Electropolymerized Poly(3,4-ethylenedioxythiophene) (PEDOT

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Biological and Medical Applications of Materials and Interfaces

Electropolymerized poly(3,4-ethylenedioxythiophene) (PEDOT) coatings for implantable deep-brain stimulating microelectrodes Côme Bodart, Nicolo' Rossetti, Jo'Elen Hagler, Pauline Chevreau, Danny Chhin, Francesca Soavi, Steen B. Schougaard, Florin Amzica, and Fabio Cicoira ACS Appl. Mater. Interfaces, Just Accepted Manuscript • DOI: 10.1021/acsami.9b03088 • Publication Date (Web): 12 Apr 2019 Downloaded from http://pubs.acs.org on April 13, 2019

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ACS Applied Materials & Interfaces

Electropolymerized Poly(3,4-ethylenedioxythiophene) (PEDOT) Coatings for Implantable Deep-Brain Stimulating Microelectrodes Côme Bodart1, Nicolò Rossetti1, Jo’Elen Hagler1, Pauline Chevreau1, Danny Chhin2, Francesca Soavi3, Steen Brian Schougaard2, Florin Amzica4, Fabio Cicoira1*

1Department

of Chemical Engineering, Polytechnique Montréal, Montréal, Québec, H3C 3J7, Canada

2Department

of Chemistry, Université du Québec à Montréal, Montréal, Québec, H3T 1J4, Canada

3Dipartimento

di Chimica Giacomo Ciamician, Alma Mater Studiorum Università di Bologna, Via Selmi 2, Bologna 40126, Italy.

4School

of Medicine/Dentistry, Université de Montréal, Montréal, Québec, H3C 3J7, Canada

Abstract

Conducting polymers have been widely explored as coating materials for metal electrodes to improve neural signal recording and stimulation because of their mixed electronic-ionic 1 ACS Paragon Plus Environment

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conduction and biocompatibility. In particular, the conducting polymer poly (3,4ethylenedioxythiophene) (PEDOT) is one of the best candidates for biomedical applications due to its high conductivity and good electrochemical stability. Coating metal electrodes with PEDOT has shown to enhance the electrode’s performance by decreasing the impedance and increasing the charge storage capacity. However, PEDOT-coated metal electrodes often have issues with delamination and stability resulting in decreased device performance and life-time. In this work, we were able to electropolymerize PEDOT coatings on sharp platinum-iridium recording and stimulating neural electrodes and demonstrated its

mechanical

and

electrochemical

stability.

Electropolymerization

of

PEDOT:tetrafluoroborate was carried out in three different solvents: propylene carbonate, acetonitrile and water. The stability of the coatings was assessed via ultrasonication, phosphate buffer solution soaking test, autoclave sterilization and electrical pulsing. Coatings prepared with propylene carbonate or acetonitrile possessed excellent electrochemical stability and survived autoclave sterilization, prolonged soaking and electrical stimulation without major changes in electrochemical properties. Stimulating microelectrodes were implanted in rats and stimulated daily, for 7 and 15 days. The electrochemical properties monitored in vivo demonstrated that the stimulation procedure for both coated and uncoated electrodes decreased the impedance.

Keywords: PEDOT, electrochemistry, in vivo stimulation, bioelectronics, neural electrodes.

Introduction

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Conducting polymers are widely investigated as coating materials for implantable recording and stimulating neural electrodes because of their high conductivity and ability to reduce foreign body response1. A common technique used to deposit conducting polymers on metal electrodes is electropolymerization. This deposition method is carried out in an electrochemical cell with a solution consisting of a monomer, a supporting electrolyte that supplies the doping ions and a solvent. Electropolymerization is generally performed via three methods: galvanostatic (constant current), potentiostatic (constant voltage) and potentiodynamic (cyclic voltage sweeps within a certain voltage window). Through finely tuning the experimental parameters, such as electropolymerization method, solvent, dopant concentration and type, this technique allows for precise control over the conductivity, morphology and thickness of the deposited coatings2. One of the most reliable conducting polymers for biomedical applications is poly(3,4ethylenedioxythiophene) (PEDOT). Due to the good electrochemical stability, ease of processing and high conductivity of PEDOT, it has been widely employed as a coating for metal recording and stimulating neural electrodes3,4. In addition to being conductive, PEDOT has been shown to be non-cytotoxic and able to promote cellular proliferation in vitro and in vivo

5–10.

