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. 1995 Dec 1;131(5):1315–1326. doi: 10.1083/jcb.131.5.1315

Axonal transport of mitochondria along microtubules and F-actin in living vertebrate neurons

PMCID: PMC2120647  PMID: 8522592

Abstract

A large body of evidence indicates that microtubules (MTs) conduct organelle transport in axons, but recent studies on extruded squid axoplasm have suggested that actin microfilaments (MFs) may also play a role in this process. To investigate the separate contributions to transport of each class of cytoskeletal element in intact vertebrate axons, we have monitored mitochondrial movements in chick sympathetic neurons experimentally manipulated to eliminate MTs, MFs, or both. First, we grew neurons in the continuous presence of: (a) cytochalasin E to create neurites which had never contained MFs; or (b) nocodazole or vinblastine to produce neurites which had never contained MTs. Mitochondria moved bidirectionally at normal velocities along the length of neurites which contained MTs and lacked MFs, but did not even enter neurites grown without MTs but containing MFs. In a second approach, we treated established neuronal cultures with cytoskeletal drugs to disrupt either MTs or MFs in axons already containing mitochondria. In cytochalasin-treated cells, which retained MTs but lacked MFs, average mitochondrial velocity increased in both directions, but net directional transport decreased. In vinblastine- treated cells, which lacked MTs but retained essentially normal levels of MFs, mitochondria continued to move bidirectionally but the average mitochondrial velocity and excursion length were reduced for both directions of movement, and the mitochondria spent threefold as much time moving in the retrograde as in the anterograde direction, resulting in net retrograde transport. Treatment of established cultures with both drugs produced neurites lacking MTs and MFs but still rich in neurofilaments; these showed a striking absence of any mitochondrial motility. These data indicate that axonal organelle transport can occur along both MTs and MFs in vivo, but with different velocities and net transport properties.

