Abstract
Hippocampal neurons growing in culture initially extend several, short minor processes that have the potential to become either axons or dendrites. The first expression of polarity occurs when one of these minor processes begins to elongate rapidly, becoming the axon. Before axonal outgrowth, the growth-associated protein GAP-43 is distributed equally among the growth cones of the minor processes; it is preferentially concentrated in the axonal growth cone once polarity has been established (Goslin, K., D. Schreyer, J. Skene, and G. Banker. 1990. J. Neurosci. 10:588-602). To determine when the selective segregation of GAP-43 begins, we followed individual cells by video microscopy, fixed them as soon as the axon could be distinguished, and localized GAP-43 by immunofluorescence microscopy. Individual minor processes acquired axonal growth characteristics within a period of 30- 60 min, and GAP-43 became selectively concentrated to the growth cones of these processes with an equally rapid time course. We also examined changes in the distribution of GAP-43 after transection of the axon. After an axonal transection that is distant from the soma, neuronal polarity is maintained, and the original axon begins to regrow almost immediately. In such cases, GAP-43 became selectively concentrated in the new axonal growth cone within 12-30 min. In contrast, when the axon is transected close to the soma, polarity is lost; the original axon rarely regrows, and there is a significant delay before a new axon emerges. Under these circumstances, GAP-43 accumulated in the new growth cone much more slowly, suggesting that its ongoing selective routing to the axon had been disrupted by the transection. These results demonstrate that the selective segregation of GAP-43 to the growth cone of a single process is closely correlated with the acquisition of axonal growth characteristics and, hence, with the expression of polarity.
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Selected References
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- Baas P. W., Black M. M., Banker G. A. Changes in microtubule polarity orientation during the development of hippocampal neurons in culture. J Cell Biol. 1989 Dec;109(6 Pt 1):3085–3094. doi: 10.1083/jcb.109.6.3085. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Banker G. A., Cowan W. M. Rat hippocampal neurons in dispersed cell culture. Brain Res. 1977 May 13;126(3):397–342. doi: 10.1016/0006-8993(77)90594-7. [DOI] [PubMed] [Google Scholar]
- Banker G. A. Trophic interactions between astroglial cells and hippocampal neurons in culture. Science. 1980 Aug 15;209(4458):809–810. doi: 10.1126/science.7403847. [DOI] [PubMed] [Google Scholar]
- Bartlett W. P., Banker G. A. An electron microscopic study of the development of axons and dendrites by hippocampal neurons in culture. I. Cells which develop without intercellular contacts. J Neurosci. 1984 Aug;4(8):1944–1953. doi: 10.1523/JNEUROSCI.04-08-01944.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Booher J., Sensenbrenner M. Growth and cultivation of dissociated neurons and glial cells from embryonic chick, rat and human brain in flask cultures. Neurobiology. 1972;2(3):97–105. [PubMed] [Google Scholar]
- Bottenstein J. E., Sato G. H. Growth of a rat neuroblastoma cell line in serum-free supplemented medium. Proc Natl Acad Sci U S A. 1979 Jan;76(1):514–517. doi: 10.1073/pnas.76.1.514. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Daniels M. P. Colchicine inhibition of nerve fiber formation in vitro. J Cell Biol. 1972 Apr;53(1):164–176. doi: 10.1083/jcb.53.1.164. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Dekker L. V., De Graan P. N., Oestreicher A. B., Versteeg D. H., Gispen W. H. Inhibition of noradrenaline release by antibodies to B-50 (GAP-43). Nature. 1989 Nov 2;342(6245):74–76. doi: 10.1038/342074a0. [DOI] [PubMed] [Google Scholar]
