Abstract
Here, we describe the use of 2-photon fluorescence lifetime imaging (2pFLIM) of a Förster resonance energy transfer (FRET) sensor to study the spatial and temporal activity pattern of the BDNF receptor TrkB. Combining 2pFLIM with laser uncaging of MNI-glutamate over single dendritic spines in organotypic hippocampal slices allows measurement of glutamate-induced changes in TrkB activity within a single spine or dendrite. This protocol covers the installation and setup of requisite hardware and software, the use of the software to acquire data, and the analysis of the data. A prerequisite for attempting this protocol is proficiency with 2-photon imaging of live cells and a microscope that is controlled by ScanImage 3.8 (Vidrio Technologies).
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Acknowledgment
This work was supported by NS05621 (NIH) to JOM.
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Hall, C.E., McNamara, J.O., Yasuda, R. (2018). Analysis of TrkB Receptor Activity Using FRET Sensors. In: Duarte, C., Tongiorgi, E. (eds) Brain-Derived Neurotrophic Factor (BDNF). Neuromethods, vol 143. Humana, New York, NY. https://doi.org/10.1007/7657_2018_12
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DOI: https://doi.org/10.1007/7657_2018_12
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Publisher Name: Humana, New York, NY
Print ISBN: 978-1-4939-8969-0
Online ISBN: 978-1-4939-8970-6
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