![[Feedback]](/icons/banner/feedbackACT.gif){kind=icon}
![[For Subscribers]](/icons/banner/subscriberACT.gif){kind=icon}
![[Archive]](/icons/banner/archiveACT.gif){kind=icon}
![[Search]](/icons/banner/searchACT.gif){kind=icon}
![[Contents]](/icons/banner/tocACT.gif){kind=icon}
|
|
|
|
|
||
Vol. 12, Issue 10, 3257-3267, October 2001
Neuroscience Program, Department of Biological Sciences, Ohio
University, Athens, Ohio 45701
Observations on naturally occurring gaps in the axonal
neurofilament array of cultured neurons have demonstrated that
neurofilament polymers move along axons in a rapid, intermittent, and
highly asynchronous manner. In contrast, studies on axonal
neurofilaments using laser photobleaching have not detected movement.
Here, we describe a modified photobleaching strategy that does permit
the direct observation of neurofilament movement. Axons of cultured neurons expressing GFP-tagged neurofilament protein were bleached by
excitation with the mercury arc lamp of a conventional epifluorescence microscope for 12-60 s. The length of the bleached region ranged from
10 to 60 µm. By bleaching thin axons, which have relatively few
neurofilaments, we were able to reduce the fluorescent intensity enough
to allow the detection of neurofilaments that moved in from the
surrounding unbleached regions. Time-lapse imaging at short intervals
revealed rapid, intermittent, and highly asynchronous movement of
fluorescent filaments through the bleached regions at peak rates of up
to 2.8 µm/s. The kinetics of movement were very similar to our
previous observations on neurofilaments moving through naturally
occurring gaps, which indicates that the movement was not impaired by
the photobleaching process. These results demonstrate that fluorescence
photobleaching can be used to study the slow axonal transport of
cytoskeletal polymers, but only if the experimental strategy is
designed to ensure that rapid asynchronous movements can be detected.
This may explain the failure of previous photobleaching studies to
reveal the movement of neurofilament proteins and other cytoskeletal
proteins in axons.
The online version of this manuscript contains video
material for Figures 2, 3, 6, and 7. The online version is available at
www.molbiolcell.org.
*
Corresponding author. E-mail address:
brown.2302{at}osu.edu. Present address: Neurobiotechnology Center and
Department of Neuroscience, The Ohio State University, Rightmire Hall,
1060 Carmack Road, Columbus OH 43210.
This article has been cited by other articles:
|
|
 |
|
  S. Y. Shim, B. A. Samuels, J. Wang, G. Neumayer, C. Belzil, R. Ayala, Y. Shi, Y. Shi, L.-H. Tsai, and M. D. Nguyen Ndel1 Controls the Dynein-mediated Transport of Vimentin during Neurite Outgrowth J. Biol. Chem., May 2, 2008; 283(18): 12232 - 12240. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 N. Trivedi, P. Jung, and A. Brown Neurofilaments Switch between Distinct Mobile and Stationary States during Their Transport along Axons J. Neurosci., January 17, 2007; 27(3): 507 - 516. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 M. A. Utton, W. J. Noble, J. E. Hill, B. H. Anderton, and D. P. Hanger Molecular motors implicated in the axonal transport of tau and {alpha}-synuclein J. Cell Sci., October 15, 2005; 118(20): 4645 - 4654. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 A. Brown, L. Wang, and P. Jung Stochastic Simulation of Neurofilament Transport in Axons: The "Stop-and-Go" Hypothesis Mol. Biol. Cell, September 1, 2005; 16(9): 4243 - 4255. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Yan and A. Brown Neurofilament Polymer Transport in Axons J. Neurosci., July 27, 2005; 25(30): 7014 - 7021. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 Y. He, F. Francis, K. A. Myers, W. Yu, M. M. Black, and P. W. Baas Role of cytoplasmic dynein in the axonal transport of microtubules and neurofilaments J. Cell Biol., February 28, 2005; 168(5): 697 - 703. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
W. K.-H. Chan, A. Dickerson, D. Ortiz, A. F. Pimenta, C. M. Moran, J. Motil, S. J. Snyder, K. Malik, H. C. Pant, and T. B. Shea Mitogen-activated protein kinase regulates neurofilament axonal transport J. Cell Sci., September 15, 2004; 117(20): 4629 - 4642. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Uchida and A. Brown Arrival, Reversal, and Departure of Neurofilaments at the Tips of Growing Axons Mol. Biol. Cell, September 1, 2004; 15(9): 4215 - 4225. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
R. Windoffer, S. Woll, P. Strnad, and R. E. Leube Identification of Novel Principles of Keratin Filament Network Turnover in Living Cells Mol. Biol. Cell, May 1, 2004; 15(5): 2436 - 2448. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. T. Helfand, L. Chang, and R. D. Goldman Intermediate filaments are dynamic and motile elements of cellular architecture J. Cell Sci., January 15, 2004; 117(2): 133 - 141. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
B. T. Helfand, P. Loomis, M. Yoon, and R. D. Goldman Rapid transport of neural intermediate filament protein J. Cell Sci., June 1, 2003; 116(11): 2345 - 2359. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. Ackerley, P. Thornhill, A. J. Grierson, J. Brownlees, B. H. Anderton, P. N. Leigh, C. E. Shaw, and C. C.J. Miller Neurofilament heavy chain side arm phosphorylation regulates axonal transport of neurofilaments J. Cell Biol., May 12, 2003; 161(3): 489 - 495. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
A. Brown Axonal transport of membranous and nonmembranous cargoes: a unified perspective J. Cell Biol., March 17, 2003; 160(6): 817 - 821. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 J. Brownlees, S. Ackerley, A. J. Grierson, N. J.O. Jacobsen, K. Shea, B. H. Anderton, P. N. Leigh, C. E. Shaw, and C. C.J. Miller Charcot-Marie-Tooth disease neurofilament mutations disrupt neurofilament assembly and axonal transport Hum. Mol. Genet., November 1, 2002; 11(23): 2837 - 2844. [Abstract] [Full Text] [PDF]
|
|
