Multifunctional Flexible Parylene-Based Intracortical Microelectrodes

Proceedings of the 2005 IEEE Engineering in Medicine and Biology 27th Annual Conference Shanghai, China, September 1-4, 2005

Multifunctional Flexible Parylene-Based Intracortical Microelectrodes D. S. Pellinen, T. Moon, R. J. Vetter, R. Miriani, D. R. Kipke Department of Biomedical Engineering, University of Michigan, MI, USA

Abstract- Delivering drugs directly to the brain tissue opens new approaches to disease treatment and improving neural interfaces. Several approaches using neural prostheses have been made to deliver drugs directly with bypassing the bloodbrain barrier (BBB) [1, 2]. In this paper, we propose a new polymer-based flexible microelectrode with drug delivery capability. The probe was fabricated and tested for electrical and fluidic functionality in early stage design. In vivo chronic recording experiments succeeded in demonstrating the in vivo reliability of the probe. Successful in vivo experiments confirm the suitability of the probes as implantable chronic recording devices with robust fluid delivery function. Keywords— flexible polymer microelectrode, parylene-C, multifunctional probe, micro drug delivery, neuroprosthesis

I. INTRODUCTION Therapeutic drugs would be able to cure diseases of the brain if only direct access to brain tissue were available. While advances in protein and peptide chemistry provide many neuro-active compounds that have therapeutic potential as therapeutic treatments, their use is limited by the difficulties in delivery. The ability to deliver these treatments systemically is hindered by the relatively short half-life of the molecules and by metabolic degradation by the liver prior to reaching the central nervous system (CNS) as a target site. In the past, the only option has been to give extremely large doses of medication; however, the patient would likely suffer serious damage elsewhere in the body. In addition, the blood-brain barrier (BBB) poses a significant barrier for the delivery of many neuro-active chemicals. The BBB is a protective cellular barrier that regulates the internal environment with a mechanism of low passive permeability combined with a highly selective transport system. If drugs could be delivered to the CNS directly, the interaction would be highly target-specific and the therapeutic effect would improve dramatically. A variety of approaches have been made to deliver the drug directly, including the use of microfabricated devices, chambers, nanoparticles, coupling drug to sensors and other implants [3]. If this direct drug delivery were to be realized, anti-inflammatory drugs for the long-term chronic recording of the electrical signal could be delivered. These directly delivered drugs would be able to prevent inflammatory reactions of brain tissue caused by microprobe insertion, and thus ensure a stable long-term chronic recording of the

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electrical signal. Rathnasingham et al. have more specifically approached the drug delivery for brain tissue and have shown the feasibility of direct drug delivery in their acute experiment [1]. Retterer et al. have shown that it takes relatively long for brain tissue to react against microprobe insertion [2]. This slow response time justifies the need for a chronic direct drug delivery system for brain tissue. The goal of this study is to develop a flexible polymer based micro system that can deliver drugs directly to brain tissue and record electrical signal at the same time. Polymer substrates expand the current design space of MEMS-based electrodes by their inherent flexibility. Flexibility allows electrodes to be formed into unique 3-dimensional shapes, and allows them to bend around structures to help reach difficult targets within the brain. Flexibility can also provide strain relief against the forces of ‘micromotion’ between the tissue and the implanted device. We have chosen parylene-C as the polymer substrate because of its long-term biocompatibility and low water absorption. The proposed drug delivery system would be able to deliver either therapeutic or anti-inflammatory drugs. II. METHODOLOGY The fluid delivery probe used in this study was designed and fabricated by our group at the NEL (Neural Engineering Lab) at the University of Michigan [4]. It is a microfabricated, multi-channel polymer probe capable of selectively delivering chemicals at the cellular level as well as electrically recording and stimulating neurons in vivo. The width and height of the fluidic channel are 9 Pm and 50 Pm, respectively. The electrical recording sites are located on the top side of the probe í designed to function for both electrical recording and stimulation (Fig. 1). The probe is mounted on a custom-built printed circuit board (PCB) with integrated electrical connections. The probe is ball-bonded onto the PCB [5], on which an electrical connector (Omnetics Connector Co., Minneapolis MN) is soldered. This assembly gives electrical connection between the electrode sites on the microprobe and the electrical signal recording system (Plexon Inc., Dallas, Texas). Access to the flow channel is provided through a polyimide tube (A-M systems, Inc.). The flow is driven by a volume flow controlled syringe pump (WPI

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Fig. 1. Photograph view of the flexible parylene-based microfluidic electrode. The total thickness of the device is nominally 20 µm, with a channel height of 5 µm.

