Abstract
The normal function of neurons depends on the integrity of microtubule-dependent transport of cellular materials and organelles to/from their cell bodies or axon terminus. In this chapter, we describe the design and implementation of a fluorescence imaging method to visualize axonal transport in neurons directly. We combine a pseudo total internal reflection microscopy, quantum dot fluorescence labeling, microfluidic neuronal culture chamber, and single molecule detection methods to achieve a high spatial and temporal resolution in tracking nerve growth factor transport in dorsal root ganglia neurons.
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References
Huang, E. J. & Reichardt, L. F. (2003) Trk receptors: roles in neuronal signal transduction. Annu Rev Biochem 72, 609–642.
Zweifel, L. S., Kuruvilla, R. & Ginty, D. D. (2005) Functions and mechanisms of retrograde neurotrophin signalling. Nat Rev Neurosci 6, 615–625.
Huang, E. J. & Reichardt, L. F. (2001) Neurotrophins: roles in neuronal development and function. Annu Rev Neurosci 24, 677–736.
Senger D. L. & Campenot, R. B. (1997) Rapid retrograde tyrosine phosphorylation of trkA and other proteins in rat sympathetic neurons in compartmented cultures. J Cell Biol 138, 411–421.
Korsching S. & Thoenen H. (1983) Quantitative demonstration of the retrograde axonal transport of endogenous nerve growth factor. Neurosci Lett 39, 1–4.
von Bartheld C. S. (2001) Tracing with radiolabeled neurotrophins. Methods Mol Biol 169, 195–216.
Jullien J., Guili, V., Derrington E. A., Darlix J., Reichardt L. F. & Rudkin, B. B. (2003) Trafficking of TrkA-green fluorescent protein chimerae during nerve growth factor-induced differentiation. J Biol Chem 278, 8706–8716.
Lalli G. & Schiavo G. (2002) Analysis of retrograde transport in motor neurons reveals common endocytic carriers for tetanus toxin and neurotrophin receptor p75. J Cell Biol 156, 233–240.
Tani T., Miyamoto Y., Fujimori K. E., Taguchi T., Yanagia T., Sako Y. & Harada Y. (2005) Trafficking a ligand-receptor complex on the growth cones as an essential step for the uptake of nerve growth factor at the distal end of the axon: a single-molecule analysis. J Neurosci 25, 2181–2191.
Cui B., Wu C., Chen L., Ramirez A., Bearer E. L., Li W., Mobley W. C. & Chu S. (2007) One at a time, live tracking of NGF axonal transport using quantum dots. Proc Natl Acad Sci U S A 104, 13666–13671.
Mobley, W. C., Schenker, A. & Shooter, E. M., (1976) Characterization and isolation of proteolytically modified nerve growth factor. Biochemistry 15, 5543–5552.
Taylor A. M., Rhee S. W., Tu C. H., Cribbs D. H., Cotman C. W. & Jeon N. L. (2003) Microfluidic multicompartment device for neuroscience research. Langmuir 19, 1551–1556.
Bronfman F. C., Tcherpakov M., Jovin T. M. & Fainzilber M. (2003) Ligand-induced internalization of the p75 neurotrophin receptor: a slow route to the signaling endosome. J Neurosci 23, 3209–3220.
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Osakada, Y., Cui, B. (2010). Real-Time Visualization of Axonal Transport in Neurons. In: Roberson, E. (eds) Alzheimer's Disease and Frontotemporal Dementia. Methods in Molecular Biology, vol 670. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-60761-744-0_16
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DOI: https://doi.org/10.1007/978-1-60761-744-0_16
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