Dopaminergic neurons from embryonic mouse mesencephalon are enriched in culture through immunoreaction with monoclonal antibody to neural specific protein 4 and flow cytometry.

Abstract

Dopaminergic neurons represent a rare neurotransmitter phenotype within the mammalian central nervous system. The mesencephalic dopaminergic neurons form the ascending dopaminergic pathways in mammals and are involved in motor and limbic functions. Here we report that about 30% of all developing mouse mesencephalic cells, including virtually all of the dopaminergic phenotype, express surface membrane determinant(s) recognized by a monoclonal antibody to neural specific protein 4 (NSP4). We have been able to isolate and culture neurons from the mesencephalon according to their expression of NSP4, using the anti-NSP4 immunoreaction in conjunction with fluorescence-activated cell sorting. Cultures of NSP4+-sorted cells showed a significant enrichment in three morphologically distinct putative dopaminergic phenotypes when compared to unsorted mesencephalic cultures, whereas the cultures of NSP4--sorted cells were virtually devoid of dopaminergic neurons. This flow cytometric enrichment in dopaminergic neurons should provide the necessary cells for multidisciplinary study of dopaminergic phenotype differentiation.