A microfluidic platform for controlled biochemical stimulation of twin neuronal networks.

Spatially and temporally resolved delivery of soluble factors is a key feature for pharmacological applications. In this framework, microfluidics coupled to multisite electrophysiology offers great advantages in neuropharmacology and toxicology. In this work, a microfluidic device for biochemical stimulation of neuronal networks was developed. A micro-chamber for cell culturing, previously developed and tested for long term neuronal growth by our group, was provided with a thin wall, which partially divided the cell culture region in two sub-compartments. The device was reversibly coupled to a flat micro electrode array and used to culture primary neurons in the same microenvironment. We demonstrated that the two fluidically connected compartments were able to originate two parallel neuronal networks with similar electrophysiological activity but functionally independent. Furthermore, the device allowed to connect the outlet port to a syringe pump and to transform the static culture chamber in a perfused one. At 14 days invitro, sub-networks were independently stimulated with a test molecule, tetrodotoxin, a neurotoxin known to block action potentials, by means of continuous delivery. Electrical activity recordings proved the ability of the device configuration to selectively stimulate each neuronal network individually. The proposed microfluidic approach represents an innovative methodology to perform biological, pharmacological, and electrophysiological experiments on neuronal networks. Indeed, it allows for controlled delivery of substances to cells, and it overcomes the limitations due to standard drug stimulation techniques. Finally, the twin network configuration reduces biological variability, which has important outcomes on pharmacological and drug screening.

[1]  Yuzuru Takamura,et al.  Investigating neuronal activity with planar microelectrode arrays: achievements and new perspectives. , 2005, Journal of bioscience and bioengineering.

[2]  M. Baudry,et al.  Advances in Network Electrophysiology , 2006 .

[3]  Giancarlo Ferrigno,et al.  A Micro-Electrode Array device coupled to a laser-based system for the local stimulation of neurons by optical release of glutamate , 2008, Journal of Neuroscience Methods.

[4]  Giancarlo Ferrigno,et al.  PhotoMEA: An opto-electronic biosensor for monitoring in vitro neuronal network activity , 2007, Biosyst..

[5]  Wytse J. Wadman,et al.  Source (or Part of the following Source): Type Article Title Dual-compartment Neurofluidic System for Electrophysiological Measurements in Physically Segregated and Functionally Connected Neuronal Cell Culture Author(s) Neuroengineering Original Research Article Dual-compartment Neurofluidic System , 2022 .

[6]  H. Robinson,et al.  Simultaneous induction of pathway-specific potentiation and depression in networks of cortical neurons. , 1999, Biophysical journal.

[7]  A. Offenhäusser,et al.  Patterning chemical stimulation of reconstructed neuronal networks. , 2006, Analytica chimica acta.

[8]  Hiroyuki Fujita,et al.  Constraining the connectivity of neuronal networks cultured on microelectrode arrays with microfluidic techniques: a step towards neuron-based functional chips. , 2006, Biosensors & bioelectronics.

[9]  Matsuhiko Nishizawa,et al.  Localized chemical stimulation to micropatterned cells using multiple laminar fluid flows. , 2003, Lab on a chip.

[10]  Marco Rasponi,et al.  Validation of long‐term primary neuronal cultures and network activity through the integration of reversibly bonded microbioreactors and MEA substrates , 2012, Biotechnology and bioengineering.

[11]  Kuihuan Jian,et al.  Neuron adhesion and strengthening , 2010 .

[12]  Wenming Liu,et al.  Microfluidics: a new cosset for neurobiology. , 2009, Lab on a chip.

[13]  Alessandro Vato,et al.  Burst detection algorithms for the analysis of spatio-temporal patterns in cortical networks of neurons , 2005, Neurocomputing.

[14]  Luca Berdondini,et al.  Network Dynamics and Synchronous Activity in cultured Cortical Neurons , 2007, Int. J. Neural Syst..

[15]  A. Scherer,et al.  Applications of microfluidics for neuronal studies , 2007, Journal of the Neurological Sciences.

[16]  Michele Giugliano,et al.  Micropatterning neural cell cultures in 3D with a multi-layered scaffold. , 2011, Biomaterials.

[17]  Eshel Ben-Jacob,et al.  Towards neuro-memory-chip: imprinting multiple memories in cultured neural networks. , 2007, Physical review. E, Statistical, nonlinear, and soft matter physics.

[18]  R. Campenot,et al.  Local control of neurite development by nerve growth factor. , 1977, Proceedings of the National Academy of Sciences of the United States of America.

[19]  O. Orwar,et al.  Microfluidic gradient-generating device for pharmacological profiling. , 2005, Analytical chemistry.

[20]  G. Ferrigno,et al.  A new cross-correlation algorithm for the analysis of “in vitro” neuronal network activity aimed at pharmacological studies , 2011, Journal of Neuroscience Methods.

[21]  G. P. Moore,et al.  Neuronal spike trains and stochastic point processes. I. The single spike train. , 1967, Biophysical journal.

[22]  George M. Whitesides,et al.  Replica molding using polymeric materials: A practical step toward nanomanufacturing , 1997 .

