Kinetic Measurements of Endocytosis and Exocytosis in Cultured Neurons

Karina J Vargas

doi:10.1007/978-1-4939-9124-2_3

Kinetic Measurements of Endocytosis and Exocytosis in Cultured Neurons

Methods Mol Biol. 2019:1948:23-33. doi: 10.1007/978-1-4939-9124-2_3.

Author

Karina J Vargas^{1

2

3}

Affiliations

¹ Department of Neurology, Yale University, New Haven, CT, USA. [email protected].
² Program in Cellular Neuroscience, Neurodegeneration and Repair, Yale University, New Haven, CT, USA. [email protected].
³ Eugene Bell Center for Regenerative Biology and Tissue Engineering, Marine Biology Laboratory, Woods Hole, MA, USA. [email protected].

PMID: 30771167
DOI: 10.1007/978-1-4939-9124-2_3

Abstract

The measurement of synaptic vesicle recycling in live neurons transfected with vesicular glutamate transporter fused to pHluorin (vGLUT-pHluorin) allows us to study exocytosis and endocytosis in neurons. When neurons are transfected with this protein we can measure the rate of vesicles fusing and internalizing from the membrane using live total internal reflection fluorescence (TIRF) imaging. Here, we describe transfection, culturing, and imaging of wild-type and αβγ-synuclein knockout hippocampal neurons. This technique can be used to evaluate the effect of different phenotypes and treatments in the physiology of synaptic vesicle recycling in cultured neurons.

Keywords: Endocytosis; Exocytosis; Imaging; Neuron culturing; Parkinson’s disease; Synaptic vesicle recycling; Synuclein; vGLUT-pHluorin.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Biomarkers
Cells, Cultured
Endocytosis*
Exocytosis*
Image Processing, Computer-Assisted
Mice
Molecular Imaging
Neurons / physiology*
Pyramidal Cells / physiology
Synaptic Vesicles / metabolism

Substances

Biomarkers