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Proceedings of the National Academy of Sciences of the United States of America logoLink to Proceedings of the National Academy of Sciences of the United States of America
. 1980 Dec;77(12):7448–7452. doi: 10.1073/pnas.77.12.7448

Analysis of the mechanism of fast axonal transport by intracellular injection of potentially inhibitory macromolecules: evidence for a possible role of actin filaments.

D J Goldberg, D A Harris, B W Lubit, J H Schwartz
PMCID: PMC350521  PMID: 6164061

Abstract

Although actin is thought to participate in several types of cell motility other than muscle contraction, no direct evidence has linked it to the force-generating mechanism for fast axonal transport. We have obtained evidence for the involvement of actin by microinjecting, into the serotonergic giant cerebral neuron of Aplysia, two preparations that have been shown to depolymerize actin filaments. One is a fraction of rabbit serum containing a heat-labile gamma globulin that affects actin polymerization in a manner similar to that of cytochalasin and several proteins that are thought to regulate the length of actin filaments. The other is bovine pancreatic DNase I which binds to actin stoichiometrically. Both preparations substantially decreased the transport of storage vesicles containing [3H]serotonin. Phalloidin, a toxic fungal peptide that binds to actin filaments but stabilizes rather than depolymerizes them, did not inhibit transport. We have not yet determined whether the inhibition od transport occurs during export of [3H]serotonin from the cell body into the axon or during translocation along the axon. Nevertheless, these observations provide a promising experimental indication that actin is involved in fast axonal transport.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Ambron R. T., Goldman J. E., Shkolnik L. J., Schwartz J. H. Synthesis and axonal transport of membrane glycoproteins in an identified serotonergic neuron of Aplysia. J Neurophysiol. 1980 Apr;43(4):929–944. doi: 10.1152/jn.1980.43.4.929. [DOI] [PubMed] [Google Scholar]
  2. Atlas S. J., Lin S. Dihydrocytochalasin B. Biological effects and binding to 3T3 cells. J Cell Biol. 1978 Feb;76(2):360–370. doi: 10.1083/jcb.76.2.360. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Banks P., Mayor D., Mraz P. Cytochalasin B and the intra-axonal movement of noradrenaline storage vesicles. Brain Res. 1973 Jan 30;49(2):417–421. doi: 10.1016/0006-8993(73)90433-2. [DOI] [PubMed] [Google Scholar]
  4. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  5. Brenner S. L., Korn E. D. The effects of cytochalasins on actin polymerization and actin ATPase provide insights into the mechanism of polymerization. J Biol Chem. 1980 Feb 10;255(3):841–844. [PubMed] [Google Scholar]
  6. Brown S. S., Spudich J. A. Cytochalasin inhibits the rate of elongation of actin filament fragments. J Cell Biol. 1979 Dec;83(3):657–662. doi: 10.1083/jcb.83.3.657. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Clarke M., Spudich J. A. Nonmuscle contractile proteins: the role of actin and myosin in cell motility and shape determination. Annu Rev Biochem. 1977;46:797–822. doi: 10.1146/annurev.bi.46.070177.004053. [DOI] [PubMed] [Google Scholar]
  8. Eisenstadt M., Goldman J. E., Kandel E. R., Koike H., Koester J., Schwartz J. H. Intrasomatic injection of radioactive precursors for studying transmitter synthesis in identified neurons of Aplysia californica. Proc Natl Acad Sci U S A. 1973 Dec;70(12):3371–3375. doi: 10.1073/pnas.70.12.3371. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Fernandez H. L., Samson F. E. Axoplasmic transport: differential inhibition by cytochalasin-B. J Neurobiol. 1973;4(3):201–206. doi: 10.1002/neu.480040305. [DOI] [PubMed] [Google Scholar]
  10. Giller E., Jr, Schwartz J. H. Choline acetyltransferase in identified neurons of abdominal ganglion of Aplysia californica. J Neurophysiol. 1971 Jan;34(1):93–107. doi: 10.1152/jn.1971.34.1.93. [DOI] [PubMed] [Google Scholar]
  11. Goldberg D. J., Schwartz J. H., Sherbany A. A. Kinetic properties of normal and perturbed axonal transport of serotonin in a single identified axon. J Physiol. 1978 Aug;281:559–579. doi: 10.1113/jphysiol.1978.sp012439. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Goldman J. E., Kim K. S., Schwartz J. H. Axonal transport of [3H]serotonin in an identified neuron of Aplysia californica. J Cell Biol. 1976 Aug;70(2 Pt 1):304–318. doi: 10.1083/jcb.70.2.304. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Goldman J. E., Schwartz J. H. Cellular specificity of serotonin storage and axonal transport in identified neurones of Aplysia californica. J Physiol. 1974 Oct;242(1):61–76. doi: 10.1113/jphysiol.1974.sp010694. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Goldman J. E., Schwartz J. H. Metabolism of [3H]serotonin in the marine mollusc, Aplysia californica. Brain Res. 1977 Nov 4;136(1):77–88. doi: 10.1016/0006-8993(77)90133-0. [DOI] [PubMed] [Google Scholar]
  15. 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]
