skip to main content
DOE PAGES title logo U.S. Department of Energy
Office of Scientific and Technical Information

Title: Controlled placement of multiple CNS cell populations to create complex neuronal cultures

Abstract

In vitro brain-on-a-chip platforms hold promise in many areas including: drug discovery, evaluating effects of toxicants and pathogens, and disease modelling. A more accurate recapitulation of the intricate organization of the brain in vivo may require a complex in vitro system including organization of multiple neuronal cell types in an anatomically-relevant manner. Most approaches for compartmentalizing or segregating multiple cell types on microfabricated substrates use either permanent physical surface features or chemical surface functionalization. This study describes a removable insert that successfully deposits neurons from different brain areas onto discrete regions of a microelectrode array (MEA) surface, achieving a separation distance of 100 μm. The regional seeding area on the substrate is significantly smaller than current platforms using comparable placement methods. The non-permanent barrier between cell populations allows the cells to remain localized and attach to the substrate while the insert is in place and interact with neighboring regions after removal. The insert was used to simultaneously seed primary rodent hippocampal and cortical neurons onto MEAs. These cells retained their morphology, viability, and function after seeding through the cell insert through 28 days in vitro (DIV). Co-cultures of the two neuron types developed processes and formed integrated networks between themore » different MEA regions. Electrophysiological data demonstrated characteristic bursting features and waveform shapes that were consistent for each neuron type in both mono- and co-culture. Additionally, hippocampal cells co-cultured with cortical neurons showed an increase in withinburst firing rate (p = 0.013) and percent spikes in bursts (p = 0.002), changes that imply communication exists between the two cell types in co-culture. The cell seeding insert described in this work is a simple but effective method of separating distinct neuronal populations on microfabricated devices, and offers a unique approach to developing the types of complex in vitro cellular environments required for anatomically-relevant brain-on-a-chip devices.« less

Authors:
ORCiD logo; ; ; ; ; ; ; ; ; ; ;
Publication Date:
Research Org.:
Lawrence Livermore National Lab. (LLNL), Livermore, CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC)
OSTI Identifier:
1409719
Alternate Identifier(s):
OSTI ID: 1627842
Grant/Contract Number:  
AC52-07NA27344
Resource Type:
Published Article
Journal Name:
PLoS ONE
Additional Journal Information:
Journal Name: PLoS ONE Journal Volume: 12 Journal Issue: 11; Journal ID: ISSN 1932-6203
Publisher:
Public Library of Science
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; Hippocampus; Neurons; Action potentials; Electrophysiology; Fluorescence imaging; Cell cultures; Electrode recording; Chemical deposition