UltraMicroPump) and a micro syringe (SEG 5 PL). The fluid is initially filtered through a 0.22 Pm in-line filter (Corning, Inc.) to remove trace particulates. The connection between the outlet of the syringe and the inlet port of the microprobe is made by polyimide tubing with a 100-Pm inner diameter. The fluid enters the inlet port of the microprobe, and flows through the microchannel inside of the probe until it comes out of the outlet ports of the microprobe. These outlet ports are placed at the tip of the probe as shown in Fig. 1. A. In vitro test - Electrical In order to validate the recording ability, impedance tests was undertaken. Impedance testing of the electrodes was done before/after electroplating and after the in vivo test using an Autolab potentiostat PGSTAT12 (Eco Chemie, Utrecht, The Netherlands) with a built-in frequency response analyzer (Brinkmann, Westbury, NY). The purpose of these repeated tests was to validate whether the electroplating or the insertion process into brain tissue changes the electrical characteristics of electrode sites. In order to lower the impedance and elevate the electrode surface, platinum black was plated on the microelectrode sites [6]. The thickness and the shape of the plating can be controlled by applying various current density and applying time. The solution used for electroplating was 0.25N HCl with 3% PtCl. The current density ranged from 6 mA/cm2 for a light smooth coat, to 30 mA/cm2 for a heavy, grainy coating. Electroplating with Pt-black is able to reduce the impedance by several orders of magnitude. B. In vitro test - Microfluidic High Performance Liquid Chromatography (HPLC) grade water was injected to visually validate the fluidic line in the device assembly. If the fluidic line has no clogging or leakage, a water bubble appears at the outlet port of the probe. The water delivered to the tip was detected visually under microscope. C. In vivo test

Initial in vivo studies of flexi-puff electrodes were conducted on three rats. The first rat was implanted with two electroplated flexi-puff electrodes, while the other two were implanted with both an electroplated flexi-puff electrode, and a 16 channel chronic University of Michigan silicon microelectrode. Both microelectrodes were implanted in the forelimb region of the motor cortex in the initial two rats (FP3 and FP4). The second electrode was inserted at a shallow angle during the surgery and the electrical integrity was compromised in FP3. In the final rat (FP5) both electrodes were implanted in the hind limb region of the motor cortex. The first two rats were studied for 28 and 40 days respectively, at which time the animals were sacrificed for histology. The third and final rat did not recover from the surgery and died of natural causes. Initial anesthesia was administered via an intraperitoneal (IP) injection of a ketamine cocktail consisting of 50 mg/mL ketamine, 5 mg/mL xylazine, and 1 mg/mL acepromazine at an injection volume of 0.125 mL/100g body weight. Updates of 0.1 mL ketamine [50 mg/mL] were delivered as needed during the surgery, approximately every hour. Animals were secured to a standard stereotaxic frame and the cranium was exposed and cleaned. Three stainless steel bone screws were inserted into the skull. The microelectrodes’ connectors were grounded to the bone screw that was over the parietal cortex using a stainless steel ground wire. A craniotomy, approximately 3x2 mm, was created over the target cortical area. Two incisions were made in the dura mater over the exposed area to create four flaps, which were then folded back over the edge of the craniotomy. Insertion of the devices can be accomplished via several different methods. Because of the extreme flexibility, hand insertion is difficult. A silastic coated tab has been placed 2 mm from the recording site at the tip. This tab is used for handling the electrode. After the dura is resected, the electrode can be implanted by hand, similar to the process for implanting Michigan silicon electrodes. The tip of the electrode has been designed to be sharp enough to pierce the pia mater. A mechanical insertion device that will accept polymer, as well as silicon electrodes is also under development. After insertion into the motor cortex, the forelimb cortex coordinates used were: 3.0 mm anterior to bregma, 2.5 mm lateral from midline, and 1.4mm deep from the surface of the brain. The hindlimb cortex coordinates were: 1.5 mm posterior to bregma, 2.0 mm lateral from midline, and 1.4 mm deep from the surface of the brain. Next, NeuroSeal (NeuroNexus Technologies, Ann Arbor, MI) was applied as a dural sealant. The silicon and parylene cable connectors were covered in a silicone elastomer (“Kwik-Sil” by World Precision Inst.), all of which were then enclosed with dental acrylic (Co-Oral-Ite Dental Mfg. Co.) for additional protection, thus leaving the omnetics connector available to plug into. Finally, sutures were used to close