A major obstacle hindering the widespread use of electropolymerized PEDOT-

coatings for neural electrodes is their poor adhesion to metal substrates. Several methods have been reported to improve PEDOT adhesion to metals or other conductive substrates. These include chemisorption of EDOT-acid, acting as an adhesion promoter on an indium thin oxide (ITO) surface prior to electrodeposition11, co-electropolymerization of EDOT-acid and EDOT on gold and gold-palladium12, electrografting of amine moieties, either through 3 ACS Paragon Plus Environment


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an EDOT derivative, EDOT-NH2 13 or through pre-electrodeposition of a diazonium-based anchoring layer exposing thiophene groups that PEDOT can bind covalently to during polymerization14. The morphologies and electrochemical properties of electrodeposited PEDOT films are strongly affected by the solvent, the dopant ions and the electropolymerization method. For instance, electropolymerizations yielded smoother layers of PEDOT when utilizing propylene carbonate as a solvent compared to acetonitrile15. When using water as the solvent and PSSas a dopant, electropolymerized PEDOT:PSS layers showed a globular, compact and smooth morphology compared to samples prepared with Cl- as a dopant, whereas the PEDOT:Cl layers were characterized by a flower-like or porous morphology16.

Potentiodynamic

PEDOT:PSS electropolymerization led to smoother layers and higher conductivity than samples prepared via potentiostatic and galvanostatic electropolymerization17. Further, the capacitance of PEDOT:ClO4 coatings has been shown to increase when switching from potentiostatic to galvanostatic electropolymerization18. Studies on the impact of the porosity of PEDOT films with different Li counter-ions, namely LiBF4, LiClO4, LiPF6 and LiN(SO2C2F5)2 (BETI), showed that ion dimensions influenced the size of the film pores19. Although PEDOT electropolymerization has been widely investigated, very few studies have been conducted on sharp microelectrodes with small surface areas, used in critical biomedical applications such as deep brain stimulation (DBS), e.g., for Parkinson’s disease or neural recording. In this work we studied the electropolymerization of PEDOT on platinum-iridium (PtIr) microelectrodes for deep brain stimulation and recording. To optimize the adhesion of 4 ACS Paragon Plus Environment

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PEDOT to the metal electrodes, we investigated several solvents. The adhesion of the coatings to the substrate and their electrochemical stability was tested through sonication, prolonged immersion in phosphate buffer solution, sterilization by autoclaving and electrical pulsing. Adhesion and electrochemical stability were evaluated using cyclic voltammetry (CV) in phosphate buffer saline and potentioelectrochemical impedance spectroscopy (EIS). Finally, PtIr stimulating microelectrodes coated with PEDOT:BF4 were implanted in the brains of two rats, which were electrically stimulated daily up to 7 and 15 days. The impedance of the electrodes was measured in vivo before and after every stimulation.

Materials and methods Parylene-C-coated platinum-iridium (PtIr) recording microelectrodes (PTM23B05) with a length of 51 mm, a shaft diameter of 231 μm and a sharp exposed tip of 18 μm (total active surface exposed 325 μm2) were purchased from World Precision Instruments. Parylene-Ccoated PtIr stimulating microelectrodes (PI201G) with a length of 51 mm, a shaft diameter of 256 μm and a sharp exposed tip of about 150 μm (total active surface exposed 6000 μm2) were purchased from Microprobes. It is worth noting that the active area exposed is much larger for the stimulating microelectrodes. The purpose of increasing the exposed active area for stimulating microelectrodes is to avoid tissue damage caused by higher charge densities.

Isopropanol (IPA, C3H8O, 70%) and acetone (C3H6O, 90%) were purchased from Honeywell Research Chemicals. Acetonitrile (ACN) non-UV and propylene carbonate (PC) (anhydrous, 99.7%) were purchased from Caledon Laboratories and Millipore Sigma, respectively. 3,45 ACS Paragon Plus Environment


ethylenedioxythiophene

(C6H6OS,

97%)

was

purchased

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from

Millipore

Sigma.

