Hamid Charkhkar

Improving the performance of poly(3,4-ethylenedioxythiophene) for brain–machine interface applications

Conducting polymers, especially poly(3,4-ethylenedioxythiophene) (PEDOT) based materials, are important for developing highly sensitive and microscale neural probes. In the present work, we show that the conductivity and stability of PEDOT can be significantly increased by switching the widely used counter anion poly(styrenesulfonate) (PSS) to the smaller tetrafluoroborate (TFB) anion during the electrodeposition of the polymer. Time-dependent impedance measurements of polymer modified implantable microwires were conducted in physiological buffer solutions under accelerated aging conditions and the relative stability of PEDOT:PSS and PEDOT:TFB modified microwires was compared over time. This study was also extended to carbon nanotube (CNT) incorporated PEDOT:PSS which, according to some reports, is claimed to enhance the stability and electrical performance of the polymer. However, no noticeable difference was observed between PEDOT:PSS and CNT:PEDOT:PSS in our measurements. At the biologically relevant frequency of 1kHz, PEDOT:TFB modified microwires exhibit approximately one order of magnitude higher conductivity and demonstrate enhanced stability over both PEDOT:PSS and CNT:PEDOT:PSS modified microwires. In addition, PEDOT:TFB is not neurotoxic and we show the proof-of-concept for both in vitro and in vivo neuronal recordings using PEDOT:TFB modified microelectrode arrays and chronic electrodes, respectively. Our findings suggest that PEDOT:TFB is a promising conductive polymer coating for the recording of neural activities.

Use of cortical neuronal networks for in vitro material biocompatibility testing

Neural interfaces aim to restore neurological function lost during disease or injury. Novel implantable neural interfaces increasingly capitalize on novel materials to achieve microscale coupling with the nervous system. Like any biomedical device, neural interfaces should consist of materials that exhibit biocompatibility in accordance with the international standard ISO10993-5, which describes in vitro testing involving fibroblasts where cytotoxicity serves as the main endpoint. In the present study, we examine the utility of living neuronal networks as functional assays for in vitro material biocompatibility, particularly for materials that comprise implantable neural interfaces. Embryonic mouse cortical tissue was cultured to form functional networks where spontaneous action potentials, or spikes, can be monitored non-invasively using a substrate-integrated microelectrode array. Taking advantage of such a platform, we exposed established positive and negative control materials to the neuronal networks in a consistent method with ISO 10993-5 guidance. Exposure to the negative controls, gold and polyethylene, did not significantly change the neuronal activity whereas the positive controls, copper and polyvinyl chloride (PVC), resulted in reduction of network spike rate. We also compared the functional assay with an established cytotoxicity measure using L929 fibroblast cells. Our findings indicate that neuronal networks exhibit enhanced sensitivity to positive control materials. In addition, we assessed functional neurotoxicity of tungsten, a common microelectrode material, and two conducting polymer formulations that have been used to modify microelectrode properties for in vivo recording and stimulation. These data suggest that cultured neuronal networks are a useful platform for evaluating the functional toxicity of materials intended for implantation in the nervous system.

Differential responses to ω-agatoxin IVA in murine frontal cortex and spinal cord derived neuronal networks.

ω-Agatoxin-IVA is a well known P/Q-type Ca(2+) channel blocker and has been shown to affect presynaptic Ca(2+) currents as well postsynaptic potentials. P/Q-type voltage gated Ca(2+) channels play a vital role in presynaptic neurotransmitter release and thus play a role in action potential generation. Monitoring spontaneous activity of neuronal networks on microelectrode arrays (MEAs) provides an important tool for examining this neurotoxin. Changes in extracellular action potentials are readily observed and are dependent on synaptic function. Given the efficacy of murine frontal cortex and spinal cord networks to detect neuroactive substances, we investigated the effects of ω-agatoxin on spontaneous action potential firing within these networks. We found that networks derived from spinal cord are more sensitive to the toxin than those from frontal cortex; a concentration of only 10nM produced statistically significant effects on activity from spinal cord networks whereas 50 nM was required to alter activity in frontal cortex networks. Furthermore, the effects of the toxin on frontal cortex are more complex as unit specific responses were observed. These manifested as either a decrease or increase in action potential firing rate which could be statistically separated as unique clusters. Administration of bicuculline, a GABAA inhibitor, isolated a single response to ω-agatoxin, which was characterized by a reduction in network activity. These data support the notion that the two clusters detected with ω-agatoxin exposure represent differential responses from excitatory and inhibitory neuronal populations.