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the skin around the acrylic and anti-bacterial ointment was applied to this region. A final IP injection of Buprenorphine at an injection volume of 0.01-0.05 mg/kg body weight was given for pain relief. Initial neural unit activity was sorted and recorded using a Multi-Channel Neural Acquisition Processor (MNAP; Plexon Inc., Dallas, TX.). The animals were then allowed to recover from the surgery for 48 hours. All procedures were performed under University of Michigan University Committee on the Use and Care of Animals (UCUCA)-approved protocols. III. RESULTS A. In vitro test - Electrical The impedance for the electrode sites were measured before/after electroplating and after surgery. The impedance of electrode sites decreased with electroplating. However, the impedance increased a little bit after surgery but still remained in the acceptable range for unit recording. These results suggest that the recording ability of the electrodes was not damaged during the probe implantation. We can deduce that a microprobe with low impedance is able to record the electro-physiological activity of neurons after implantation. B. In vitro test - Microfluidic Fig. 2 shows a sequential process of a water bubble growing at the fluidic outlet port of the polymer microprobe. HPLC grade water from the syringe pump passed through the fluidic channel in the microprobe and came out through the outlet port of the probe. C. In vivo test Units were detected on all bonded channels and the activity of a single unit was followed consistently out to for 8 days. Fig. 3 shows the recording capabilities of the parylene electrode. Data snapshots in Fig. 3 show 3 different units on 3 different channels on Day 4 after the implantation for rat FP4. The data set includes waveforms, raw data samples, and action potentials. Since the scales are different on each picture, the time and amplitude scales are included along each axis. The data shows simultaneous recording of at least three different units from three of the channels of the flexi-puff electrode implanted in rat FP4. The plots in the first row in Fig. 3 show series of overlayed, thresholded action potentials recorded from rat cortex from sites 7, 8 and 12 of a parylene flexi-puff electrode array (site size 40 x 40 um). The unit activities from the recording sites 7, 8 and 12 are shown in second row. The plots in third row are magnified view of the action potential in second row.

Fig. 2. Pumping through a microfluidic electrode. Figure A shows the whole electrode, up to the bonding pads. The fluid connection (not shown) is to the far left of the probe. Figure B shows the implant portion of the electrode prior to fluid delivery. Figure C show the initiation of flow, which quickly develops into a large bubble of fluid in Figure D.

IV. DISCUSSION The results of pilot experiment set up support the functionality of the probe. In vitro water tests showed that the fluidic function of the probe was robust as to be used for in vivo drug delivery. The fluidic channel tests are very important because of the intrinsic problems in microfluidics like channel clogging or air bubble encapsulation. The difficulties in working with microfluidic devices has been pointed out by Rathnasingham et al.[1] and Ren et al.[7]. The electroplating decreased the impedance of the electrode sites and hence improved the electrical recoding ability. The recoding ability was still maintained even after in vivo testing, indicating that the recoding ability of the polymer probe is so robust that the probe is able to be used not only for acute experiments but also for chronic recordings with implanted in the brain tissue. The results from in vivo tests verified the recoding ability of the polymer probe for chronic recoding. V. CONCLUSION A custom designed polymer probe was successfully fabricated and tested for electrical and fluidic functionality in early stage design. Chronic experiments succeeded in demonstrating the in vivo reliability of the probe. ACKNOWLEDGMENT

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Fig. 3. Recording capabilities of the electrode demonstrated with data snapshots: waveforms, raw data samples, and action potentials of 3 different units on 3 different channels on Day 4 after the implantation for FP4.

The authors would like to thank the Neural Engineering Laboratory. Probes were fabricated and assembled at the Michigan Nanofabrication Facility (MNF). This work was supported by the center for neural communication technology (CNCT) and the Engineering Research Centers program of the National Science Foundation under NSF Award Number EEC-9986866. REFERENCES [1]

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