Citation Formats

Soscia, D., Belle, A., Fischer, N., Enright, H., Sales, A., Osburn, J., Benett, W., Mukerjee, E., Kulp, K., Pannu, S., Wheeler, E., and Lytton, ed., William W.. Controlled placement of multiple CNS cell populations to create complex neuronal cultures. United States: N. p., 2017. Web. https://doi.org/10.1371/journal.pone.0188146.
Soscia, D., Belle, A., Fischer, N., Enright, H., Sales, A., Osburn, J., Benett, W., Mukerjee, E., Kulp, K., Pannu, S., Wheeler, E., & Lytton, ed., William W.. Controlled placement of multiple CNS cell populations to create complex neuronal cultures. United States. https://doi.org/10.1371/journal.pone.0188146
Soscia, D., Belle, A., Fischer, N., Enright, H., Sales, A., Osburn, J., Benett, W., Mukerjee, E., Kulp, K., Pannu, S., Wheeler, E., and Lytton, ed., William W.. Tue . "Controlled placement of multiple CNS cell populations to create complex neuronal cultures". United States. https://doi.org/10.1371/journal.pone.0188146.
@article{osti_1409719,
title = {Controlled placement of multiple CNS cell populations to create complex neuronal cultures},
author = {Soscia, D. and Belle, A. and Fischer, N. and Enright, H. and Sales, A. and Osburn, J. and Benett, W. and Mukerjee, E. and Kulp, K. and Pannu, S. and Wheeler, E. and Lytton, ed., William W.},
abstractNote = {In vitro brain-on-a-chip platforms hold promise in many areas including: drug discovery, evaluating effects of toxicants and pathogens, and disease modelling. A more accurate recapitulation of the intricate organization of the brain in vivo may require a complex in vitro system including organization of multiple neuronal cell types in an anatomically-relevant manner. Most approaches for compartmentalizing or segregating multiple cell types on microfabricated substrates use either permanent physical surface features or chemical surface functionalization. This study describes a removable insert that successfully deposits neurons from different brain areas onto discrete regions of a microelectrode array (MEA) surface, achieving a separation distance of 100 μm. The regional seeding area on the substrate is significantly smaller than current platforms using comparable placement methods. The non-permanent barrier between cell populations allows the cells to remain localized and attach to the substrate while the insert is in place and interact with neighboring regions after removal. The insert was used to simultaneously seed primary rodent hippocampal and cortical neurons onto MEAs. These cells retained their morphology, viability, and function after seeding through the cell insert through 28 days in vitro (DIV). Co-cultures of the two neuron types developed processes and formed integrated networks between the different MEA regions. Electrophysiological data demonstrated characteristic bursting features and waveform shapes that were consistent for each neuron type in both mono- and co-culture. Additionally, hippocampal cells co-cultured with cortical neurons showed an increase in withinburst firing rate (p = 0.013) and percent spikes in bursts (p = 0.002), changes that imply communication exists between the two cell types in co-culture. The cell seeding insert described in this work is a simple but effective method of separating distinct neuronal populations on microfabricated devices, and offers a unique approach to developing the types of complex in vitro cellular environments required for anatomically-relevant brain-on-a-chip devices.},
doi = {10.1371/journal.pone.0188146},
journal = {PLoS ONE},
number = 11,
volume = 12,
place = {United States},
year = {2017},
month = {11}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record
https://doi.org/10.1371/journal.pone.0188146

Citation Metrics:
Cited by: 3 works
Citation information provided by
Web of Science

Figures / Tables:

Fig 1 Fig 1: Microelectrode array design. (A) Completed device with well and connectors attached. (B) Cell-containing area of the device. The cell culture well is placed just inside the SU-8 alignment ring. (C) Brightfield micrograph of microelectrode region containing 60 electrodes. Dashed line denotes border between inner and outer region. Themore » inner region is further separated into three subregions, labeled A, B, and C. (D) Electrode detail and interstitial space between two cell placement areas. The metal used for visualization of borders between regions does not intersect any of the electrodes or traces.« less

Save / Share:

Works referenced in this record:

Dissociated cortical networks show spontaneously correlated activity patterns during in vitro development
journal, June 2006


Tracking axonal action potential propagation on a high-density microelectrode array across hundreds of sites
journal, July 2013

  • Bakkum, Douglas J.; Frey, Urs; Radivojevic, Milos
  • Nature Communications, Vol. 4, Issue 1
  • DOI: 10.1038/ncomms3181

Hippocampal-neocortical interaction: A hierarchy of associativity
journal, January 2000


Endogenous cholinergic tone modulates spontaneous network level neuronal activity in primary cortical cultures grown on multi-electrode arrays
journal, January 2013

  • Hammond, Mark W.; Xydas, Dimitris; Downes, Julia H.
  • BMC Neuroscience, Vol. 14, Issue 1
  • DOI: 10.1186/1471-2202-14-38

Long-term non-invasive interrogation of human dorsal root ganglion neuronal cultures on an integrated microfluidic multielectrode array platform
journal, January 2016

  • Enright, H. A.; Felix, S. H.; Fischer, N. O.
  • The Analyst, Vol. 141, Issue 18
  • DOI: 10.1039/C5AN01728A