- Dotti C. G., Banker G. A. Experimentally induced alteration in the polarity of developing neurons. Nature. 1987 Nov 19;330(6145):254–256. doi: 10.1038/330254a0. [DOI] [PubMed] [Google Scholar]
- Dotti C. G., Sullivan C. A., Banker G. A. The establishment of polarity by hippocampal neurons in culture. J Neurosci. 1988 Apr;8(4):1454–1468. doi: 10.1523/JNEUROSCI.08-04-01454.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Gordon-Weeks P. R. Growth at the growth cone. Trends Neurosci. 1989 Jul;12(7):238–240. doi: 10.1016/0166-2236(89)90018-0. [DOI] [PubMed] [Google Scholar]
- Goslin K., Banker G. Experimental observations on the development of polarity by hippocampal neurons in culture. J Cell Biol. 1989 Apr;108(4):1507–1516. doi: 10.1083/jcb.108.4.1507. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Goslin K., Schreyer D. J., Skene J. H., Banker G. Changes in the distribution of GAP-43 during the development of neuronal polarity. J Neurosci. 1990 Feb;10(2):588–602. doi: 10.1523/JNEUROSCI.10-02-00588.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Goslin K., Schreyer D. J., Skene J. H., Banker G. Development of neuronal polarity: GAP-43 distinguishes axonal from dendritic growth cones. Nature. 1988 Dec 15;336(6200):672–674. doi: 10.1038/336672a0. [DOI] [PubMed] [Google Scholar]
- Johnson G. D., Nogueira Araujo G. M. A simple method of reducing the fading of immunofluorescence during microscopy. J Immunol Methods. 1981;43(3):349–350. doi: 10.1016/0022-1759(81)90183-6. [DOI] [PubMed] [Google Scholar]
- LaBate M. E., Skene J. H. Selective conservation of GAP-43 structure in vertebrate evolution. Neuron. 1989 Sep;3(3):299–310. doi: 10.1016/0896-6273(89)90254-7. [DOI] [PubMed] [Google Scholar]
- Meiri K. F., Pfenninger K. H., Willard M. B. Growth-associated protein, GAP-43, a polypeptide that is induced when neurons extend axons, is a component of growth cones and corresponds to pp46, a major polypeptide of a subcellular fraction enriched in growth cones. Proc Natl Acad Sci U S A. 1986 May;83(10):3537–3541. doi: 10.1073/pnas.83.10.3537. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Meiri K. F., Willard M., Johnson M. I. Distribution and phosphorylation of the growth-associated protein GAP-43 in regenerating sympathetic neurons in culture. J Neurosci. 1988 Jul;8(7):2571–2581. doi: 10.1523/JNEUROSCI.08-07-02571.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Selak I., Skaper S. D., Varon S. Pyruvate participation in the low molecular weight trophic activity for central nervous system neurons in glia-conditioned media. J Neurosci. 1985 Jan;5(1):23–28. doi: 10.1523/JNEUROSCI.05-01-00023.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Simons K., Fuller S. D. Cell surface polarity in epithelia. Annu Rev Cell Biol. 1985;1:243–288. doi: 10.1146/annurev.cb.01.110185.001331. [DOI] [PubMed] [Google Scholar]
- Skene J. H. Axonal growth-associated proteins. Annu Rev Neurosci. 1989;12:127–156. doi: 10.1146/annurev.ne.12.030189.001015. [DOI] [PubMed] [Google Scholar]
- Skene J. H., Jacobson R. D., Snipes G. J., McGuire C. B., Norden J. J., Freeman J. A. A protein induced during nerve growth (GAP-43) is a major component of growth-cone membranes. Science. 1986 Aug 15;233(4765):783–786. doi: 10.1126/science.3738509. [DOI] [PubMed] [Google Scholar]
- Skene J. H., Virág I. Posttranslational membrane attachment and dynamic fatty acylation of a neuronal growth cone protein, GAP-43. J Cell Biol. 1989 Feb;108(2):613–624. doi: 10.1083/jcb.108.2.613. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Skene J. H., Willard M. Changes in axonally transported proteins during axon regeneration in toad retinal ganglion cells. J Cell Biol. 1981 Apr;89(1):86–95. doi: 10.1083/jcb.89.1.86. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Zuber M. X., Strittmatter S. M., Fishman M. C. A membrane-targeting signal in the amino terminus of the neuronal protein GAP-43. Nature. 1989 Sep 28;341(6240):345–348. doi: 10.1038/341345a0. [DOI] [PubMed] [Google Scholar]