Tetraethylammonium tetrafluoroborate (TEABF4, 99%) was purchased from Acros Organics. Phosphate buffer solution (PBS) of pH 7.4 was prepared using PBS tablets purchased from Millipore Sigma. All chemicals were used as received. Electropolymerization was carried out in a three-electrode electrochemical cell using a Bio-Logic VSP-300 Potentiostat equipped with the EC-Lab software. Silver/silver chloride (Ag/AgCl) was used as the reference electrode, a Pt wire as the counter electrode, and the PtIr microelectrodes as the working electrodes. All working electrodes were rinsed in acetone,

IPA

and

deionized

water

(DW)

and

dried

with

nitrogen

prior

to

electropolymerization. Electropolymerization was carried out galvanostatically in ACN, PC, or DW in solutions containing the EDOT monomer in the presence of 120 mM of TEABF4. The monomer concentration was 20 mM in the organic solvents and 10 mM in DW, due to the lower aqueous solubility of the monomer. Electropolymerization parameters are listed in Table 1. Prior to electropolymerization, all solutions were degassed for 10 minutes using nitrogen and a nitrogen blanket was maintained during the electropolymerization, in order to limit the oxygen concentration in the solution and avoid unwanted oxidation reactions. After electrodeposition, the electrodes were rinsed with deionized water, gently dried with nitrogen, and stored in ambient conditions. Table 1. Parameters for galvanostatic electropolymerization of PEDOT. Electrode

[TEABF4] [EDOT] Solvent Current Polymerization Polymerization applied time charge density

PtIr recording 120mM

20 mM PC

17 nA

50 s


260 mC/cm2

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microelectrode

PtIr stimulating microelectrode

20 mM ACN

17 nA

50 s

10 mM DW

6 nA

150 s

20 mM PC

315 nA

50 s

Electrochemical characterizations were carried out in PBS pH 7.4 using a Bio-Logic VSP300 Potentiostat equipped with the EC-Lab software. Cyclic voltammetry (CV) was conducted within a -0.6 V – 0.6 V window (100 mV/s) for both uncoated and coated microelectrodes to prevent overoxidation of PEDOT and water electrolysis. Potentioelectrochemical impedance spectroscopy (EIS) was performed using a sine wave of 10 mV amplitude between 1 Hz and 100 kHz with a 0 V bias vs the reference electrode. Optical microscope images were obtained with an Axio Imager M1 from Zeiss, equipped with an AxioCam Mrm CCD camera. Scanning electron microscopy (SEM) was carried out using a scanning electron microscope equipped with a field emission gun (JEOL JSM7600F), at an acceleration voltage of 2.00 kV. Adhesion tests were performed by placing the PEDOT-coated microelectrodes inside a beaker filled with deionized water and immersed in an ultrasonic bath (Eumax ultrasonic cleaner (ud100sh-4l) with 100W ultrasonic power). The sonication time was limited to 5 minutes to avoid the destruction of the insulating parylene layer. For PBS soaking tests, the coated electrodes were placed in PBS solution stored in ambient conditions to monitor the evolution of our PEDOT:BF4 coatings in a physiological solution. The solutions were changed every week. Sterilization tests were performed by autoclaving the PEDOT-coated electrodes at 121 °C for 30 minutes in a Steris/AMSCO Century Sterilizer V136. 7 ACS Paragon Plus Environment


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Surgery and in vivo stimulations Two female Wistar rats were implanted with two stimulating PtIr microelectrodes, a PEDOT-coated one and an uncoated one, in their subthalamic nucleus (STN). During surgery, the rats were kept under a 2% isoflurane anesthesia. The head of the animal was placed in a stereotaxic frame. The shaved skin was cleaned using an iodine solution and bupivacaine was injected under the scalp. The scalp was incised for approximately 2 cm and tissues and muscles were removed until the skull was clearly visible. The alignment of the skull was adjusted by measuring the vertical coordinates of the lambda and bregma points of the skull. Two holes were pierced in the skull at the implantation coordinates necessary to reach the STN (anteroposterior: - 3.3 mm, lateral: 2 mm bilateral, from the bregma). Four stainless steel screws were also inserted into the skull around the implantation sites and tied together with a silver wire terminated by a connector to create a counter/reference electrode. Finally, the two electrodes were implanted in the two drilled holes (at a depth of 6 mm, from the top of the skull) and bonded to the skull with dental cement. An antibiotic cream was spread around the wound at the end of the surgery to prevent infections, and buprenorphine and fluids were injected prior to the rat waking up. The rats were left to recover for one week after the surgery before any manipulation. For the electrical stimulations, we used symmetrical biphasic pulses of 20 μA and 90 μs for the cathodic phase and an interphase period of 10 μs at a frequency of 130 Hz. This corresponds to roughly 30 μC/cm2 and 1.8 nC/phase. Prior to implantation, the electrodes were immersed in a PBS pH 7.4 solution and stimulated for 2 hours and electrochemical properties were monitored before and after the stimulation. The biphasic pulses were 8 ACS Paragon Plus Environment

