The Fabrication Of Novel Implantable Microwire Microelectrode Array Of Neural Interface And Its Modification

Neural interface is a functional interface possessing the ability of communication and control between brain(including human brain, animal brain and in vitro neural cells) and outside devices(such as computer, electronic equipment and mechanical device). It can be classified as implantable neural interface, nonimplantable neural interface and in vitro neural interface according to different invasive methods. And implantable neural interface has the advantages of good signal quality, high spatial resolution and precise control accuracy which makes it widely applied in the fundamental research of neurobiology. Implantable microelectrode is the most important part of implantable neural interface. It connects human nervous system to outside devices, and its development and modification has been the key technique in discovering brain function, treating neural diseases and developing human-machine-combined devices.The first implantable microelectrode is a kind of simple microwire electrode that fabricated in 1950. And as the related researches go deeper, the properties of implantable microelectrode need to be better. In recent years, as a lot of progresses have been made in nanotechnology and MEMS(micro electro mechanical system), which have promoted the development of implantable microelectrode, some commercial neural implantable microelectrodes arose, such as the Michigan electrode and Utah electrode which are made of silicon and possess the advantages of microsize and little invasiveness, but these electrodes are difficult to manufacture and the cost is high. While due to the fabrication conveniency and low cost, the microwire microelectrode has become the first choice of many laboratories researching in neurosciences.In the preparation of microelectrode, the main issue focusses on how to promote its electrochemical properties and biocompatibility. Coating electrodes with suitable materials could change its electrochemical properties dramatically. Among the numerous modification materials, carbon nanotubes can form rough and porous structure which readily increase the surface area, reduce the impedance, so it attracts much attention from researchers all over the world. However, the adhesion between sole CNT and substrate is generally poor. Poly(3,4-ethylenedioxythiophene)(PEDOT) is easy to prepared on the neural electrodes, and it has been wildly used as a coating material. PEDOT/CNT composite, a intriguing coating material which can solve the adhension problem of CNT, is becoming a hot research area in improving neural electrode. But PEDOT/CNT composite isn’t the best coating material, its biocompatibility can be further improved with bioactive materials.To improve biocompatibility of neural electrode, it’s intrinsic method is to mimic the extracellular environment on the neural electrode, so as to minimize inflammatory response, and even promote the development of neural tissues. PEDOT has been doped with various bioactive materials including laminin(DCDPGYIGSR and DEDEDYFQRYLI), fibronectin(DCDPGYIGSR) and NGF yielding functional materials which used to improve the adhension, differentiation and proliferation of cells respectively. But bioactive peptides immobilized on CNT or CP surface by simple adsorption or entrapment are not stable because the surrounding extracellular matrix may displace them after implantation, while the function time of bioactive peptides can be prolonged by covalent bonding methods. It is reported that RGD can facilitate the adhesion of many kinds of cells and YIGSR can promote the growing of synpse which are required urgently by implanted electrodes. As a result, it is meaningful to covalently link these biomolecules to the coating surface.The mechanical property of coating materials also influences its biocompatibility. As is known to all that the micromotion always exists in the brain, and the inflammatory response would be worse for the inconformity of mechanical property between brain tissue and neural electrode. Using soft materials such as SU-8 and PDMS to fabricate neural electrode could solve this problem, but almost all coating materials are tough materials, it is reported that PVA/PEDOT conductive hydrogel can be prepared with elastic modulus that very closed to brain tissue, it has great potential to become a soft coating material and researches on the electrochemical properties and biocompatibility of this coating material is meaningful to the improvement of neural electrode.Based on the analysis above, the first subject of this study is to develop a kind of microwire electrode array and a kind of bioactive coating materials featuring a poly(3, 4-ethylenedioxythiophene)(PEDOT)/carbon nanotube(CNT) composite and covalent bonded YIGSR