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Axon Myelination and Electrical Stimulation in a Microfluidic, Compartmentalized Cell Culture Platform

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Abstract

Axon demyelination contributes to the loss of sensory and motor function following injury or disease in the central nervous system. Numerous reports have demonstrated that myelination can be achieved in neuron/oligodendrocyte co-cultures. However, the ability to selectively treat neuron or oligodendrocyte (OL) cell bodies in co-cultures improves the value of these systems when designing mechanism-based therapeutics. We have developed a microfluidic-based compartmentalized culture system to achieve segregation of neuron and OL cell bodies while simultaneously allowing the formation of myelin sheaths. Our microfluidic platform allows for a high replicate number, minimal leakage, and high flexibility. Using a custom built lid, fit with platinum electrodes for electrical stimulation (10-Hz pulses at a constant 3 V with ~190 kΩ impedance), we employed the microfluidic platform to achieve activity-dependent myelin segment formation. Electrical stimulation of dorsal root ganglia resulted in a fivefold increase in the number of myelinated segments/mm2 when compared to unstimulated controls (19.6 ± 3.0 vs. 3.6 ± 2.3 MBP+ segments/mm2). This work describes the modification of a microfluidic, multi-chamber system so that electrical stimulation can be used to achieve increased levels of myelination while maintaining control of the cell culture microenvironment.

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References

  • Balkowiec, A., & Katz, D. M. (2000). Activity-dependent release of endogenous brain-derived neurotrophic factor from primary sensory neurons detected by ELISA in situ. Journal of Neuroscience, 20(19), 7417–7423.

    PubMed  CAS  Google Scholar 

  • Barres, B. A., Jacobson, M. D., Schmid, R., Sendtner, M., & Raff, M. C. (1993). Does oligodendrocyte survival depend on axons? Current Biology, 3(8), 489–497. doi:10.0960-9822(93)90039-Q.

    Article  PubMed  CAS  Google Scholar 

  • Barres, B. A., & Raff, M. C. (1993). Proliferation of oligodendrocyte precursor cells depends on electrical activity in axons. Nature, 361(6409), 258–260. doi:10.1038/361258a0.

    Article  PubMed  CAS  Google Scholar 

  • Becker, D., Gary, D. S., Rosenzweig, E. S., Grill, W. M., & McDonald, J. W. (2010). Functional electrical stimulation helps replenish progenitor cells in the injured spinal cord of adult rats. Experimental Neurology, 222(2), 211–218. doi:10.1016/j.expneurol.2009.12.029.

    Article  PubMed  Google Scholar 

  • Bozzali, M., & Wrabetz, L. (2004). Axonal signals and oligodendrocyte differentiation. Neurochemical Research, 29(5), 979–988.

    Article  PubMed  CAS  Google Scholar 

  • Campenot, R. B. (1977). Local control of neurite development by nerve growth factor. Proceedings of the National Academy of Sciences of the United States of America, 74(10), 4516–4519.

    Article  PubMed  CAS  Google Scholar 

  • Campenot, R. B. (1981). Regeneration of neurites on long-term cultures of sympathetic neurons deprived of nerve growth factor. Science, 214(4520), 579–581.

    Article  PubMed  CAS  Google Scholar 

  • Campenot, R. B. (1982a). Development of sympathetic neurons in compartmentalized cultures. I. Local control of neurite growth by nerve growth factor. Developmental Biology, 93(1), 1–12.

    Article  PubMed  CAS  Google Scholar 

  • Campenot, R. B. (1982b). Development of sympathetic neurons in compartmentalized cultures. II. Local control of neurite survival by nerve growth factor. Developmental Biology, 93(1), 13–21. doi:10.0012-1606(82)90233-0.

    Article  PubMed  CAS  Google Scholar 

  • Chen, Y., Balasubramaniyan, V., Peng, J., Hurlock, E. C., Tallquist, M., Li, J., et al. (2007). Isolation and culture of rat and mouse oligodendrocyte precursor cells. Nature Protocols, 2(5), 1044–1051. doi:10.1038/nprot.2007.149.

    Article  PubMed  CAS  Google Scholar 

  • Claude, P., Hawrot, E., Dunis, D. A., & Campenot, R. B. (1982). Binding, internalization, and retrograde transport of 125I-nerve growth factor in cultured rat sympathetic neurons. Journal of Neuroscience, 2(4), 431–442.

    PubMed  CAS  Google Scholar 

  • Demerens, C., Stankoff, B., Logak, M., Anglade, P., Allinquant, B., Couraud, F., et al. (1996). Induction of myelination in the central nervous system by electrical activity. Proceedings of the National Academy of Sciences of the United States of America, 93(18), 9887–9892.

