UNIT 2.7 Characterizing Synaptic Vesicle Proteins Using Synaptosomal Fractions and Cultured Hippocampal Neurons
Published Online: 1 APR 2012
Copyright © 2012 by John Wiley & Sons, Inc.
Lab Protocol Title
Current Protocols in Neuroscience
How to Cite
DiGiovanni, J., Sun, T. and Sheng, Z.-H. 2012. Characterizing Synaptic Vesicle Proteins Using Synaptosomal Fractions and Cultured Hippocampal Neurons. Current Protocols in Neuroscience. 59:2.7.1–2.7.22.
- Published Online: 1 APR 2012
- Published Print: APR 2012
Cloning and characterization of synaptic vesicle proteins and their binding counterparts on the presynaptic plasma membrane have greatly advanced our understanding of the molecular mechanisms involved in the synaptic vesicle cycle and neurotransmitter release. This unit discusses multidisciplinary approaches to characterize proteins from synaptosome-enriched subcellular fractions and localize them within cultured neurons. The first approach regroups methods used to isolate synaptic vesicles from rat brain synaptosomal preparations, allowing for specific biochemical investigation of synaptic vesicle proteins. The second is a detailed procedure for pre-embedding immunogold staining and electron microscopic observation, which permits the morphological identification of proteins in individual vesicles at intact synapses. Additionally, this chapter proposes methods for light microscopic examination of hippocampal neurons. It includes procedures for embryonic and postnatal hippocampal neuron culture and describes an immunocytochemical staining protocol used to investigate synaptic vesicle protein localization with respect to other proteins or subcellular structures. Curr. Protoc. Neurosci. 59:2.7.1-2.7.22. © 2012 by John Wiley & Sons, Inc.
- hippocampal neurons;
- light microscopy;
- electron microscopy;