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generated by coupling two monophasic FHC pulsar 6i, one delivering the first negative phase and the second delivering the positive phase with a delay of 100 μs (duration of the first phase plus the delay). The waveform and the maximum intensity amplitude were monitored using a DSO9104A Infiniium Oscilloscope, 1 GHz, Megazoom from Agilent Technologies. In vivo stimulation was carried out for 90 minutes per day for 7 days for the first rat and for 15 days for the second, starting after the recovering week. PEDOT-coated and uncoated PtIr microelectrodes were used one after the other. The rats were freely moving during the stimulation. The waveform and maximum applied current were checked before every stimulation. The voltage excursion observed at the microelectrodes during stimulation was recorded at the beginning and the end of every stimulation session and regularly monitored.

Results and discussion

We selected the galvanostatic electropolymerization mode, since it allowed for more precise control over the quantity of deposited material with respect to potentiostatic and potentiodynamic modes. The potential stabilizes after an initial peak in current versus time plots (Figure 1) indicating that galvanostatic electropolymerization takes place without overoxidation of the polymer. The wider electrochemical stability window of the organic solvents and the higher monomer concentration permitted the process to be carried out at higher currents than in water, resulting in faster kinetics (Figure 1). The electrodeposited PEDOT appears as a dark coating on the conductive tip of the electrodes (Figure S1). As expected, the different solvents lead to different morphologies of the coatings. 9 ACS Paragon Plus Environment


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Figure 1. Potential versus time plots for galvanostatic electropolymerization of PEDOT:BF4 on PtIr recording microelectrodes carried out in solutions of TEABF4 and acetonitrile, propylene carbonate or deionized water. PEDOT coatings obtained in ACN and water appear homogeneous, while those obtained in PC display additional overgrowing regions (Figure 2 a, b, c and d). SEM images at higher magnification revealed that coatings processed in organic solvents have a porous morphology, with ACN providing larger pores than PC. On the other hand, PEDOT processed in DW showed a compact cauliflower morphology (Figure 2 e and f).


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Figure 2. SEM images of PEDOT:BF4 galvanostatically electropolymerized on PtIr recording microelectrodes using solvents of propylene carbonate (a and b), acetonitrile (c and d) and water (e and f). SEM voltage: 2.00 kV. 11 ACS Paragon Plus Environment


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In order to gain insight into the electrochemical properties of our PEDOT-coated electrodes, we performed CV and EIS measurements in PBS (Figure 3). In cyclic voltammetry, PEDOT is doped during the anodic scan and dedoped during the cathodic scan. Assuming a reversible process, the de-doping charge equals the doping one. The PEDOT doping charge can therefore be evaluated by calculating the time integral of the cathodic current extracted from a cyclic voltammogram, and is known as the charge storage capacity (CSC)20. The PEDOT-coated recording microelectrodes showed an enhanced CSC (Figure 3 a) (about 147±42 μC for water-processing and 215±45 μC for PC-processing and 214.7±23 μC for ACN-processing) with respect to the bare PtIr (about 20 μC) The PEDOTcoated stimulating microelectrodes displayed a similar behavior: the PEDOT:BF4 coatings prepared in PC increased by 3.8 times the CSC (from 0.66 mC to 2.5 mC) (Figure S7 a). EIS gives important information about charge transfer and electrical double-layer or pseudo-capacitive processes, at different time scales and frequencies. For deep brain stimulation, a low impedance is required to facilitate charge transfer at the electrode-tissue interface and to minimize the energy consumption required for stimulation21. The electropolymerization of PEDOT:BF4 on PtIr recording microelectrodes led to a significant decrease of the impedance within the investigated frequency range (1 to 105 Hz), regardless of the processing solvent (Figure 3 b). For instance, at a frequency of 1 kHz, a commonly used benchmark value for biological processes, we observed an impedance decrease of about 2 orders of magnitude. For PEDOT-coated stimulating microelectrodes, only PC was used as solvent and the impedance at 1 kHz was reduced by about 1 order of magnitude


