Cellular and Molecular Neurobiology

, Volume 27, Issue 6, pp 701–716 | Cite as

Axon Viability and Mitochondrial Function are Dependent on Local Protein Synthesis in Sympathetic Neurons

  • Mi Hillefors
  • Anthony E. Gioio
  • Marie G. Mameza
  • Barry B. Kaplan
Original Paper


(1) Axons contain numerous mRNAs and a local protein synthetic system that can be regulated independently of the cell body. (2) In this study, cultured primary sympathetic neurons were employed, to assess the effect of local protein synthesis blockade on axon viability and mitochondrial function. (3) Inhibition of local protein synthesis reduced newly synthesized axonal proteins by 65% and resulted in axon retraction after 6 h. Acute inhibition of local protein synthesis also resulted in a significant decrease in the membrane potential of axonal mitochondria. Likewise, blockade of local protein transport into the mitochondria by transfection of the axons with Hsp90 C-terminal domain decreased the mitochondrial membrane potential by 65%. Moreover, inhibition of the local protein synthetic system also reduced the ability of mitochondria to restore axonal levels of ATP after KCl-induced depolarization. (4) Taken together, these results indicate that the local protein synthetic system plays an important role in mitochondrial function and the maintenance of the axon.


Superior cervical ganglia Axon Local protein synthesis Mitochondrial membrane potential ATP levels 



We thank Dr. J. Young (McGill Faculty of Medicine, Montreal, Quebec, Canada) for providing the C90 plasmid.


