Structuring a multi-nodal neural network in vitro within a novel design microfluidic chip

  • Rosanne van de Wijdeven
  • Ola Huse Ramstad
  • Ulrich Stefan Bauer
  • Øyvind Halaas
  • Axel Sandvig
  • Ioanna Sandvig


Neural network formation is a complex process involving axon outgrowth and guidance. Axon guidance is facilitated by structural and molecular cues from the surrounding microenvironment. Micro-fabrication techniques can be employed to produce microfluidic chips with a highly controlled microenvironment for neural cells enabling longitudinal studies of complex processes associated with network formation. In this work, we demonstrate a novel open microfluidic chip design that encompasses a freely variable number of nodes interconnected by axon-permissible tunnels, enabling structuring of multi-nodal neural networks in vitro. The chip employs a partially open design to allow high level of control and reproducibility of cell seeding, while reducing shear stress on the cells. We show that by culturing dorsal root ganglion cells (DRGs) in our microfluidic chip, we were able to structure a neural network in vitro. These neurons were compartmentalized within six nodes interconnected through axon growth tunnels. Furthermore, we demonstrate the additional benefit of open top design by establishing a 3D neural culture in matrigel and a neuronal aggregate 3D culture within the chips. In conclusion, our results demonstrate a novel microfluidic chip design applicable to structuring complex neural networks in vitro, thus providing a versatile, highly relevant platform for the study of neural network dynamics applicable to developmental and regenerative neuroscience.


Microfabrication Maskless aligner Lab-on-chip Axon growth and guidance 3D cell culture Organoid 



This work was supported by the Research Council of Norway, Norwegian Micro- and Nano-Fabrication Facility, NorFab, project number 245963/F50, NTNU program for Enabling Technologies, (Nanotechnology), and the Liaison Committee between the Central Norway Health Authority and NTNU (Samarbeidsorganet HMN-NTNU).

Compliance with ethical standards

All animal procedures were in accordance with the EU Directive 86/609/EEC and the Norwegian laws and regulations controlling procedures on experimental animals.

Conflict of interest

There are no conflicts to declare.