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Selected References

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  1. Allen R. D., Weiss D. G., Hayden J. H., Brown D. T., Fujiwake H., Simpson M. Gliding movement of and bidirectional transport along single native microtubules from squid axoplasm: evidence for an active role of microtubules in cytoplasmic transport. J Cell Biol. 1985 May;100(5):1736–1752. doi: 10.1083/jcb.100.5.1736. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bearer E. L., DeGiorgis J. A., Bodner R. A., Kao A. W., Reese T. S. Evidence for myosin motors on organelles in squid axoplasm. Proc Natl Acad Sci U S A. 1993 Dec 1;90(23):11252–11256. doi: 10.1073/pnas.90.23.11252. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bradley T. J., Satir P. Evidence of microfilament-associated mitochondrial movement. J Supramol Struct. 1979;12(2):165–175. doi: 10.1002/jss.400120203. [DOI] [PubMed] [Google Scholar]
  4. Brady S. T. A novel brain ATPase with properties expected for the fast axonal transport motor. Nature. 1985 Sep 5;317(6032):73–75. doi: 10.1038/317073a0. [DOI] [PubMed] [Google Scholar]
  5. Brady S. T., Crothers S. D., Nosal C., McClure W. O. Fast axonal transport in the presence of high Ca2+: evidence that microtubules are not required. Proc Natl Acad Sci U S A. 1980 Oct;77(10):5909–5913. doi: 10.1073/pnas.77.10.5909. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Brady S. T., Lasek R. J., Allen R. D. Video microscopy of fast axonal transport in extruded axoplasm: a new model for study of molecular mechanisms. Cell Motil. 1985;5(2):81–101. doi: 10.1002/cm.970050203. [DOI] [PubMed] [Google Scholar]
  7. Brady S. T., Lasek R. J., Allen R. D., Yin H. L., Stossel T. P. Gelsolin inhibition of fast axonal transport indicates a requirement for actin microfilaments. Nature. 1984 Jul 5;310(5972):56–58. doi: 10.1038/310056a0. [DOI] [PubMed] [Google Scholar]
  8. Bray D., Bunge M. B. Serial analysis of microtubules in cultured rat sensory axons. J Neurocytol. 1981 Aug;10(4):589–605. doi: 10.1007/BF01262592. [DOI] [PubMed] [Google Scholar]
  9. Breitling F., Little M. Carboxy-terminal regions on the surface of tubulin and microtubules. Epitope locations of YOL1/34, DM1A and DM1B. J Mol Biol. 1986 May 20;189(2):367–370. doi: 10.1016/0022-2836(86)90517-6. [DOI] [PubMed] [Google Scholar]
  10. Burridge K., Bray D. Purification and structural analysis of myosins from brain and other non-muscle tissues. J Mol Biol. 1975 Nov 25;99(1):1–14. doi: 10.1016/s0022-2836(75)80154-9. [DOI] [PubMed] [Google Scholar]
  11. Cheney R. E., O'Shea M. K., Heuser J. E., Coelho M. V., Wolenski J. S., Espreafico E. M., Forscher P., Larson R. E., Mooseker M. S. Brain myosin-V is a two-headed unconventional myosin with motor activity. Cell. 1993 Oct 8;75(1):13–23. doi: 10.1016/S0092-8674(05)80080-7. [DOI] [PubMed] [Google Scholar]
  12. Cheney R. E., Riley M. A., Mooseker M. S. Phylogenetic analysis of the myosin superfamily. Cell Motil Cytoskeleton. 1993;24(4):215–223. doi: 10.1002/cm.970240402. [DOI] [PubMed] [Google Scholar]
  13. Corthésy-Theulaz I., Pauloin A., Pfeffer S. R. Cytoplasmic dynein participates in the centrosomal localization of the Golgi complex. J Cell Biol. 1992 Sep;118(6):1333–1345. doi: 10.1083/jcb.118.6.1333. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Espreafico E. M., Cheney R. E., Matteoli M., Nascimento A. A., De Camilli P. V., Larson R. E., Mooseker M. S. Primary structure and cellular localization of chicken brain myosin-V (p190), an unconventional myosin with calmodulin light chains. J Cell Biol. 1992 Dec;119(6):1541–1557. doi: 10.1083/jcb.119.6.1541. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Fine R. E., Bray D. Actin in growing nerve cells. Nat New Biol. 1971 Nov 24;234(47):115–118. doi: 10.1038/newbio234115a0. [DOI] [PubMed] [Google Scholar]
  16. Forman D. S., Lynch K. J., Smith R. S. Organelle dynamics in lobster axons: anterograde, retrograde and stationary mitochondria. Brain Res. 1987 May 26;412(1):96–106. doi: 10.1016/0006-8993(87)91443-0. [DOI] [PubMed] [Google Scholar]