[23]  A. Habets,et al.  Spontaneous neuronal firing patterns in fetal rat cortical networks during development in vitro: a quantitative analysis , 2004, Experimental Brain Research.

[24]  Marco Rasponi,et al.  Reliable magnetic reversible assembly of complex microfluidic devices: fabrication, characterization, and biological validation , 2011 .

[25]  G Shahaf,et al.  Learning in Networks of Cortical Neurons , 2001, The Journal of Neuroscience.

[26]  Daniel A. Wagenaar,et al.  The Neurally Controlled Animat: Biological Brains Acting with Simulated Bodies , 2001, Auton. Robots.

[27]  Alessandro Vato,et al.  Dissociated cortical networks show spontaneously correlated activity patterns during in vitro development , 2006, Brain Research.

[28]  G. Gross.  Simultaneous Single Unit Recording in vitro with a Photoetched Laser Deinsulated Gold Multimicroelectrode Surface , 1979, IEEE Transactions on Biomedical Engineering.

[29]  J. Pine.  Recording action potentials from cultured neurons with extracellular microcircuit electrodes , 1980, Journal of Neuroscience Methods.

[30]  Surendra K. Ravula,et al.  Spatiotemporal localization of injury potentials in DRG neurons during vincristine-induced axonal degeneration , 2007, Neuroscience Letters.

[31]  Deyu Li,et al.  A versatile valve-enabled microfluidic cell co-culture platform and demonstration of its applications to neurobiology and cancer biology , 2011, Biomedical microdevices.

[32]  Roger D Kamm,et al.  A high-throughput microfluidic assay to study neurite response to growth factor gradients. , 2011, Lab on a chip.

[33]  Nitish Thakor,et al.  Circular compartmentalized microfluidic platform: Study of axon-glia interactions. , 2010, Lab on a chip.

[34]  Deyu Li,et al.  Co-culture of neurons and glia in a novel microfluidic platform , 2011, Journal of Neuroscience Methods.

[35]  Ran Ginosar,et al.  An Integrated System for Multichannel Neuronal Recording With Spike/LFP Separation, Integrated A/D Conversion and Threshold Detection , 2007, IEEE Trans. Biomed. Eng..

[36]  S. Takayama,et al.  Arrays of horizontally-oriented mini-reservoirs generate steady microfluidic flows for continuous perfusion cell culture and gradient generation. , 2004, The Analyst.

[37]  Thomas M Pearce,et al.  Integrated microelectrode array and microfluidics for temperature clamp of sensory neurons in culture. , 2005, Lab on a chip.

[38]  B R Ringeisen,et al.  Biochip∕laser cell deposition system to assess polarized axonal growth from single neurons and neuron∕glia pairs in microchannels with novel asymmetrical geometries. , 2011, Biomicrofluidics.

[39]  Teruo Fujii,et al.  Characterization of a microfluidic dispensing system for localised stimulation of cellular networks. , 2006, Lab on a chip.

[40]  Shaoqun Zeng,et al.  Characterization of synchronized bursts in cultured hippocampal neuronal networks with learning training on microelectrode arrays. , 2007, Biosensors & bioelectronics.

[41]  Bruce C Wheeler,et al.  Added astroglia promote greater synapse density and higher activity in neuronal networks. , 2007, Neuron glia biology.

[42]  Hynek Wichterle,et al.  Combined microfluidics/protein patterning platform for pharmacological interrogation of axon pathfinding. , 2010, Lab on a chip.

[43]  Erin M. Schuman,et al.  Microfluidic Local Perfusion Chambers for the Visualization and Manipulation of Synapses , 2010, Neuron.

[44]  Yoonkey Nam,et al.  Active 3-D microscaffold system with fluid perfusion for culturing in vitro neuronal networks. , 2007, Lab on a chip.

[45]  Anja Kunze,et al.  Co-pathological connected primary neurons in a microfluidic device for Alzheimer studies. , 2011, Biotechnology and bioengineering.

[46]  Noo Li Jeon,et al.  Micro-scale and microfluidic devices for neurobiology , 2010, Current Opinion in Neurobiology.

[47]  Arum Han,et al.  Microfluidic compartmentalized co-culture platform for CNS axon myelination research , 2009, Biomedical microdevices.

[48]  Andrew F M Johnstone,et al.  Microelectrode arrays: a physiologically based neurotoxicity testing platform for the 21st century. , 2010, Neurotoxicology.

[49]  Bruce C Wheeler,et al.  Novel MEA platform with PDMS microtunnels enables the detection of action potential propagation from isolated axons in culture. , 2009, Lab on a chip.

[50]  Jong Hwan Sung,et al.  A microfluidic device for a pharmacokinetic-pharmacodynamic (PK-PD) model on a chip. , 2010, Lab on a chip.

[51]  Wim L. C. Rutten,et al.  Long-term characterization of firing dynamics of spontaneous bursts in cultured neural networks , 2004, IEEE Transactions on Biomedical Engineering.

[52]  Thomas M Pearce,et al.  Microtechnology: meet neurobiology. , 2007, Lab on a chip.

[53]  M. Chiappalone,et al.  Networks of neurons coupled to microelectrode arrays: a neuronal sensory system for pharmacological applications. , 2003, Biosensors & bioelectronics.