Optical coherence tomography imaging of retinal damage in real time under a stimulus electrode

We have developed a novel method to study the effects of electrical stimulation of the local retina directly under an epiretinal stimulus electrode in real time. Using optical coherence tomography (OCT) and a superfused retinal eyecup preparation, we obtained high-resolution images of the rabbit retina directly under an optically transparent saline-filled fluoropolymer stimulation tube electrode. During OCT imaging, 50 Hz trains of biphasic current pulses 1 ms/phase (23-749 µC cm(-2) ph(-1)) were applied to the retinal surface for 5 min. After imaging, the stimulated regions were stained with the dye propidium iodide (PI) to reveal cytotoxic damage. Pulse train stimulation at 44-133 µC cm(-2) ph(-1) had little effect on the retina; however, trains ≥442 µC cm(-2) ph(-1) caused increases in the reflectance of the inner plexiform layer (IPL) and edema. The damage seen in retinal OCT images matched the pattern observed in histological sections, and in the PI staining. With pulse trains ≥442 µC cm(-2) ph(-1), rapid increases in the reflectivity of the IPL could be observed under the stimulus electrode. Below the electrode, we observed a ring-like pattern of retinal detachment in the subretinal space. The OCT imaging method may be useful for analyzing overstimulation of neuronal tissue by electrodes in many brain regions.

Design and demonstration of an intracortical probe technology with tunable modulus.

Intracortical probe technology, consisting of arrays of microelectrodes, offers a means of recording the bioelectrical activity from neural tissue. A major limitation of existing intracortical probe technology pertains to limited lifetime of 6 months to a year of recording after implantation. A major contributor to device failure is widely believed to be the interfacial mechanical mismatch of conventional stiff intracortical devices and the surrounding brain tissue. We describe the design, development, and demonstration of a novel functional intracortical probe technology that has a tunable Youngs modulus from ∼2 GPa to ∼50 MPa. This technology leverages advances in dynamically softening materials, specifically thiol-ene/acrylate thermoset polymers, which exhibit minimal swelling of < 3% weight upon softening in vitro. We demonstrate that a shape memory polymer-based multichannel intracortical probe can be fabricated, that the mechanical properties are stable for at least 2 months and that the device is capable of single unit recordings for durations up to 77 days in vivo. This novel technology, which is amenable to processes suitable for manufacturing via standard semiconductor fabrication techniques, offers the capability of softening in vivo to reduce the tissue-device modulus mismatch to ultimately improve long term viability of neural recordings.

Chronic intracortical neural recordings using microelectrode arrays coated with PEDOT-TFB.

UNLABELLED Microelectrode arrays have been extensively utilized to record extracellular neuronal activity for brain-machine interface applications. Modifying the microelectrodes with conductive polymers such as poly(3,4-ethylenedioxythiophene) (PEDOT) has been reported to be advantageous because it increases the effective surface area of the microelectrodes, thereby decreasing impedance and enhancing charge transfer capacity. However, the long term stability and integrity of such coatings for chronic recordings remains unclear. Previously, our group has demonstrated that use of the smaller counter ion tetrafluoroborate (TFB) during electrodeposition increased the stability of the PEDOT coatings in vitro compared to the commonly used counter ion poly(styrenesulfonate) (PSS). In the current work, we examined the long-term in vivo performance of PEDOT-TFB coated microelectrodes. To do so, we selectively modified half of the microelectrodes on NeuroNexus single shank probes with PEDOT-TFB while the other half of the microelectrodes were modified with gold as a control. The modified probes were then implanted into the primary motor cortex of rats. Single unit recordings were observed on both PEDOT-TFB and gold control microelectrodes for more than 12 weeks. Compared to the gold-coated microelectrodes, the PEDOT-TFB coated microelectrodes exhibited an overall significantly lower impedance and higher number of units per microelectrode specifically for the first four weeks. The majority of PEDOT-TFB microelectrodes with activity had an impedance magnitude lower than 400 kΩ at 1 kHz. Our equivalent circuit modeling of the impedance data suggests stability in the polymer-related parameters for the duration of the study. In addition, when comparing PEDOT-TFB microelectrodes with and without long-term activity, we observed a distinction in certain circuit parameters for these microelectrodes derived from equivalent circuit modeling prior to implantation. This observation may prove useful in qualifying PEDOT-TFB microelectrodes with a greater likelihood of registering long-term activity. Overall, our findings confirm that PEDOT-TFB is a chronically stable coating for microelectrodes to enable neural recording. STATEMENT OF SIGNIFICANCE Microelectrode arrays have been extensively utilized to record extracellular neuronal activity for brain-machine interface applications. Poly(3,4-ethylenedioxythiophene) (PEDOT) has gained interest because of its unique electrochemical characteristics and its excellent intrinsic electrical conductivity. However, the long-term stability of the PEDOT film, especially for chronic neural applications, is unclear. In this manuscript, we report for the first time the use of highly stable PEDOT doped with tetrafluoroborate (TFB) for long-term neural recordings. We show that PEDOT-TFB coated microelectrodes on average register more units compared to control gold microelectrodes for at least first four weeks post implantation. We collected the in vivo impedance data over a wide frequency spectrum and developed an equivalent circuit model which helped us determine certain parameters to distinguish between PEDOT-TFB microelectrodes with and without long-term activity. Our findings suggest that PEDOT-TFB is a chronically stable coating for neural recording microelectrodes. As such, PEDOT-TFB could facilitate chronic recordings with ultra-small and high-density neural arrays.

