title: Studying endosomes in cultured neurons by live-cell imaging.
abstract: Endosomes play critical roles on regulating surface receptor levels as well as signaling cascades in all cell types, including neurons. Endocytosis and endosomal trafficking is routinely studied after fixation, but live imaging is increasingly being used to capture the dynamic nature of endosomes and is allowing increasingly sophisticated glimpses into trafficking processes in live neurons. In this chapter, we describe the basics of neuronal primary cultures, methods for expressing fluorescent proteins, and live imaging of cargos and endosomal regulators.
authors.affiliation: Children's National Medical Center, Washington, DC, USA.
authors.forename: Zofia M
authors.initials: ZM
authors.lastname: Lasiecka
authors.affiliation: Department of Neuroscience, University of Virginia Medical School, Charlottesville, VA, USA.
authors.forename: Bettina
authors.initials: B
authors.lastname: Winckler
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D018274:Electroporation / Q000379:methods; D004622:Embryo, Mammalian / Q000166:cytology / Q000294:innervation; D004705:Endocytosis / Q000502:physiology*; D011992:Endosomes / Q000378:metabolism*; D005456:Fluorescent Dyes; D006624:Hippocampus / Q000166:cytology*; D016086:Lentivirus / Q000235:genetics; D008565:Membrane Proteins / Q000378:metabolism; D051379:Mice; D018613:Microscopy, Confocal; D061251:Primary Cell Culture; D021381:Protein Transport / Q000502:physiology*; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D051381:Rats; D013194:Staining and Labeling; D014162:Transfection / Q000379:methods
chemical_list: D005456:Fluorescent Dyes; D008565:Membrane Proteins; D034741:RNA, Small Interfering
keywords: Endocytic cargo; Endocytosis; Endosome dynamics; Internalization; Live imaging; Live imaging chamber; Neuronal culture; Primary neurons; Pulse-chase experiments; Rab proteins