  1. Anderson S, Bankier AT, Barrell BG, de Bruijn MHL, Coulson AR, Drouin J, Eperon IC, Nierlich DP, Roe BA, Sanger F, Schreier PH, Smith AJH, Staden R, Young IG (1981) Sequence and organization of the human mitochondrial genome. Nature 290:457–465PubMedCrossRefGoogle Scholar
  2. Attardi G, Schatz G (1988) Biogenesis of mitochondria. Annu Rev Cell Biol 4:289–331PubMedCrossRefGoogle Scholar
  3. Bauer MF, Hofmann S (2006) Import of mitochondrial proteins. In: Schapira AHV (ed) Mitochondrial function and dysfunction. Academic Press, San Diego, CA, pp 57–90Google Scholar
  4. Beaumont V, Zhong N, Fletcher R, Froemke RC, Zucker RS (2001) Phosphorylation and local presynaptic protein synthesis in calcium- and calcineurin-dependent induction of crayfish long-term facilitation. Neuron 32:489–501PubMedCrossRefGoogle Scholar
  5. Bleher R, Martin R (2001) Ribosomes in the squid giant axon. Neuroscience 107:527–534PubMedCrossRefGoogle Scholar
  6. Brittis PA, Lu Q, Flanagan JG (2002) Axonal protein synthesis provides a mechanism for localized regulation at an intermediate target. Cell 110:223–235PubMedCrossRefGoogle Scholar
  7. Campbell DS, Holt CE (2001) Chemotropic responses of retinal growth cones mediated by rapid local protein synthesis and degradation. Neuron 32:1013–1026PubMedCrossRefGoogle Scholar
  8. Campenot RB, Martin G (2001) Construction and use of compartmented cultures for studies of cell biology in neurons. In: Federoff S, Richadrson A (ed) Protocols for neural cell culture. Humana Press, Inc., Totowa, NJ, pp 49–57CrossRefGoogle Scholar
  9. Casadio A, Martin KC, Giustetto M, Zhu H, Chen M, Bartsch D, Bailey CH, Kandel ER (1999) A transient, neuron-wide form of CREB-mediated long-term facilitation can be stabilized at specific synapses by local protein synthesis. Cell 99:221–237PubMedCrossRefGoogle Scholar
  10. Cossarizza A, Baccaranicontri M, Kalashnikova G, Franceschi C (1993) A new method for the cytofluorometric analysis of mitochondrial membrane potential using the J-aggregate forming lipophilic cation 5,5′,6,6′-tetrachloro-1,1′,3,3′-tetraethylbenzimidazolcarbocyanine iodide (JC-1). Biochem Biophys Res Commun 197:40–45PubMedCrossRefGoogle Scholar
  11. Ehrenberg B, Montana V, Wei MD, Wuskell JP, Loew LM (1988) Membrane potential can be determined in individual cells from the nernstian distribution of cationic dyes. Biophys J 53:785–794PubMedCrossRefGoogle Scholar
  12. Eng H, Lund K, Campenot RB (1999) Synthesis of beta-tubulin, actin, and other proteins in axons of sympathetic neurons in compartmented cultures. J Neurosci 19:1–9PubMedGoogle Scholar
  13. Gioio AE, Eyman M, Zhang H, Lavina ZS, Giuditta A, Kaplan BB (2001) Local synthesis of nuclear-encoded mitochondrial proteins in the presynaptic nerve terminal. J Neurosci Res 64:447–453PubMedCrossRefGoogle Scholar
  14. Gioio AE, Lavina ZS, Jurkovicova D, Zhang H, Eyman M, Giuditta A, Kaplan BB (2004) Nerve terminals of squid photoreceptor neurons contain a heterogeneous population of mRNAs and translate a transfected reporter mRNA. Eur J Neurosci 20:865–872PubMedCrossRefGoogle Scholar
  15. Giuditta A, Kaplan BB, van Minnen J, Alvarez J, Koenig E (2002) Axonal and presynaptic protein synthesis: new insights into the biology of the neuron. Trends Neurosci 25:400–404PubMedCrossRefGoogle Scholar
  16. Giuditta A, Menichini E, Perrone Capano C, Langella M, Martin R, Castigli E, Kaplan BB (1991) Active polysomes in the axoplasm of the squid giant axon. J Neurosci Res 28:18–28PubMedCrossRefGoogle Scholar
  17. Hanz S, Perlson E, Willis D, Zheng JQ, Massarwa R, Huerta JJ, Koltzenburg M, Kohler M, van-Minnen J, Twiss JL, Fainzilber M (2003) Axoplasmic importins enable retrograde injury signaling in lesioned nerve. Neuron 40:1095–1104PubMedCrossRefGoogle Scholar
  18. Heales SJR, Gegg ME, Clark JB (2006) Oxidative phosporylation: structure, function, and intermediary metabolism. In: Schapira AHV (ed) Mitochondria function and dysfuntion. Academic Press, San Diego, CA, pp 25–56Google Scholar
  19. Hollenbeck PJ, Saxton WM (2005) The axonal transport of mitochondria. J Cell Sci 118:5411–5419PubMedCrossRefGoogle Scholar
  20. Johnson MI (2001) Primary cultures of sympathetic ganglia. In: Federoff S, Richadrson A (eds) Protocols for neural cell culture. Humana Press, Inc., Totowa, NJ, pp 71–94CrossRefGoogle Scholar
  21. Kaplan BB, Gioio AE, Perrone Capano C, Crispino M, Giuditta A (1992) β-Actin and β-Tubulin are components of a heterogeneous mRNA population present in the squid giant axon. Mol Cell Neurosci 3:133–144CrossRefGoogle Scholar