Complex heterogeneous tissue constructs containing multiple cell types prepared by inkjet printing technology
journal, January 2013


Advances and Challenges in Recapitulating Human Pulmonary Systems: At the Cusp of Biology and Materials
journal, March 2016


Microfabricated elastomeric stencils for micropatterning cell cultures
journal, January 2000


Organs-on-chips: breaking the in vitro impasse
journal, January 2012

  • Meer, Andries D. van der; Berg, Albert van den
  • Integrative Biology, Vol. 4, Issue 5
  • DOI: 10.1039/c2ib00176d

Male-specific Volume Expansion of the Human Hippocampus during Adolescence
journal, July 2004


Perfused multiwell plate for 3D liver tissue engineering
journal, January 2010

  • Domansky, Karel; Inman, Walker; Serdy, James
  • Lab Chip, Vol. 10, Issue 1
  • DOI: 10.1039/B913221J

A ring barrier–based migration assay to assess cell migration in vitro
journal, May 2015

  • Das, Asha M.; Eggermont, Alexander M. M.; ten Hagen, Timo L. M.
  • Nature Protocols, Vol. 10, Issue 6
  • DOI: 10.1038/nprot.2015.056

Fabrication of transferable micropatterned-co-cultured cell sheets with microcontact printing
journal, October 2009


Building and manipulating neural pathways with microfluidics
journal, January 2010

  • Berdichevsky, Yevgeny; Staley, Kevin J.; Yarmush, Martin L.
  • Lab on a Chip, Vol. 10, Issue 8
  • DOI: 10.1039/b922365g

A microfluidic culture platform for CNS axonal injury, regeneration and transport
journal, July 2005

  • Taylor, Anne M.; Blurton-Jones, Mathew; Rhee, Seog Woo
  • Nature Methods, Vol. 2, Issue 8
  • DOI: 10.1038/nmeth777

Negative dielectrophoretic patterning with different cell types
journal, December 2008

  • Suzuki, Masato; Yasukawa, Tomoyuki; Shiku, Hitoshi
  • Biosensors and Bioelectronics, Vol. 24, Issue 4
  • DOI: 10.1016/j.bios.2008.06.051

Continuous monitoring of developmental activity changes in cultured cortical networks
journal, September 2003

  • Mukai, Yoshitaka; Shiina, Tsuyoshi; Jimbo, Yasuhiko
  • Electrical Engineering in Japan, Vol. 145, Issue 4
  • DOI: 10.1002/eej.10216

Directed cell growth on protein-functionalized hydrogel surfaces
journal, May 2007

  • Hynd, Matthew R.; Frampton, John P.; Dowell-Mesfin, Natalie
  • Journal of Neuroscience Methods, Vol. 162, Issue 1-2
  • DOI: 10.1016/j.jneumeth.2007.01.024

Patterning microscale extracellular matrices to study endothelial and cancer cell interactions in vitro
journal, January 2012

  • Dickinson, Laura E.; Lütgebaucks, Cornelis; Lewis, Daniel M.
  • Lab on a Chip, Vol. 12, Issue 21
  • DOI: 10.1039/c2lc40819h

Microengineering of Cellular Interactions
journal, August 2000


Role for a cortical input to hippocampal area CA1 in the consolidation of a long-term memory
journal, October 2004


Reconstituting Organ-Level Lung Functions on a Chip
journal, June 2010


Biomimetic tissues on a chip for drug discovery
journal, February 2012


The Role of Body-on-a-Chip Devices in Drug and Toxicity Studies
journal, August 2011


Controlling cell interactions by micropatterning in co-cultures: Hepatocytes and 3T3 fibroblasts
journal, February 1997


What is the brain?
journal, November 2000


The Spiking Component of Oscillatory Extracellular Potentials in the Rat Hippocampus
journal, August 2012