    Article  PubMed  CAS  Google Scholar 

  • Gong, S., Zheng, C., Doughty, M. L., Losos, K., Didkovsky, N., Schambra, U. B., et al. (2003). A gene expression atlas of the central nervous system based on bacterial artificial chromosomes. Nature, 425(6961), 917–925. doi:10.1038/nature02033nature02033.

    Article  PubMed  CAS  Google Scholar 

  • Gyllensten, L., & Malmfors, T. (1963). Myelinization of the optic nerve and its dependence on visual function—a quantitative investigation in mice. Journal of Embryology and Experimental Morphology, 11, 255–266.

    PubMed  CAS  Google Scholar 

  • Hosmane, S., Yang, I. H., Ruffin, A., Thakor, N., & Venkatesan, A. (2010). Circular compartmentalized microfluidic platform: Study of axon-glia interactions. Lab Chip, 10(6), 741–747. doi:10.1039/b918640a.

    Article  PubMed  CAS  Google Scholar 

  • Hur, E. M., Yang, I. H., Kim, D. H., Byun, J., Saijilafu, Xu, W. L., et al. (2011). Engineering neuronal growth cones to promote axon regeneration over inhibitory molecules. Proceedings of the National Academy of Sciences of the United States of America, 108(12), 5057–5062. doi:10.1073/pnas.1011258108.

    Article  PubMed  CAS  Google Scholar 

  • Ishibashi, T., Dakin, K. A., Stevens, B., Lee, P. R., Kozlov, S. V., Stewart, C. L., et al. (2006). Astrocytes promote myelination in response to electrical impulses. Neuron, 49(6), 823–832. doi:10.1016/j.neuron.2006.02.006.

    Article  PubMed  CAS  Google Scholar 

  • Kimpinski, K., Campenot, R. B., & Mearow, K. (1997). Effects of the neurotrophins nerve growth factor, neurotrophin-3, and brain-derived neurotrophic factor (BDNF) on neurite growth from adult sensory neurons in compartmented cultures. Journal of Neurobiology, 33(4), 395–410. doi:10.1002/(SICI)1097-4695(199710)33:4<395:AID-NEU5>3.0.CO;2-5.

    Article  PubMed  CAS  Google Scholar 

  • Li, Q., Brus-Ramer, M., Martin, J. H., & McDonald, J. W. (2010). Electrical stimulation of the medullary pyramid promotes proliferation and differentiation of oligodendrocyte progenitor cells in the corticospinal tract of the adult rat. Neuroscience Letters, 479(2), 128–133. doi:10.1016/j.neulet.2010.05.043.

    Article  PubMed  CAS  Google Scholar 

  • Mehta, N. R., Lopez, P. H., Vyas, A. A., & Schnaar, R. L. (2007). Gangliosides and Nogo receptors independently mediate myelin-associated glycoprotein inhibition of neurite outgrowth in different nerve cells. Journal of Biological Chemistry, 282(38), 27875–27886. doi:10.1074/jbc.M704055200.

    Article  PubMed  CAS  Google Scholar 

  • Millet, L. J., Stewart, M. E., Sweedler, J. V., Nuzzo, R. G., & Gillette, M. U. (2007). Microfluidic devices for culturing primary mammalian neurons at low densities. Lab Chip, 7(8), 987–994. doi:10.1039/b705266a.

    Article  PubMed  CAS  Google Scholar 

  • Omlin, F. X. (1997). Optic disc and optic nerve of the blind cape mole-rat (Georychus capensis): A proposed model for naturally occurring reactive gliosis. Brain Research Bulletin, 44(5), 627–632.

    Article  PubMed  CAS  Google Scholar 

  • Park, J., Koito, H., Li, J., & Han, A. (2009). Microfluidic compartmentalized co-culture platform for CNS axon myelination research. Biomedical Microdevices, 11(6), 1145–1153. doi:10.1007/s10544-009-9331-7.

    Article  PubMed  CAS  Google Scholar 

  • Riccio, A., Pierchala, B. A., Ciarallo, C. L., & Ginty, D. D. (1997). An NGF-TrkA-mediated retrograde signal to transcription factor CREB in sympathetic neurons. Science, 277(5329), 1097–1100.

    Article  PubMed  CAS  Google Scholar 

  • Sadowsky, C. L., & McDonald, J. W. (2009). Activity-based restorative therapies: Concepts and applications in spinal cord injury-related neurorehabilitation. Developmental Disabilities Research Reviews, 15(2), 112–116. doi:10.1002/ddrr.61.

    Article  PubMed  Google Scholar 

  • Senger, D. L., & Campenot, R. B. (1997). Rapid retrograde tyrosine phosphorylation of trkA and other proteins in rat sympathetic neurons in compartmented cultures. Journal of Cell Biology, 138(2), 411–421.

    Article  PubMed  CAS  Google Scholar 

  • Stevens, B., & Fields, R. D. (2000). Response of Schwann cells to action potentials in development. Science, 287(5461), 2267–2271.

