title: Generation of embryonic stem cells and transgenic mice expressing green fluorescence protein in midbrain dopaminergic neurons.
abstract: We have generated embryonic stem (ES) cells and transgenic mice with green fluorescent protein (GFP) inserted into the Pitx3 locus via homologous recombination. In the central nervous system, Pitx3-directed GFP was visualized in dopaminergic (DA) neurons in the substantia nigra and ventral tegmental area. Live primary DA neurons can be isolated by fluorescence-activated cell sorting from these transgenic mouse embryos. In culture, Pitx3-GFP is coexpressed in a proportion of ES-derived DA neurons. Furthermore, ES cell-derived Pitx3-GFP expressing DA neurons responded to neurotrophic factors and were sensitive to DA-specific neurotoxin N-4-methyl-1, 2, 3, 6-tetrahydropyridine. We anticipate that the Pitx3-GFP ES cells could be used as a powerful model system for functional identification of molecules governing mDA neuron differentiation and for preclinical research including pharmaceutical drug screening and transplantation. The Pitx3 knock-in mice, on the other hand, could be used for purifying primary neurons for molecular studies associated with the midbrain-specific DA phenotype at a level not previously feasible. These mice would also provide a useful tool to study DA fate determination from embryo- or adult-derived neural stem cells.
authors.affiliation: Institute for Stem Cell Research, University of Edinburgh, King's Buildings, West Mains Road, Edinburgh, United Kingdom, EH9 3JQ, UK.
authors.forename: Suling
authors.initials: S
authors.lastname: Zhao
authors.forename: Sarah
authors.initials: S
authors.lastname: Maxwell
authors.forename: Antonio
authors.initials: A
authors.lastname: Jimenez-Beristain
authors.forename: Joaquim
authors.initials: J
authors.lastname: Vives
authors.forename: Eva
authors.initials: E
authors.lastname: Kuehner
authors.forename: Jiexin
authors.initials: J
authors.lastname: Zhao
authors.forename: Carmel
authors.initials: C
authors.lastname: O'Brien
authors.forename: Carmen
authors.initials: C
authors.lastname: de Felipe
authors.forename: Elena
authors.initials: E
authors.lastname: Semina
authors.forename: Meng
authors.initials: M
authors.lastname: Li
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D004298:Dopamine / Q000096:biosynthesis* / Q000235:genetics; D004622:Embryo, Mammalian; D005260:Female; D018507:Gene Expression Regulation, Developmental / Q000502:physiology; D049452:Green Fluorescent Proteins; D018398:Homeodomain Proteins / Q000096:biosynthesis / Q000235:genetics; D008164:Luminescent Proteins / Q000096:biosynthesis* / Q000235:genetics; D008297:Male; D008636:Mesencephalon / Q000166:cytology / Q000196:embryology / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D009474:Neurons / Q000166:cytology / Q000378:metabolism*; D011247:Pregnancy; D013234:Stem Cells / Q000166:cytology / Q000378:metabolism*; D014157:Transcription Factors / Q000096:biosynthesis / Q000235:genetics
chemical_list: D018398:Homeodomain Proteins; D008164:Luminescent Proteins; D014157:Transcription Factors; C433629:homeobox protein PITX3; D049452:Green Fluorescent Proteins; D004298:Dopamine