Regionalization of plasma membrane-bound flavoproteins of cerebellar granule neurons in culture by fluorescence energy transfer imaging

Journal of Fluorescence
Alejandro K Samhan-AriasCarlos Gutiérrez-Merino

Abstract

Flavoproteins are components of plasma membrane redox chains, which have been suggested to play major roles in neuronal activity and survival. We found that the red/orange autofluorescence of mature primary cultures of cerebellar granule neurons (8-9 days in vitro) was largely quenched by millimolar concentrations of dithionite added to the extracellular medium, and pointed out that nearly 50% of this autofluorescence was due to plasma membrane-bound flavoproteins. We report in this work that the lipophilic neuronal plasma membrane markers N-(3-triethylammoniumpropyl)-4-(4-(4-(diethylamino)phenyl)butadienyl)-pyridinium dibromide (RH-414) and N-(3-triethylammoniumpropyl)-4-(6-(4-(diethylamino)phenyl)hexatrienyl)pyridinium dibromide (FM4-64) can form fluorescence energy transfer donor-acceptor pairs with flavoproteins with calculated R (0) values between 3.7 and 4.2 nm. The quantification of the efficiency of fluorescence energy transfer with different concentrations of acceptor dyes has been worked out with re-suspended neurons. Using quantitative images of the neurons in culture, acquired with a CCD camera attached to an epifluorescence microscope, regionalization of the plasma membrane-bound flavoproteins of cerebellar granule...Continue Reading

References

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Citations

Nov 1, 2008·Molecular and Cellular Neurosciences·Alejandro K Samhan-AriasCarlos Gutierrez-Merino
Mar 7, 2019·Journal of Biological Inorganic Chemistry : JBIC : a Publication of the Society of Biological Inorganic Chemistry·Alejandro K Samhan-AriasJosé J G Moura

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