Abstract
In this protocol, we describe the procedures we have developed to optimize the performance of voltage-sensitive dyes for recording changes in neuronal electrical activity. We emphasize our experience in finding the best dye conditions for recording backpropagating action potentials from individual dendritic spines in a neuron within a brain slice. We fully describe procedures for loading the dye through a patch pipette and for finding excitation and emission wavelengths for the best sensitivity of the fluorescence signal to membrane voltage. Many of these approaches can be adapted to in vivo preparations and to experiments on mapping brain activity via optical recording.
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Acknowledgements
This work was supported by National Institutes of Health grants No. R01 EB001963 and No. P41 RR013186. We are grateful for the dedication of the people responsible for the synthesis of the voltage-sensitive dyes, Ping Yan and Joseph Wuskell.
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Acker, C.D., Loew, L.M. (2013). Characterization of Voltage-Sensitive Dyes in Living Cells Using Two-Photon Excitation. In: Banghart, M. (eds) Chemical Neurobiology. Methods in Molecular Biology, vol 995. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-345-9_11
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DOI: https://doi.org/10.1007/978-1-62703-345-9_11
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Publisher Name: Humana Press, Totowa, NJ
Print ISBN: 978-1-62703-344-2
Online ISBN: 978-1-62703-345-9
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