title: Dynamic visualization of membrane-inserted fraction of pHluorin-tagged channels using repetitive acidification technique.
abstract: BACKGROUND
  Changes in neuronal excitability, synaptic efficacy and generally in cell signaling often result from insertion of key molecules into plasma membrane (PM). Many of the techniques used for monitoring PM insertion lack either spatial or temporal resolution.
  
  
  RESULTS
  We improved the imaging method based on time-lapse total internal reflection fluorescence (TIRF) microscopy and pHluorin tagging by supplementing it with a repetitive extracellular acidification protocol. We illustrate the applicability of this method by showing that brief activation of NMDA receptors ("chemical LTP") in cultured hippocampal neurons induced a persistent PM insertion of glutamate receptors containing the pHluorin-tagged GluR-A(flip) subunits.
  
  
  CONCLUSIONS
  The repetitive acidification technique provides a more accurate way of monitoring the PM-inserted fraction of fluorescently tagged molecules and offers a good temporal and spatial resolution.
authors: Neuroscience Center, University of Helsinki, FIN-00014, Helsinki, Finland. sekhirug@mappi.helsinki.fi
authors: Serguei S
authors: SS
authors: Khiroug
authors: Evgeny
authors: E
authors: Pryazhnikov
authors: Sarah K
authors: SK
authors: Coleman
authors: Andreas
authors: A
authors: Jeromin
authors: Kari
authors: K
authors: Keinänen
authors: Leonard
authors: L
authors: Khiroug
mesh_terms: D002462:Cell Membrane / Q000648:ultrastructure*; D002478:Cells, Cultured; D049452:Green Fluorescent Proteins / Q000032:analysis*; D006624:Hippocampus / Q000166:cytology*; D008856:Microscopy, Fluorescence / Q000379:methods*; D009473:Neuronal Plasticity; D009474:Neurons; D016194:Receptors, N-Methyl-D-Aspartate / Q000648:ultrastructure*; D009435:Synaptic Transmission
chemical_list: C485187:PHluorin; D016194:Receptors, N-Methyl-D-Aspartate; D049452:Green Fluorescent Proteins