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(Figure S5 b). These results are in accordance with the literature when using BF4 as a dopant for PEDOT22. The increase in capacitance and the decrease in impedance induced by the presence of the conducting polymer coatings are due to the faradaic (pseudo capacitive) nature of the doping/dedoping process of the conducting polymer coating and their higher active surface area of the polymer with respect to the metal23. The origin of the difference in modulus impedance between PEDOT coatings processed in organic solvents and water can be clarified by examining the Nyquist plots (Figure S2). At low frequencies the imaginary impedance, associated with the capacitive behavior of the electrodes, is largely predominant over the real impedance (Figure S2 a), associated with charge transfer processes. Coatings processed in organic solvents showed higher capacitance with respect to those processed in water. At high frequencies (Figure S2 b), the real impedance increased since the charge-transfer mechanism related to PEDOT doping/dedoping dominates the process. Interestingly, the real impedances for coatings processed in organic solvents were similar, whereas the real part of coatings processed in DW was higher, indicating a higher charge-transfer resistance. Overall, coatings processed in organic solvents led to improved electrochemical properties than those processed in water, i.e., higher CSC, higher capacitance and lower charge-transfer resistance.



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¬¬¬¬

Figure 3. Cyclic voltammetry at 100 mV/s (a) and impedance (b) of bare PtIr and PEDOTcoated recording microelectrodes, with PEDOT galvanostatically electropolymerized in different solvents (propylene carbonate, acetonitrile and water). Number of samples: 4 for uncoated Pt/Ir, 10 for PEDOT processed in PC, 6 for PEDOT processed in ACN, 3 for PEDOT processed in water. Evolution of the impedance of galvanostatically deposited PEDOT:BF4 coatings on PtIr recording microelectrodes, processed in ACN, PC and DW, after 2 weeks of soaking in PBS pH 7.4 (c), and after being steam sterilized at 121 °C for 30 minutes (d). The superior electrochemical properties of coatings processed in organic solvents can be explained by their porous and open morphology, with respect to the globular and compact morphology of those processed in water, as well as by the higher monomer concentration during polymerization. The difference between coatings processed in ACN and PC could be due to the lower solubility of short PEDOT oligomers in ACN with respect to PC15. Low solubility leads to a higher number of nucleation centres and to electropolymerized coatings 14 ACS Paragon Plus Environment

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consisting mainly of short oligomers on the electrodes. On the other hand, high solubility leads to electropolymerization of coatings characterized by longer polymeric chains, as the shorter oligomers dissolve more easily into the solution. In accordance with our observations, Belaidi et al. observed a clear porous morphology for PEDOT:ClO4 films processed in ACN and a compact and smooth morphology for the PEDOT:PSS films processed in water24. These results were attributed to the higher dielectric constant of water (around 80, compared to around 36 for ACN) and to the possibly higher solubility of the oligomers in water due to the presence of PSS-, expected to lead to slower deposition rates and to the formation of larger polymeric chain, resulting in smooth films. Water is also known to facilitate the elimination of protons during PEDOT growth and therefore to allow for a more compact film25. We believe that these factors led to the compact and smoother coatings when using water, and the open porous morphology when using PC and ACN. On the basis of the similarity of the results obtained on the deposition of PEDOT on PtIr recording microelectrodes in organic solvents, which show better performance with respect to those processed on water, PEDOT electropolymerization on PtIr stimulating microelectrodes was realized only in PC. Before implantation in animals, the stability tests were repeated on the PtIr stimulating microelectrodes to ensure the difference in electrode dimension would not affect adhesion or electrical stability.

The stability of the PEDOT:BF4 coatings was evaluated by ultrasonication in water. The coatings processed in organic solvents showed a significantly higher stability versus ultrasonication of samples processed in water (Figure S3). For instance, PEDOT:BF4 coatings 15 ACS Paragon Plus Environment


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deposited in organic solvents on PtIr microelectrodes remained physically stable after 5 minutes of sonication and retained more than 80% of their CSC. Coatings deposited in DW detached from the PtIr microelectrodes after 2 to 3 minutes of sonication. It has been previously observed that for polymerizations carried out in 1:1 ACN/water mixture, PEDOT:PSS coatings delaminated more easily than PEDOT:ClO4 and PEDOT:pTS coatings, due to the higher rigidity induced by the stiffness of PSS9, preventing an efficient attachment to the electrode surface.