  1. M.W. Amoroso, G.F. Croft, D.J. Williams, S. O’Keeffe, M.A. Carrasco, A.R. Davis, et al., J Neurosci : Official J Soc Neurosci. 33, 2 (2013)CrossRefGoogle Scholar
  2. M.-C. Chuang, H.-Y. Lai, J.-A. Annie Ho, Y.-Y. Chen, Biosens. Bioelectron. 41 (2013)Google Scholar
  3. B. Deleglise, S. Magnifico, E. Duplus, P. Vaur, V. Soubeyre, M. Belle, et al., Acta Neuropathol. Commun. 2 (2014)Google Scholar
  4. J.-P. Dollé, B. Morrison, R.R. Schloss, M.L. Yarmush, Lab Chip 13, 3 (2013)CrossRefGoogle Scholar
  5. S. Hosmane, A. Fournier, R. Wright, L. Rajbhandari, R. Siddique, I.H. Yang, et al., Lab Chip 11, 22 (2011)CrossRefGoogle Scholar
  6. T.T. Kanagasabapathi, D. Ciliberti, S. Martinoia, W.J. Wadman, M.M.J. Decré, Front. Neuroeng. 4 (2011)Google Scholar
  7. H.K. Kleinman, G.R. Martin, Semin. Cancer Biol. 15, 5 (2005)CrossRefGoogle Scholar
  8. M.A. Lancaster, N.S. Corsini, S. Wolfinger, E.H. Gustafson, A.W. Phillips, T.R. Burkard, et al., Nat. Biotechnol. 35, 7 (2017)CrossRefGoogle Scholar
  9. H.U. Lee, S. Nag, A. Blasiak, Y. Jin, N. Thakor, I.H. Yang, ACS Chem. Neurosci. 7, 10 (2016)Google Scholar
  10. Y. Lei, J. Li, N. Wang, X. Yang, Y. Hamada, Q. Li, et al., Integr. Biol. 8, 3 (2016)CrossRefGoogle Scholar
  11. X. Lu, J.S. Kim-Han, K.L. O’Malley, S.E. Sakiyama-Elbert, J. Neurosci. Methods 209, 1 (2012)CrossRefGoogle Scholar
  12. J.W. Park, B. Vahidi, A.M. Taylor, S.W. Rhee, N.L. Jeon, Nat. Protoc. 1, 4 (2006)CrossRefGoogle Scholar
  13. J.-M. Peyrin, B. Deleglise, L. Saias, M. Vignes, P. Gougis, S. Magnifico, et al., Lab Chip 11, 21 (2011)CrossRefGoogle Scholar
  14. W.W. Poon, M. Blurton-Jones, C.H. Tu, L.M. Feinberg, M.A. Chabrier, J.W. Harris, et al., Neurobiol. Aging 32, 5 (2011)CrossRefGoogle Scholar
  15. S.K. Ravula, M.S. Wang, S.A. Asress, J.D. Glass, A. Bruno Frazier, J. Neurosci. Methods 159, 1 (2007)CrossRefGoogle Scholar
  16. R. Renault, N. Sukenik, S. Descroix, L. Malaquin, J.-L. Viovy, J.-M. Peyrin, et al., PLoS One 10, 4 (2015)Google Scholar
  17. O. Sporns, R. Kötter, PLoS Biol. 2, 11 (2004)CrossRefGoogle Scholar
  18. A.M. Taylor, S.W. Rhee, C.H. Tu, D.H. Cribbs, C.W. Cotman, N.L. Jeon, Langmuir: ACS J Surfaces Colloids 19, 5 (2003)Google Scholar
  19. A.M. Taylor, M. Blurton-Jones, S.W. Rhee, D.H. Cribbs, C.W. Cotman, N.L. Jeon, Nat. Methods 2, 8 (2005)CrossRefGoogle Scholar
  20. A.M. Taylor, S. Menon, S.L. Gupton, Lab Chip 15, 13 (2015)CrossRefGoogle Scholar
  21. M. Tessier-Lavigne, M. Placzek, A.G.S. Lumsden, J. Dodd, T.M. Jessell, Nature 336, 6201 (1988)CrossRefGoogle Scholar
  22. C. Tsantoulas, C. Farmer, P. Machado, K. Baba, S.B. McMahon, R. Raouf, PLoS One 8, 11 (2013)CrossRefGoogle Scholar
  23. B. Vahidi, J.W. Park, H.J. Kim, N.L. Jeon, J. Neurosci. Methods 170, 2 (2008)CrossRefGoogle Scholar
  24. Y. Wang, V. Balaji, S. Kaniyappan, L. Krüger, S. Irsen, K. Tepper, et al., Mol. Neurodegener. 12 (2017)Google Scholar
  25. G.M. Whitesides, Nature 442, 7101 (2006)CrossRefGoogle Scholar
  26. Y. Yu, M.H. Shamsi, D.L. Krastev, M.D.M. Dryden, Y. Leung, A.R. Wheeler, Lab Chip 16, 3 (2016)CrossRefGoogle Scholar
  27. E.E. Zahavi, A. Ionescu, S. Gluska, T. Gradus, K. Ben-Yaakov, E. Perlson, J. Cell Sci. 128, 6 (2015)CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2017

Authors and Affiliations

  • Rosanne van de Wijdeven
    • 1
  • Ola Huse Ramstad
    • 2
  • Ulrich Stefan Bauer
    • 2
  • Øyvind Halaas
    • 1
  • Axel Sandvig
    • 2
    • 3
    • 4
  • Ioanna Sandvig
    • 2
  1. 1.Department of Clinical and Molecular Medicine, Faculty of Medicine and Health SciencesNorwegian University of Science and TechnologyTrondheimNorway
  2. 2.Department of Neuromedicine and Movement Science, Faculty of Medicine and Health SciencesNorwegian University of Science and TechnologyTrondheimNorway
  3. 3.Department of Pharmacology and Clinical Neurosciences, Division of Neuro, Head and NeckUmeå University HospitalUmeåSweden
  4. 4.Department of Neurology and Clinical NeurophysiologySt Olavs HospitalTrondheimNorway

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