  17. Goldberg D. J., Harris D. A., Lubit B. W., Schwartz J. H. Analysis of the mechanism of fast axonal transport by intracellular injection of potentially inhibitory macromolecules: evidence for a possible role of actin filaments. Proc Natl Acad Sci U S A. 1980 Dec;77(12):7448–7452. doi: 10.1073/pnas.77.12.7448. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Goldberg D. J. Microinjection into an identified axon to study the mechanism of fast axonal transport. Proc Natl Acad Sci U S A. 1982 Aug;79(15):4818–4822. doi: 10.1073/pnas.79.15.4818. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Grafstein B., Forman D. S. Intracellular transport in neurons. Physiol Rev. 1980 Oct;60(4):1167–1283. doi: 10.1152/physrev.1980.60.4.1167. [DOI] [PubMed] [Google Scholar]
  20. Hasson T., Mooseker M. S. Porcine myosin-VI: characterization of a new mammalian unconventional myosin. J Cell Biol. 1994 Oct;127(2):425–440. doi: 10.1083/jcb.127.2.425. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hirokawa N. Cross-linker system between neurofilaments, microtubules, and membranous organelles in frog axons revealed by the quick-freeze, deep-etching method. J Cell Biol. 1982 Jul;94(1):129–142. doi: 10.1083/jcb.94.1.129. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Ho W. C., Allan V. J., van Meer G., Berger E. G., Kreis T. E. Reclustering of scattered Golgi elements occurs along microtubules. Eur J Cell Biol. 1989 Apr;48(2):250–263. [PubMed] [Google Scholar]
  23. Hollenbeck P. J., Bray D. Rapidly transported organelles containing membrane and cytoskeletal components: their relation to axonal growth. J Cell Biol. 1987 Dec;105(6 Pt 1):2827–2835. doi: 10.1083/jcb.105.6.2827. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Hollenbeck P. J. Products of endocytosis and autophagy are retrieved from axons by regulated retrograde organelle transport. J Cell Biol. 1993 Apr;121(2):305–315. doi: 10.1083/jcb.121.2.305. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Hollenbeck P. J., Swanson J. A. Radial extension of macrophage tubular lysosomes supported by kinesin. Nature. 1990 Aug 30;346(6287):864–866. doi: 10.1038/346864a0. [DOI] [PubMed] [Google Scholar]
  26. 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]
  27. Johnson L. V., Walsh M. L., Bockus B. J., Chen L. B. Monitoring of relative mitochondrial membrane potential in living cells by fluorescence microscopy. J Cell Biol. 1981 Mar;88(3):526–535. doi: 10.1083/jcb.88.3.526. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Kuznetsov S. A., Langford G. M., Weiss D. G. Actin-dependent organelle movement in squid axoplasm. Nature. 1992 Apr 23;356(6371):722–725. doi: 10.1038/356722a0. [DOI] [PubMed] [Google Scholar]
  29. Kuznetsov S. A., Rivera D. T., Severin F. F., Weiss D. G., Langford G. M. Movement of axoplasmic organelles on actin filaments from skeletal muscle. Cell Motil Cytoskeleton. 1994;28(3):231–242. doi: 10.1002/cm.970280306. [DOI] [PubMed] [Google Scholar]
  30. Langford G. M., Kuznetsov S. A., Johnson D., Cohen D. L., Weiss D. G. Movement of axoplasmic organelles on actin filaments assembled on acrosomal processes: evidence for a barbed-end-directed organelle motor. J Cell Sci. 1994 Aug;107(Pt 8):2291–2298. doi: 10.1242/jcs.107.8.2291. [DOI] [PubMed] [Google Scholar]
  31. Li D., Chantler P. D. Evidence for a new member of the myosin I family from mammalian brain. J Neurochem. 1992 Oct;59(4):1344–1351. doi: 10.1111/j.1471-4159.1992.tb08446.x. [DOI] [PubMed] [Google Scholar]
  32. Li D., Miller M., Chantler P. D. Association of a cellular myosin II with anionic phospholipids and the neuronal plasma membrane. Proc Natl Acad Sci U S A. 1994 Feb 1;91(3):853–857. doi: 10.1073/pnas.91.3.853. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Magrassi L., Purves D., Lichtman J. W. Fluorescent probes that stain living nerve terminals. J Neurosci. 1987 Apr;7(4):1207–1214. doi: 10.1523/JNEUROSCI.07-04-01207.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Marsh L., Letourneau P. C. Growth of neurites without filopodial or lamellipodial activity in the presence of cytochalasin B. J Cell Biol. 1984 Dec;99(6):2041–2047. doi: 10.1083/jcb.99.6.2041. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Martz D., Lasek R. J., Brady S. T., Allen R. D. Mitochondrial motility in axons: membranous organelles may interact with the force generating system through multiple surface binding sites. Cell Motil. 1984;4(2):89–101. doi: 10.1002/cm.970040203. [DOI] [PubMed] [Google Scholar]