This explanation can be extended to our work,

specifically to the delamination of coatings from electrodes observed in samples processed in water. Our coatings processed in water were more rigid and had a more compact morphology than those processed in organic solvents.

Recording and stimulating microelectrodes must possess good electrochemical stability to remain functional over long periods of time after implantation. To test the long-term stability of the electrodes, PEDOT-coated electrodes were immersed in a PBS pH 7.4 solution for two weeks, as recording operations are usually short and sometimes even limited to a few hours, and the electrochemical cyclic voltammetry and impedance were measured. For PEDOT-coated recording microelectrodes processed in PC and ACN, the impedance was only affected by a factor of 2-3 by immersion in PBS and remained considerably lower than that of the bare electrodes, as shown in Figure 3c (see Figure S4 for data extracted on multiple samples). At 1 kHz, the PBS immersion led to a slight increase for electrodes processed in organic solvents (about + 6 kΩ for the PC-processed electrode and about + 4 kΩ for the ACN-processed electrode) and to a larger increase for electrodes processed in 16 ACS Paragon Plus Environment

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DW (about + 25 kΩ) (Figure 3 c). The CSC of the electrodes decreased over the two weeks (between 20 and 25% for all electrodes). For PEDOT-coated stimulating microelectrodes, over the 8 weeks of soaking (a longer period of immersion as they are supposed to stay chronically implanted) the impedance had a small increase (136 Ω), and the CSC retained 82% of its initial value. PEDOT-coated electrodes were sterilized by autoclaving at 121 °C for 30 minutes. Electrochemical characterizations before and after the treatment were conducted. As shown by CSC values extracted from CVs, the CSCs for PEDOT-coated PtIr electrodes were 4 to 5 times higher than the CSC of bare PtIr (roughly 20 μC). Coatings processed in PC and ACN retained about 85% of their CSC, while those processed in DW retained about 60% (Figure S5). The impedances after sterilization of all coatings in PBS remained much lower with respect to bare PtIr. However, the impedance of PEDOT:BF4 processed in organic solvents was stable while an increase (15 kΩ) was noticed for the one prepared in water (Figure 3 d). Electrochemical impedance tests after autoclave sterilization were also performed on samples previously subjected to a soaking test (Figure S6). The results confirm the trend: low increase for organic solvents and larger increase for water. Similar to the sonication test, we may advance the higher rigidity and more compact character of the water-processed coatings leads to a more brittle structure, whereas a more porous morphology would be more resilient against harsh treatments. Overall, our coatings were stable and did not delaminate heavily during steam sterilized, as reported in the literature for PEDOT:PSS9.

In vitro stimulations 17 ACS Paragon Plus Environment


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Prior to implantation, a 2-hour stimulation period in PBS pH 7.4 was done using PEDOT stimulating microelectrodes, coated with PEDOT:BF4 processed in PC, to ensure the stability of the coatings to electrical stimulations. The electrochemical properties remained stable, i.e., no differences were observed before and after the stimulation period (Figure S7).

In vivo stimulations In vivo experiments were performed with stimulating electrodes only. Two rats were stimulated daily for 15 and 7 days. After this period, the rats were sacrificed because of deterioration of the cement holding the electrodes to the skull. EIS measurements were conducted before and after stimulation and the voltage excursion was monitored. The uncoated microelectrode showed a consistent impedance increase (measured at 1 kHz) over time (Figure 4 a). However, after 90 minutes of stimulation, the impedance returned approximately to the initial value and always remained between 20 and 50 kΩ. The PEDOT-coated stimulating microelectrode also showed an overall increase in impedance over time. In addition, we observed that the relative change in impedance pre- and poststimulation always varied. When considering the impedance-frequency relationship for the electrodes studied, an interesting observation is that a decrease in impedance was present only at high frequencies for the bare PtIr stimulating microelectrode, whereas for the PEDOT-coated microelectrode, the impedance decrease was present throughout the entire frequency range studied (Figure 4 b). This was less noticeable but still measurable for the second animal (Figure S8 b).