Voice controlled wheelchairs

People without disabilities seamlessly control devices with their hands. Interestingly, their hands can perform coarse and fine control. Implementing smooth control for computerized systems is not straightforward and most of the time it is not intuitive either. Here we offer a solution to that problem: smooth control through humming. Voice commands have become ubiquitous in modern technology. Speech-to-text applications abound. Smooth control, on the other hand, has not been tackled yet. Here we design and implement a humming control technique, and demonstrate a hardware implementation with a powered wheelchair. Once actuated, the speed with which the chair moves will depend on the subtle variation on the fundamental frequency of the users humming, acquired through an accelerometer measuring vocal cord vibration. We also discuss two signal processing techniques that handle commonly encountered issues when trying to resolve frequencies in real time data. The hardware implementation shows performance of 80% and higher in speech recognition for signal-to-noise ratio (SNR) higher than 8dB and 100% in smooth control and frequency detection for all tested SNRs. We also discuss potential applications of smooth humming control to other assistive technology.

Amyloid beta modulation of neuronal network activity in vitro.

In vitro assays offer a means of screening potential therapeutics and accelerating the drug development process. Here, we utilized neuronal cultures on planar microelectrode arrays (MEA) as a functional assay to assess the neurotoxicity of amyloid-β 1-42 (Aβ42), a biomolecule implicated in the Alzheimer׳s disease (AD). In this approach, neurons harvested from embryonic mice were seeded on the substrate-integrated microelectrode arrays. The cultured neurons form a spontaneously active network, and the spiking activity as a functional endpoint could be detected via the MEA. Aβ42 oligomer, but not monomer, significantly reduced network spike rate. In addition, we demonstrated that the ionotropic glutamate receptors, NMDA and AMPA/kainate, play a role in the effects of Aβ42 on neuronal activity in vitro. To examine the utility of the MEA-based assay for AD drug discovery, we tested two model therapeutics for AD, methylene blue (MB) and memantine. Our results show an almost full recovery in the activity within 24h after administration of Aβ42 in the cultures pre-treated with either MB or memantine. Our findings suggest that cultured neuronal networks may be a useful platform in screening potential therapeutics for Aβ induced changes in neurological function.

Amorphous Silicon Carbide for Neural Interface Applications

Abstract Implantable neural interfaces offer promise as prosthetic devices that enable communication and control of artificial limbs for individuals suffering from paralysis and amputation. While proof-of-concept recordings from microelectrode arrays (MEAs) have been promising, work in animal models demonstrates that the obtained signals degrade over time until device failure. Amorphous silicon carbide ( a -SiC), a robust material that is corrosion resistant, has emerged as an alternative encapsulation layer for biomedical devices. The purpose of this chapter is to discuss mechanisms involved in the degradation of MEA performance, the role of material choice in the tissue response, the biological response of neural tissue to a -SiC both in vitro and in vivo , and the utility of a -SiC as an encapsulant for implantable neural interfaces.

Effects of Carbon Nanotube and Conducting Polymer Coated Microelectrodes on Single-unit Recordings in vitro

Neuronal networks cultured on microelectrode arrays (MEAs) have been utilized as biosensors that can detect all or nothing extracellular action potentials, or spikes. Coating the microelectrodes with carbon nanotubes (CNTs), either pristine or conjugated with a conductive polymer, has been previously reported to improve extracellular recordings presumably via reduction in microelectrode impedance. The goal of this work was to examine the basis of such improvement in vitro. Every other microelectrode of in vitro MEAs was electrochemically modified with either conducting polymer, poly-3,4-ethylenedioxythiophene (PEDOT) or a blend of CNT and PEDOT. Mouse cortical tissue was dissociated and cultured on the MEAs to form functional neuronal networks. The performance of the modified and unmodified microelectrodes was evaluated by activity measures such as spike rate, spike amplitude, burst duration and burst rate. We observed that the yield, defined as percentage of microelectrodes with neuronal activity, was significantly higher by 55% for modified microelectrodes compared to the unmodified sites. However, the spike rate and burst parameters were similar for modified and unmodified microelectrodes suggesting that neuronal networks were not physiologically altered by presence of PEDOT or PEDOT-CNT. Our observations from immunocytochemistry indicated that neuronal cells were more abundant in proximity to modified microelectrodes.