  36. Mochida S., Kobayashi H., Matsuda Y., Yuda Y., Muramoto K., Nonomura Y. Myosin II is involved in transmitter release at synapses formed between rat sympathetic neurons in culture. Neuron. 1994 Nov;13(5):1131–1142. doi: 10.1016/0896-6273(94)90051-5. [DOI] [PubMed] [Google Scholar]
  37. Mooseker M. Myosin superfamily: a multitude of myosins. Curr Biol. 1993 Apr 1;3(4):245–248. doi: 10.1016/0960-9822(93)90346-p. [DOI] [PubMed] [Google Scholar]
  38. Morris R. L., Hollenbeck P. J. The regulation of bidirectional mitochondrial transport is coordinated with axonal outgrowth. J Cell Sci. 1993 Mar;104(Pt 3):917–927. doi: 10.1242/jcs.104.3.917. [DOI] [PubMed] [Google Scholar]
  39. Osborn M., Webster R. E., Weber K. Individual microtubules viewed by immunofluorescence and electron microscopy in the same PtK2 cell. J Cell Biol. 1978 Jun;77(3):R27–R34. doi: 10.1083/jcb.77.3.r27. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Provance D. W., Jr, McDowall A., Marko M., Luby-Phelps K. Cytoarchitecture of size-excluding compartments in living cells. J Cell Sci. 1993 Oct;106(Pt 2):565–577. doi: 10.1242/jcs.106.2.565. [DOI] [PubMed] [Google Scholar]
  41. Schnapp B. J., Reese T. S. Dynein is the motor for retrograde axonal transport of organelles. Proc Natl Acad Sci U S A. 1989 Mar;86(5):1548–1552. doi: 10.1073/pnas.86.5.1548. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Schnapp B. J., Vale R. D., Sheetz M. P., Reese T. S. Single microtubules from squid axoplasm support bidirectional movement of organelles. Cell. 1985 Feb;40(2):455–462. doi: 10.1016/0092-8674(85)90160-6. [DOI] [PubMed] [Google Scholar]
  43. Schroer T. A., Steuer E. R., Sheetz M. P. Cytoplasmic dynein is a minus end-directed motor for membranous organelles. Cell. 1989 Mar 24;56(6):937–946. doi: 10.1016/0092-8674(89)90627-2. [DOI] [PubMed] [Google Scholar]
  44. Sheetz M. P., Spudich J. A. Movement of myosin-coated fluorescent beads on actin cables in vitro. Nature. 1983 May 5;303(5912):31–35. doi: 10.1038/303031a0. [DOI] [PubMed] [Google Scholar]
  45. Shpetner H. S., Paschal B. M., Vallee R. B. Characterization of the microtubule-activated ATPase of brain cytoplasmic dynein (MAP 1C). J Cell Biol. 1988 Sep;107(3):1001–1009. doi: 10.1083/jcb.107.3.1001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Sun W. D., Chantler P. D. A unique cellular myosin II exhibiting differential expression in the cerebral cortex. Biochem Biophys Res Commun. 1991 Feb 28;175(1):244–249. doi: 10.1016/s0006-291x(05)81226-4. [DOI] [PubMed] [Google Scholar]
  47. Sun W., Chantler P. D. Cloning of the cDNA encoding a neuronal myosin heavy chain from mammalian brain and its differential expression within the central nervous system. J Mol Biol. 1992 Apr 20;224(4):1185–1193. doi: 10.1016/0022-2836(92)90482-y. [DOI] [PubMed] [Google Scholar]
  48. Terasaki M., Chen L. B., Fujiwara K. Microtubules and the endoplasmic reticulum are highly interdependent structures. J Cell Biol. 1986 Oct;103(4):1557–1568. doi: 10.1083/jcb.103.4.1557. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Terasaki M., Song J., Wong J. R., Weiss M. J., Chen L. B. Localization of endoplasmic reticulum in living and glutaraldehyde-fixed cells with fluorescent dyes. Cell. 1984 Aug;38(1):101–108. doi: 10.1016/0092-8674(84)90530-0. [DOI] [PubMed] [Google Scholar]
  50. Vale R. D., Reese T. S., Sheetz M. P. Identification of a novel force-generating protein, kinesin, involved in microtubule-based motility. Cell. 1985 Aug;42(1):39–50. doi: 10.1016/s0092-8674(85)80099-4. [DOI] [PMC free article] [PubMed] [Google Scholar]
  51. Vallee R. B., Wall J. S., Paschal B. M., Shpetner H. S. Microtubule-associated protein 1C from brain is a two-headed cytosolic dynein. Nature. 1988 Apr 7;332(6164):561–563. doi: 10.1038/332561a0. [DOI] [PubMed] [Google Scholar]

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