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The gradual increase of impedance over time is most likely a consequence of the foreign body reaction, i.e., the accumulation of immune cells that encapsulates an intruding device26, that can also be observed when using PEDOT:BF4 as a coating27. However, this increase is neutralized during the stimulation periods. This behavior of the impedance has already been observed for metallic electrodes used in deep brain stimulation applications28,29. As mentioned previously, most studies using PEDOT-coated neural electrodes either tested their stimulation protocol in vitro or did not stimulate in vivo for a long enough period to alter the surrounding environment and notice a change in the electrochemical properties of the PEDOT-coating. A study by Kolarcik et al. observed a change in impedance for PEDOT after a stimulation period, however, their other results were very different from ours: the impedance measured in vivo of their coated and uncoated microelectrodes before stimulation did not show any significant differences, whereas our coated and uncoated electrodes before stimulation showed larger differences in relative impedance in vivo30. A common explanation for the impedance decreases measured in metallic electrodes is that the stimulation ‘cleans’ the interface by removing the adhered cells and adsorbed proteins. This mechanism may also explain the reduction of impedance for PEDOT-coated microelectrodes, however, the interactions between PEDOT-coatings and tissues differ from those between metals and tissues, thus we speculate that the ‘cleaning’ mechanism of the microelectrodes’ surface is similar, despite differences between the materials. In particular, we noticed several differences when considering the repartition of the impedance between real and imaginary parts for coated and uncoated stimulating microelectrodes, (Figure 4 c and d). For the PEDOT-coated microelectrode, the predominant component was the real 19 ACS Paragon Plus Environment


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impedance, mostly due to the really high capacitance that reduces the imaginary component and the encapsulation of immune cells that increases the real component (Figure 4 d). However, after stimulation, both components were reduced, but most significantly the real impedance. At low frequencies the capacitive behavior became dominant, as observed in vitro (Figure S2a). Hence, the stimulation tended to restore the initial behavior observed in vitro for the microelectrode: capacitive behavior at low frequencies and charge resistance transfer at high frequencies, whereas before the stimulation the main component was the real part, associated with the insulating immune cell layer. Similarly, for the PtIr uncoated microelectrode before stimulation, charge transfer dominated on the whole frequency range (Figure 4 c), the semi-circle observed at high frequencies, also noticeable for the PEDOTcoated microelectrode (Figure 4 d), is considered to be the tissue component of the impedance31. After the stimulation, a significant reduction is noticed for both the real and imaginary impedances at low frequencies and mainly the real impedance at high frequencies (Figure 4 c). However, the capacitive behavior did not become dominant at low frequencies after the stimulation, most likely due to the low capacitance of the bare PtIr and possibly a stronger foreign body reaction. In summary, stimulation in vivo reduced both components of the impedance but mostly the resistive or tissue component, i.e., the semi-circle in the Nyquist plots, even to the point that the capacitive component became dominant at low frequencies for the PEDOT-coated stimulating microelectrode. This supports the ‘cleaning’ hypothesis: an insulating layer, represented electrochemically by a highly resistive component, created by an accumulation of immune cells, would be partially removed by the stimulation, leading to a lowering of the 20 ACS Paragon Plus Environment

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real impedance due to an easier charge-transfer and a smaller decrease of the imaginary impedance due to a more accessible surface and thus a higher capacitance. Also, no significant changes were noticed after the stimulation realized in vitro, indicating that the decrease is specific to the biological environment. Nevertheless, the main observation is that after 90 minutes of stimulation, the difference in impedance modulus pre- and poststimulation between PEDOT-coated and uncoated stimulating microelectrodes is reduced. Despite PEDOT:BF4 coatings providing the lowest impedance (Figure 4 a), the gap in impedance between coated and uncoated microelectrodes was smaller after several days of stimulation. The same conclusion could be drawn from results obtained with the second animal (Figure S8).



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Figure 4. Impedance modulus at 1 kHz for PEDOT-coated and uncoated PtIr stimulating microelectrodes before and after 90 minutes of stimulation over 15 days in vivo (a), impedance modulus for PEDOT-coated and uncoated microelectrodes, after being implanted in vivo for 15 days (one week of recovery and 8 days of stimulation), before and after 90 minutes of stimulation in vivo (b) and Nyquist plots of the impedance of PtIr uncoated stimulating microelectrode (c) with an inset showing a zoom on the high frequencies and PEDOT-coated PtIr stimulating microelectrode, before and after 90 minutes of stimulation (d).

