.forename: Ulrich
authors.initials: U
authors.lastname: Eisel
authors.forename: Emília
authors.initials: E
authors.lastname: Madarász
mesh_terms: D017398:Alternative Splicing / Q000502:physiology; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002454:Cell Differentiation / Q000502:physiology; D002478:Cells, Cultured; D018507:Gene Expression Regulation, Developmental; D066298:In Vitro Techniques; D051379:Mice; D009474:Neurons / Q000737:chemistry / Q000166:cytology* / Q000502:physiology; D012333:RNA, Messenger / Q000032:analysis; D016194:Receptors, N-Methyl-D-Aspartate / Q000032:analysis / Q000235:genetics* / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013234:Stem Cells / Q000737:chemistry / Q000166:cytology / Q000502:physiology
chemical_list: C120997:NR2A NMDA receptor; C121001:NR2B NMDA receptor; D012333:RNA, Messenger; D016194:Receptors, N-Methyl-D-Aspartate; D002118:Calcium