Ensembles of engineered cardiac tissues for physiological and pharmacological study: Heart on a chip
journal, January 2011

  • Grosberg, Anna; Alford, Patrick W.; McCain, Megan L.
  • Lab on a Chip, Vol. 11, Issue 24
  • DOI: 10.1039/c1lc20557a

3D printed nervous system on a chip
journal, January 2016

  • Johnson, Blake N.; Lancaster, Karen Z.; Hogue, Ian B.
  • Lab on a Chip, Vol. 16, Issue 8
  • DOI: 10.1039/C5LC01270H

Quantitative differences in developmental profiles of spontaneous activity in cortical and hippocampal cultures
journal, January 2015


Three-dimensional atlas system for mouse and rat brain imaging data
journal, January 2007


Differential Attention-Dependent Response Modulation across Cell Classes in Macaque Visual Area V4
journal, July 2007


Physiologically relevant organs on chips
journal, December 2013

  • Yum, Kyungsuk; Hong, Soon Gweon; Healy, Kevin E.
  • Biotechnology Journal, Vol. 9, Issue 1
  • DOI: 10.1002/biot.201300187

Rebuilding a realistic corticostriatal “social network” from dissociated cells
journal, April 2015

  • Garcia-Munoz, Marianela; Taillefer, Eddy; Pnini, Reuven
  • Frontiers in Systems Neuroscience, Vol. 9
  • DOI: 10.3389/fnsys.2015.00063

Primary support cultures of hippocampal and substantia nigra neurons
journal, December 2008


Neuronal Diversity and Temporal Dynamics: The Unity of Hippocampal Circuit Operations
journal, July 2008


Scaling and systems biology for integrating multiple organs-on-a-chip
journal, January 2013

  • Wikswo, John P.; Curtis, Erica L.; Eagleton, Zachary E.
  • Lab on a Chip, Vol. 13, Issue 18
  • DOI: 10.1039/c3lc50243k

Hippocampal networks on reliable patterned substrates
journal, January 2012

  • Boehler, Michael D.; Leondopulos, Stathis S.; Wheeler, Bruce C.
  • Journal of Neuroscience Methods, Vol. 203, Issue 2
  • DOI: 10.1016/j.jneumeth.2011.09.020

Characterization of a microfluidic in vitro model of the blood-brain barrier (μBBB)
journal, January 2012


    Works referencing / citing this record:

    Evaluation of in vitro neuronal platforms as surrogates for in vivo whole brain systems
    journal, July 2018


    A flexible 3-dimensional microelectrode array for in vitro brain models
    journal, January 2020

    • Soscia, David A.; Lam, Doris; Tooker, Angela C.
    • Lab on a Chip, Vol. 20, Issue 5
    • DOI: 10.1039/c9lc01148j

    A micro-fabricated in vitro complex neuronal circuit platform
    journal, June 2019

    • Kamudzandu, M.; Köse-Dunn, M.; Evans, M. G.
    • Biomedical Physics & Engineering Express, Vol. 5, Issue 4
    • DOI: 10.1088/2057-1976/ab2307

    Disease-modifying effects of metabolic perturbations in ALS/FTLD
    journal, December 2018

    • Jawaid, Ali; Khan, Romesa; Polymenidou, Magdalini
    • Molecular Neurodegeneration, Vol. 13, Issue 1
    • DOI: 10.1186/s13024-018-0294-0

    Organ-On-A-Chip in vitro Models of the Brain and the Blood-Brain Barrier and Their Value to Study the Microbiota-Gut-Brain Axis in Neurodegeneration
    journal, January 2020

    • Raimondi, Ilaria; Izzo, Luca; Tunesi, Marta
    • Frontiers in Bioengineering and Biotechnology, Vol. 7
    • DOI: 10.3389/fbioe.2019.00435

      Figures/Tables have been extracted from DOE-funded journal article accepted manuscripts.