Conclusions We carried out electropolymerization of PEDOT:BF4 on PtIr microelectrodes in PC, ACN and water. In order to assess the stability of the coatings the electrodes were subjected to ultrasonication, PBS pH 7.4 soaking, steam sterilization and electrical pulsing. PEDOTcoatings processed in organic solvents showed better stability under ultrasonication, soaking in PBS pH 7.4 and steam sterilization. PtIr stimulating microelectrodes coated with PEDOT:BF4 and uncoated (for comparison) were implanted and tested in vivo for stimulation purposes and proved to be stable in vivo for 15 days. The implanted electrodes showed an increase in impedance over time. However, the stimulation process led to a decrease in impedance for both coated and uncoated microelectrodes. We interpreted this phenomenon as a ‘cleaning’ of the surface of the microelectrodes, although some differences were noticed between the behaviour of the coated microelectrodes and the uncoated control microelectrodes. Our results call for a 22 ACS Paragon Plus Environment

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deeper investigation on the specific interactions between tissues and PEDOT:BF4 in a cerebral environment, e.g. by histological analysis to detect PEDOT release towards living tissues during prolonged implantation periods. A characterization of these interactions would provide the necessary tools to understand the behaviour of conducting polymer in vivo in deep brain stimulation conditions.

ASSOCIATED CONTENT Supporting Information. The Supporting Information is available free of charge on the ACS Publications website. The section contains an optical image of the microelectrodes tips, coated and uncoated, Nyquist plots comparing the influence of the solvent on the impedance, several Bode plots displaying the results of sonication, soaking and autoclaving tests, with standard deviation included, several cyclic voltammograms on the influence of soaking, and the impedance results of the in vivo stimulation of the second rat. AUTHOR INFORMATION Corresponding Author: [email protected]* Author Contributions The manuscript was written through contributions of all authors. All authors have given approval to the final version of the manuscript. ACKNOWLEDGMENT This work is supported by grants FRQNT Team Grant, awarded to FC, SBS and FA, NSERC Discovery, National Defence Discovery supplement (NSERC) awarded to FC. NR is grateful to 23 ACS Paragon Plus Environment


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Canada First Research Excellence Fund for financial support through a TransMedTech Master Excellence Scholarship. We have also benefited from the support of FRQNT and its Regroupement strategique program through a grant awarded to RQMP. F.S. acknowledges the Italy-Quebec Mobility program (MRIF) for financial support.

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In this work, we were able to electropolymerize PEDOT coatings on sharp platinum-iridium recording and stimulating neural electrodes and demonstrated its mechanical and electrochemical stability. 467x98mm (120 x 120 DPI)

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Figure 1. Potential versus time plots for galvanostatic electropolymerization of PEDOT:BF4 on PtIr recording microelectrodes carried out in solutions of TEABF4 and acetonitrile, propylene carbonate or deionized water.



Figure 2. SEM images of PEDOT:BF4 galvanostatically electropolymerized on PtIr recording microelectrodes using solvents of propylene carbonate (a and b), acetonitrile (c and d) and water (e and f). SEM voltage: 2.00 kV. 269x421mm (120 x 120 DPI)


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Figure 3. Cyclic voltammetry at 100 mV/s (a) and impedance (b) of bare PtIr and PEDOT-coated recording microelectrodes, with PEDOT galvanostatically electropolymerized in different solvents (propylene carbonate, acetonitrile and water). Number of samples: 4 for uncoated Pt/Ir, 10 for PEDOT processed in PC, 6 for PEDOT processed in ACN, 3 for PEDOT processed in water. Evolution of the impedance of galvanostatically deposited PEDOT:BF4 coatings on PtIr recording microelectrodes, processed in ACN, PC and DW, after 2 weeks of soaking in PBS pH 7.4 (c), and after being steam sterilized at 121 °C for 30 minutes (d). 789x408mm (120 x 120 DPI)



Figure 4. Impedance modulus at 1 kHz for PEDOT-coated and uncoated PtIr stimulating microelectrodes before and after 90 minutes of stimulation over 15 days in vivo (a), impedance modulus for PEDOT-coated and uncoated microelectrodes, after being implanted in vivo for 15 days (one week of recovery and 8 days of stimulation), before and after 90 minutes of stimulation in vivo (b) and Nyquist plots of the impedance of PtIr uncoated stimulating microelectrode (c) zoom in on the high frequencies and PEDOT-coated PtIr stimulating microelectrode, before and after 90 minutes of stimulation (d). 795x582mm (120 x 120 DPI)


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Electropolymerized Poly(3,4-ethylenedioxythiophene) (PEDOT

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