and RGD. Then based on PDMS and PVA/MWCNT hydrogel, we further developed a soft optrode.Part I: The fabrication of microwire microelectrode array and its modification by PEDOT/PSS/MWCNT-peptide.Method: in this part, we developed a kind of coating composite on our home made microwire electrodes and ITO glass. Extracellular peptide RGD and YIGSR were covalent bond to the PEDOT/PSS/MWCNT composite by EDC. We used fluorescamine to react with the PEDOT/PSS/MWCNT-peptide film to identify if the polypeptide had been modified on the composite. SEM was used to investigate the surface morphology and microstructure of the coating materials. Electrochemical impedance spectroscopy(EIS), cyclic voltammetry(CV), and potential transient methods were used to characterize the electrode–electrolyte interface of the modified electrodes. We analyzed the reasons of electrochemical properties improvement by combining SEM result with an equivalent circuit model. The stability of the composites was characterized by the variation of the impedance at 1 k Hz after ultrasonic testing. PC12 cells were used for the study of biocompatibility. The advantages of the improved electrochemical impedance and microstructure surface were analysed by acute and chornic neural recording. And the state of neural cells on the neural interface were investigated to further evaluate the biocompatibility of the coating materials through combining neural recording result with in vivo impedance.Result: the fluorescamine experiment indicates that RGD and YIGSR peptide were successfully covalent bonded to PEDOT/ PSS/MWCNT film. The composite surface is rough and porous, and this phenomenon becomes more serious after being modified by polypeptide. The coated electrodes exhibited significantly higher cathodic charge storage capacity(CSCC=7.5032 m C/cm2) and lower electrochemical impedance at 1 k Hz(reduced from 600kΩ to 15 kΩ) when the test area is 706.5μm2. Through the equivalent circuit model we find that MWCNT increased the efficient surface and porous property of electrodes which makes the impedance decreased dramatically. The ultrasonic testing result shows good adhesion ability of the coating composites. PC12 cells experiment suggests that these coated substrates are favorable to cell attachment and growth. Neural recording result indcates that the increased information content of the coated electrode acquired data with more active channels, higher mean maximal amplitude, higher SNR and reduced noise. Considering the in vivo impedance, the neural recording results also suggests that there are more active neural cells around the electrode interface and the neural cells are closer to the electrode modified by PEDOT/PSS/MWCNT-peptide composite.Conclusion: in this part, YIGSR and RGD were covalent bonded to PEDOT/PSS/MWCNT composite to improve the electrochemical properties and biocompatibility of neural interface. And we firstly combined the in vivo impedance with neural recording results to analyse the biocompatibility of neural interface.Part I I: The fabrication of soft optrode and its modification by soft conductive hydrogel.Method: in this part, we fabricated a kind of soft optrode with soft coating materials. The optrode is based on PDMS, which was prepared by an injection method using glass micropipettes and microwires to form microchannels in the PDMS base. Then another microwires were inserted in the microchannels under microscope, and PVA/MWCNT conductive hydrogel was used to fill the microchannels by the same injection method. We took freeze-thaw method to accomplish the cross-linking and solidity of conductive hydrogel. In the next step, PEDOT/PSS was polymerized in the conductive hydrogel to improve its electrochemical properties. Electrochemical impedance spectroscopy(EIS), cyclic voltammetry(CV), and potential transient methods were used to optimize its preparation technics. And the performance of delivering light and recording neural signal of the optrode were tested to know whether it could be used in optogenetics experiment.Result: upon testing, the optrode could deliver enough light intensity and record neural signal. And we find that optrode with thinner coating materials possessed better electrical properties. The injection method could increase the efficient surface area which also improved the electrical properties. After PEDOT/PSS being added to PVA/MWCNT hydrogel, the optrode impedance at 1 k Hz decreased from ~30kΩ to ~10kΩ, and the optrode has a higher charge injection capability.Conclusion: in this part, we fabricated a kind of soft optrode based on PDMS and PVA/MWCNT. After PEDOT/PSS polymerized in the PVA/MWCNT, the electrochemical properties of the optrode were significantly improved. But the advantages of this optrode in neural recording and its biocompatibility need to be further investigated.