  16. Hasegawa T., Takahashi S., Hayashi H., Hatano S. Fragmin: a calcium ion sensitive regulatory factor on the formation of actin filaments. Biochemistry. 1980 Jun 10;19(12):2677–2683. doi: 10.1021/bi00553a021. [DOI] [PubMed] [Google Scholar]
  17. Hitchcock S. E., Carisson L., Lindberg U. Depolymerization of F-actin by deoxyribonuclease I. Cell. 1976 Apr;7(4):531–542. doi: 10.1016/0092-8674(76)90203-8. [DOI] [PubMed] [Google Scholar]
  18. Isenberg G., Schubert P., Kreutzberg G. W. Experimental approach to test the role of actin in axonal transport. Brain Res. 1980 Aug 4;194(2):588–593. doi: 10.1016/0006-8993(80)91247-0. [DOI] [PubMed] [Google Scholar]
  19. KUNITZ M. Crystalline desoxyribonuclease; isolation and general properties; spectrophotometric method for the measurement of desoxyribonuclease activity. J Gen Physiol. 1950 Mar;33(4):349–362. doi: 10.1085/jgp.33.4.349. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Kamiya R., Maruyama K., Kuroda M., Kawamura M., Kikuchi M. Mg-polymer of actin formed under the influence of -actinin. Biochim Biophys Acta. 1972 Jan 21;256(1):120–131. doi: 10.1016/0005-2728(72)90167-3. [DOI] [PubMed] [Google Scholar]
  21. Karsenti E., Guilbert B., Bornens M., Avrameas S. Antibodies to tubulin in normal nonimmunized animals. Proc Natl Acad Sci U S A. 1977 Sep;74(9):3997–4001. doi: 10.1073/pnas.74.9.3997. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Lazarides E., Lindberg U. Actin is the naturally occurring inhibitor of deoxyribonuclease I. Proc Natl Acad Sci U S A. 1974 Dec;71(12):4742–4746. doi: 10.1073/pnas.71.12.4742. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Lin D. C., Tobin K. D., Grumet M., Lin S. Cytochalasins inhibit nuclei-induced actin polymerization by blocking filament elongation. J Cell Biol. 1980 Feb;84(2):455–460. doi: 10.1083/jcb.84.2.455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Lubit B. W., Schwartz J. H. An antiactin antibody that distinguishes between cytoplasmic and skeletal muscle actins. J Cell Biol. 1980 Sep;86(3):891–897. doi: 10.1083/jcb.86.3.891. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Maruyama K. A study of beta-actinin, myofibrillar protein from rabbit skeletal muscle. J Biochem. 1971 Feb;69(2):369–386. doi: 10.1093/oxfordjournals.jbchem.a129476. [DOI] [PubMed] [Google Scholar]
  26. Maruyama K., Kamiya R., Kimura S., Hatano S. Beta-actinin-like protein from plasmodium. J Biochem. 1976 Apr;79(4):709–715. doi: 10.1093/oxfordjournals.jbchem.a131122. [DOI] [PubMed] [Google Scholar]
  27. Muszbek L., Laki K. Cleavage of actin by thrombin. Proc Natl Acad Sci U S A. 1974 Jun;71(6):2208–2211. doi: 10.1073/pnas.71.6.2208. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Norberg R., Thorstensson R., Utter G., Fagraeus A. F-Actin-depolymerizing activity of human serum. Eur J Biochem. 1979 Oct 15;100(2):575–583. doi: 10.1111/j.1432-1033.1979.tb04204.x. [DOI] [PubMed] [Google Scholar]
  29. Pollard T. D., Weihing R. R. Actin and myosin and cell movement. CRC Crit Rev Biochem. 1974 Jan;2(1):1–65. doi: 10.3109/10409237409105443. [DOI] [PubMed] [Google Scholar]
  30. Price P. A., Liu T. Y., Stein W. H., Moore S. Properties of chromatographically purified bovine pancreatic deoxyribonuclease. J Biol Chem. 1969 Feb 10;244(3):917–923. [PubMed] [Google Scholar]
  31. Schwartz J. H. Axonal transport: components, mechanisms, and specificity. Annu Rev Neurosci. 1979;2:467–504. doi: 10.1146/annurev.ne.02.030179.002343. [DOI] [PubMed] [Google Scholar]
  32. Spudich J. A., Watt S. The regulation of rabbit skeletal muscle contraction. I. Biochemical studies of the interaction of the tropomyosin-troponin complex with actin and the proteolytic fragments of myosin. J Biol Chem. 1971 Aug 10;246(15):4866–4871. [PubMed] [Google Scholar]
  33. Treistman S. N., Schwartz J. H. Metabolism of acetylcholine in the nervous system of Aplysia californica. IV. Studies of an identified cholinergic axon. J Gen Physiol. 1977 Jun;69(6):725–741. doi: 10.1085/jgp.69.6.725. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Wang D., Moore S. Preparation of protease-free and ribonuclease-free pancreatic deoxyribonuclease. J Biol Chem. 1978 Oct 25;253(20):7216–7219. [PubMed] [Google Scholar]
  35. Wehland J., Osborn M., Weber K. Phalloidin-induced actin polymerization in the cytoplasm of cultured cells interferes with cell locomotion and growth. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5613–5617. doi: 10.1073/pnas.74.12.5613. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Wehland J., Stockem W., Weber K. Cytoplasmic streaming in Amoeba proteus is inhibited by the actin-specific drug phalloidin. Exp Cell Res. 1978 Sep;115(2):451–454. doi: 10.1016/0014-4827(78)90307-5. [DOI] [PubMed] [Google Scholar]
  37. Wehland J., Weber K., Gawlitta W., Stockem W. Effects of the actin-binding protein DNAase I on cytoplasmic streaming and ultrastructure of Amoeba proteus. An attempt to explain amoeboid movement. Cell Tissue Res. 1979 Jul 17;199(3):353–372. doi: 10.1007/BF00236075. [DOI] [PubMed] [Google Scholar]
  38. Wieland T. Modification of actins by phallotoxins. Naturwissenschaften. 1977 Jun;64(6):303–309. doi: 10.1007/BF00446784. [DOI] [PubMed] [Google Scholar]

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