    Article  PubMed  CAS  Google Scholar 

  • Stevens, B., Porta, S., Haak, L. L., Gallo, V., & Fields, R. D. (2002). Adenosine: A neuron-glial transmitter promoting myelination in the CNS in response to action potentials. Neuron, 36(5), 855–868.

    Article  PubMed  CAS  Google Scholar 

  • Steward, O., & Worley, P. F. (2001). Selective targeting of newly synthesized Arc mRNA to active synapses requires NMDA receptor activation. Neuron, 30(1), 227–240.

    Article  PubMed  CAS  Google Scholar 

  • Tauber, H., Waehneldt, T. V., & Neuhoff, V. (1980). Myelination in rabbit optic nerves is accelerated by artificial eye opening. Neuroscience Letters, 16(3), 235–238.

    Article  PubMed  CAS  Google Scholar 

  • Taylor, A. M., Blurton-Jones, M., Rhee, S. W., Cribbs, D. H., Cotman, C. W., & Jeon, N. L. (2005). A microfluidic culture platform for CNS axonal injury, regeneration and transport. Nature Methods, 2(8), 599–605. doi:10.1038/nmeth777.

    Article  PubMed  CAS  Google Scholar 

  • Tsui-Pierchala, B. A., & Ginty, D. D. (1999). Characterization of an NGF-P-TrkA retrograde-signaling complex and age-dependent regulation of TrkA phosphorylation in sympathetic neurons. Journal of Neuroscience, 19(19), 8207–8218.

    PubMed  CAS  Google Scholar 

  • Ure, D. R., & Campenot, R. B. (1997). Retrograde transport and steady-state distribution of 125I-nerve growth factor in rat sympathetic neurons in compartmented cultures. Journal of Neuroscience, 17(4), 1282–1290.

    PubMed  CAS  Google Scholar 

  • Vazdarjanova, A., Ramirez-Amaya, V., Insel, N., Plummer, T. K., Rosi, S., Chowdhury, S., et al. (2006). Spatial exploration induces ARC, a plasticity-related immediate-early gene, only in calcium/calmodulin-dependent protein kinase II-positive principal excitatory and inhibitory neurons of the rat forebrain. Journal of Comparative Neurology, 498(3), 317–329. doi:10.1002/cne.21003.

    Article  PubMed  CAS  Google Scholar 

  • Wake, H., Lee, P. R., & Fields, R. D. (2011). Control of local protein synthesis and initial events in myelination by action potentials. Science, 333(6049), 1647–1651. doi:10.1126/science.1206998.

    Article  PubMed  CAS  Google Scholar 

  • Yang, I. H., Siddique, R., Hosmane, S., Thakor, N., & Hoke, A. (2009). Compartmentalized microfluidic culture platform to study mechanism of paclitaxel-induced axonal degeneration. Experimental Neurology, 218(1), 124–128. doi:10.1016/j.expneurol.2009.04.017.

    Article  PubMed  CAS  Google Scholar 

  • Zalc, B., & Fields, R. D. (2000). Do action potentials regulate myelination? Neuroscientist, 6(1), 5–13. doi:10.1177/107385840000600109.

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

This work was funded by US Department of Defense USAMRMC/TATRC/USAMRAA contracts W81XWH-08-2-0192, W81XWH-09-2-0186, W81XWH-10-BCRP-IDEA, and Maryland Stem Cell Research Fund.

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Authors and Affiliations

  1. Department of Biomedical Engineering, School of Medicine, Johns Hopkins University, Baltimore, MD, USA

    In Hong Yang, Misti Malone, Stephen Dria & Nitish Thakor

  2. The International Center for Spinal Cord Injury, Hugo Moser Research Institute, Kennedy Krieger Institute, Baltimore, MD, USA

    Devin Gary, Misti Malone, Thierry Houdayer, Visar Belegu & John W. Mcdonald

  3. Department of Neurology, School of Medicine, Johns Hopkins University, Baltimore, MD, USA

    Devin Gary, Visar Belegu & John W. Mcdonald

  4. Department of Physical Medicine and Rehabilitation, School of Medicine, Johns Hopkins University, Baltimore, MD, USA

    John W. Mcdonald

  5. SiNAPSE, National University of Singapore, Singapore, Singapore

    In Hong Yang & Nitish Thakor

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  1. In Hong Yang
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Corresponding authors

Correspondence to John W. Mcdonald or Nitish Thakor.

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In Hong Yang and Devin Gary contributed equally to the work.

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Yang, I.H., Gary, D., Malone, M. et al. Axon Myelination and Electrical Stimulation in a Microfluidic, Compartmentalized Cell Culture Platform. Neuromol Med 14, 112–118 (2012). https://doi.org/10.1007/s12017-012-8170-5

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  • DOI: https://doi.org/10.1007/s12017-012-8170-5

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