  22. Lee SK, Hollenbeck PJ (2003) Organization and translation of mRNA in sympathetic axons. J Cell Sci 116:4467–4478PubMedCrossRefGoogle Scholar
  23. Li C, Sasaki Y, Takei K, Yamamoto H, Shouji M, Sugiyama Y, Kawakami T, Nakamura F, Yagi T, Ohshima T, Goshima Y (2004) Correlation between Semaphorin3A-induced facilitation of axonal transport and local activation of a translation initiation factor eukaryotic translation initiation factor 4E. J Neurosci 24:6161–6170PubMedCrossRefGoogle Scholar
  24. Li J-Y, Volknandt W, Dahlstrom A, Herrmann C, Blasi J, Das B, Zimmermann H (1999) Axonal transport of ribonucleoprotein particles (Vaults). Neuroscience 91:1055–1065PubMedCrossRefGoogle Scholar
  25. Liu K, Hu J-Y, Wang D, Schacher S (2003) Protein synthesis at synapse versus cell body: enhanced but transient expression of long-term facilitation at isolated synapses. J Neurobiol 56:275–286PubMedCrossRefGoogle Scholar
  26. Liu S, Wong-Riley M (1994) Nuclear-encoded mitochondrial precursor protein: intramitochondrial delivery to dendrites and axon terminals of neurons and regulation by neuronal activity. J Neurosci 14:5338–5351PubMedGoogle Scholar
  27. Margeot A, Garcia M, Wang W, Tetaud E, di Rago JP, Jacq C (2005) Why are many mRNAs translated to the vicinity of mitochondria: a role in protein complex assembly? Gene 354:64–71PubMedCrossRefGoogle Scholar
  28. Martin KC, Barad M, Kandel ER (2000) Local protein synthesis and its role in synapse-specific plasticity. Curr Opin Neurobiol 10:587–592PubMedCrossRefGoogle Scholar
  29. Martin KC, Casadio A, Zhu H, E Y, Rose JC, Chen M, Bailey CH, Kandel ER (1997) Synapse-specific, long-term facilitation of Aplysia sensory to motor synapses: a function for local protein synthesis in memory storage. Cell 91:927–938PubMedCrossRefGoogle Scholar
  30. Ming GL, Wong ST, Henley J, Yuan XB, Song HJ, Spitzer NC, Poo M (2002) Adaptation in the chemotactic guidance of nerve growth cones. Nature 417:411–418PubMedCrossRefGoogle Scholar
  31. Moccia R, Chen D, Lyles V, Kapuya E, E Y, Kalachikov S, Spahn CMT, Frank J, Kandel ER, Barad M, Martin KC (2003) An unbiased cDNA library prepared from isolated Aplysia sensory neuron processes is enriched for cytoskeletal and translational mRNAs. J Neurosci 23:9409–9417PubMedGoogle Scholar
  32. Olink-Coux M, Hollenbeck PJ (1996) Localization and active transport of mRNA in axons of sympathetic neurons in culture. J Neurosci 16:1346–1358PubMedGoogle Scholar
  33. Perrone Capano C, Giuditta A, Castigli E, Kaplan BB (1987) Occurrence and sequence complexity of polyadenylated RNA in squid axoplasm. J Neurochem 49:698–704CrossRefGoogle Scholar
  34. Piper M, Holt C (2004) RNA translations in axons. Annu Rev Cell Dev Biol 20:505–523PubMedCrossRefGoogle Scholar
  35. Schacher S, Wu F (2002) Synapse formation in the absence of cell bodies requires protein synthesis. J Neurosci 22:1831–1839PubMedGoogle Scholar
  36. Schuman EM, Dynes JL, Steward O (2006) Synaptic regulation of translation of dendritic mRNAs. J Neurosci 26:7143–7146PubMedCrossRefGoogle Scholar
  37. Si K, Giustetto M, Etkin A, Hsu R, Janisiewicz AM, Miniaci MC, Kim JH, Zhu H, Kandel ER (2003) A neuronal isoform of CPEB regulates local protein synthesis and stabilizes synapse-specific long-term facilitation in Aplysia. Cell 115:893–904PubMedCrossRefGoogle Scholar
  38. Truscott KN, Brandner K, Pfanner N (2003) Mechanisms of protein import into mitochondria. Curr Biol 13:R326–R337PubMedCrossRefGoogle Scholar
  39. Young JC, Hoogenraad NJ, Hartl FU (2003) Molecular chaperones Hsp90 and Hsp70 deliver preproteins to the mitochondrial import receptor Tom70. Cell 112:41–50PubMedCrossRefGoogle Scholar
  40. Young JC, Obermann WM, Hartl FU (1998) Specific binding of tetratricopeptide repeat proteins to the C-terminal 12-kDa domain of Hsp90. J Biol Chem 273:18007–18010PubMedCrossRefGoogle Scholar
  41. Zhang X, Poo M (2002) Localized synaptic potentiation by BDNF requires local protein synthesis in the developing axon. Neuron 36:675–688PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2007

Authors and Affiliations

  • Mi Hillefors
    • 1
  • Anthony E. Gioio
    • 1
  • Marie G. Mameza
    • 1
  • Barry B. Kaplan
    • 1
  1. 1.Laboratory of Molecular Biology, National Institute of Mental HealthNational Institutes of HealthBethesdaUSA

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