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CONTENTS
Impedance spectroscopy
Its need and uses
Experimental set up
Circuit model for
Bare electrodes
Cell coated electrodes

Results
Conclusion
References

What is Impedance spectroscopy
EIS measures the dielectric properties of the medium as a function of frequency.
It is based on the interaction of electric field with the electric dipole moment of the sample.
This technique measures the frequency response of the system.

NEED FOR EIS
Chronic integration of microfabricated neuroprosthetic devices is currently limited by the development of reactive cell and tissue responses after device implantation
Few tools are available that permit realtime assessment of reactive response development and the impact on device electrical performance.
EIS provides a method for the real time acquisition of electrical data that can be used to evaluate the composition and structure of neural tissue
EIS provides an impedance based system for modelling the electrical properties of 3-D neural tissue cultures surrounding neuroprosthetic devices
Mpreover Impedance measurements can be made over time, and modelling of the corresponding impedance spectra is possible

An in vitro System for Modeling Brain Reactive Responses andChanges in Neuroprosthetic Device Impedance
 Experiments in cellular biology

EXPERIMENTAL SET UP
Electrodes composed of iridium oxide due to its charge storage properties , with an area of 177 um².
Electrodes were arranged in a 1x16 configuration along the silicon shank with a center-to-center spacing of 100 μm.
Deposition of electrode materials and insulation layers achieved through photolithographic processes.
An Ag/AgCl columnar electrode, filled with 2M KCl and sealed with a Vycor tip was used as a reference electrode.
Platinum foil (2 cm long, 3mm wide) was used as a counter electrode.
Individual device electrodes were connected to the working lead of the potentiostat using a customized 16-prong adapter
Impedance measurements were recorded using a Gamry potentiostat





Custom made chambers were designed to permit impedance measurement from recording devices and 3-D cell cultures

Culture chambers were fabricated around the prepackaged probe and PCB


Recording medium, HEPES buffered Hanks’ saline (HBHS).
Cultures of primary glial cells were obtained using cell isolation procedure.
3-D neural tissue constructs were formed using a micromolding process.
Constructs were modified in terms of cell density and cell.
Confocal microscopy was used to collect images from 3-D cultures labeled with nuclear stains in order to demonstrate the 3-D distribution of glia.
Confocal microscopy is a technique used to form 3–D images from micrographs


Impedance modeling and image analysis
Impedance spectra, displayed as Bode and Nyquist plots
These were fitted to equivalent circuits using a simplex non-linear least squares algorithm and GEA software
The circuit elements can be separated into two groups. Elements that can be attributed to
the electrode and electrode/solution interface
the cell construct coating material
The circuit elements used to model the cellular contributions to impedance were selected based on
capacitive properties of lipid bilayers
reduction in extracellular space in high density cell



Cell constructs contribute additional impedance in two ways
(i) by contributing capacitance in the form of plasma membrane lipid, and
(ii) by reducing the amount of free space for chemical species and electrical current to flow, resulting in a Warburg impedance.
Data from bare and cell-coated electrodes were fitted to equivalent circuit models
The capacitance contributed by cell plasma membranes,and the porous bounded Warburg impedance used to model diffusion limitations in the system.
Values for Rsol and CPE were fixed with respect to the bare electrode measurements, thus permitting accurate fitting of the remaining parameters.
Rct was not fixed due to the relatively large variations observed during measurement and possible interrelation with diffusion controlled processes



Circuit model for bare electrodes



Fitted spectra for an uncoated iridium electrode



Circuit model for cell coated electrodes

Circuit model for cell coated electrodes
Consists of several additional circuit elements
Ccells - capacitance contributed by cell plasma membranes
ZWarburg - porous bounded Warburg impedance to model diffusion limitations in the system.
It could be possible to improve the fit slightly by adding additional circuit elements.
However, it was found that additional elements did not reduce the value (goodness of fit) enough to justify the additions

RESULTS
Data from both bare electrodes and cell-coated electrodes could be accurately fitted to the equivalent circuits depicted
The curve fitting routine for both bare and cell coated electrodes converged in less than 50 iterations for all spectra sampled. These results demonstrate that EIS data from bare electrodes and electrodes surrounded by cells can be interpreted in terms of defined circuit elements.


USES
The ability to use automated image analysis and feature mapping provides a valuable tool for correlating the modeled impedance changes with changes in cell distribution around the electrodes
This measurement system will be useful for monitoring the changes in impedance that accompany glial response to neuroprosthetic devices and can also be applied for use in a variety of biosensor applications

CONCLUSION
The results from impedance characterization and EIS data modelling confirm that EIS can be used to quantify the electrical properties of electrodes and tissues with minimal disruption
The sensitivity, stability and reproducibility of the impedance measurements permit accurate analysis over several time points and could potentially be used to record time lapse impedance data. The ability to model data using standard fitting algorithms adapted for use with equivalent circuit parameters provides a method to describe the physical basis of EIS data.

REFERENCES
Banker G a G, K. 1998 Culturing Nerve Cells (Cambridge: MIT Press)
Bard A J, and Faulkner, L.R. 2001 Electrochemical Methods: Fundamentals and Applications (New York: John Wiley & Sons)
Bertrand C A, Durand D M, Saidel G M, Laboisse C and Hopfer U 1998 System for dynamic measurements of membrane capacitance in intact epithelial monolayers Biophys J 75 2743-56
Crone C and Olesen S P 1982 Electrical resistance of brain microvascular endothelium Brain Res 241 49-55