Implantable neural electrode for long-term electrophysiological signal monitoring and preparation method and application thereof

Abstract

The invention relates to the technical field of medical instruments, and discloses an implantable neural electrode for long-term electrophysiological signal monitoring and a preparation method and application thereof. The neural electrode comprises an upper hydrogel insulation packaging layer, a lower hydrogel insulation packaging layer, an electrode site, a connecting line and a bonding pad, wherein the electrode site, the connecting line and the bonding pad are arranged between the upper hydrogel insulation packaging layer and the lower hydrogel insulation packaging layer, and the electrode site and the bonding pad are connected through the connecting line. The electrode sites, the connecting lines and the bonding pads are made of conductive hydrogel; the upper hydrogel insulation packaging layer and the lower hydrogel insulation packaging layer are made of insulation polymers and hydrogel. The insulating polymer is a plasma surface-modified insulating polymer. The preparation method of the neural electrode is simple and efficient, biotoxic materials are not introduced in the preparation process, good biocompatibility is achieved, continuous monitoring in a living body can be achieved for one year, important application value is achieved for studying signal response and neural circuits of all brain regions, and a beneficial tool is provided for long-term in-vivo monitoring.

Description

Technical Field

[0001] The present invention relates to the technical field of medical devices, and in particular to an implantable neural electrode for long-term electrophysiological signal monitoring, a preparation method thereof and an application thereof. Background Art

[0002] At the intersection of biomedicine and neuroscience, there are few biomaterials and electronic interfaces that can be chronically implanted and accurately decode neural activity. Currently, two-dimensional planar electrode arrays have been developed for minimally invasive electrophysiological recording and stimulation of brain activity, commonly known as electrocorticogram (ECoG) neural electrodes. However, traditional ECoG electrode arrays are usually based on metals such as silicon (Si) or platinum (Pt). Although they can effectively capture signals, they face challenges in long-term implantation. The long-term presence of these foreign materials can trigger a persistent immune response and even cause damage and death of neurons near the electrodes. The Young's modulus of traditional electrodes is usually measured in GPa, while the modulus of the brain is 0.5-1.2MPa and the modulus of gray matter / white matter is 1-1.5kPa. The mechanical difference between the two exacerbates the challenge of implantable biocompatibility. In addition, the mechanical mismatch between the electrode and the brain can also lead to uneven strain distribution, shear motion and positioning disorder during implantation, thereby changing physiological responses and compromising signal fidelity. In addition, traditional packaging materials such as polyimide and polyparaxylene have good insulation properties, but due to their hydrophobic nature, they do not have adhesion to brain tissue. Insufficient adhesion often exacerbates the damage to tissue caused by mismatched mechanical properties during long-term implantation of neural electrodes, thereby destroying the stability and service life of the neural interface.

[0003] In addition, how to prepare implantable neural electrode arrays with high precision and high electrode density is another difficulty. Conductive polymer-based hydrogels, such as poly(ethylenedioxythiophene):poly(styrenesulfonic acid) (PEDOT:PSS), have become excellent materials for the conductive layer of neural electrodes due to their controllable mechanical and electrical properties and structural stability in biological environments. However, how to pattern conductive hydrogels to prepare neural electrodes with line widths in the micron range is also a challenge. Recent studies have shown that 3D printing and photolithography technologies have shown a certain degree of help in patterning conductive hydrogels, but photoresists and developers may affect the biocompatibility of the material. Summary of the invention

[0004] In order to overcome at least one of the problems existing in the above-mentioned prior art, one of the purposes of the present invention is to provide an implantable neural electrode for long-term electrophysiological signal monitoring. The second purpose of the present invention is to provide a method for preparing the implantable neural electrode for long-term electrophysiological signal monitoring. The third purpose of the present invention is to provide an electrophysiological signal monitoring device, including the neural electrode or the neural electrode prepared by the method for preparing the neural electrode. The implantable neural electrode for long-term electrophysiological signal monitoring of the present invention includes two layers of hydrogel insulation packaging layers, as well as electrode sites, connecting wires, and pads between the upper and lower layers of hydrogel insulation packaging layers, and the electrode sites and the pads are connected by the connecting wires. The electrode sites, connecting wires, and pads formed by aerosol printing technology provide reliable electrical connections for the neural electrode, the conductive hydrogel provides the neural electrode with the ability to capture tiny electrophysiological signals and apply electrical stimulation, and the hydrogel insulation packaging layer with good hydrophilicity and wet tissue adhesion provides the neural electrode with good biocompatibility and long-term recording stability.

[0005] To this end, the present invention adopts the following technical solutions:

[0006] The first aspect of the present invention provides an implantable neural electrode for long-term electrophysiological signal monitoring, wherein the neural electrode comprises two upper and lower hydrogel insulating packaging layers, as well as electrode sites, connecting wires, and welding pads between the upper and lower hydrogel insulating packaging layers, wherein the electrode sites and the welding pads are connected by the connecting wires.

[0007] Preferably, the electrode sites, connecting wires and pads are all made of conductive hydrogel;

[0008] Preferably, the upper and lower hydrogel insulating packaging layers have the same material composition;

[0009] Preferably, the material composition of the upper and lower hydrogel insulation packaging layers includes insulating polymer and hydrogel;

[0010] Preferably, the insulating polymer is an insulating polymer surface-modified by plasma.

[0011] Preferably, the raw material composition of the conductive hydrogel includes conductive ink and hydrogel;

[0012] The conductive ink, the hydrogel and the insulating polymer can all be obtained through existing technologies, and the present invention does not limit this.

[0013] Preferably, the conductivity of the conductive ink is 4800-5200 S / cm. Further preferably, the conductivity of the conductive ink is about 5000 S / cm.

[0014] Preferably, the conductive ink is a modified poly(3,4-ethylenedioxythiophene) (PEDOT):poly(styrene sulfonate) (PSS). Further preferably, the conductive ink is the EG and LiTFSI modified PEDOT:PSS conductive ink disclosed in the invention patent application "PEDOT:PSS / EG / LiTFSI conductive ink and ultra-flexible electrode and monitoring method of electrophysiological signals" (CN114574040A). The preparation method of the PEDOT:PSS / EG / LiTFSI conductive ink comprises: modifying the PEDOT:PSS solution with ethylene glycol and lithium bistrifluoromethanesulfonyl imide, wherein the viscosity of the PEDOT:PSS / EG / LiTFSI conductive ink is 1 to 1000 cP and the surface tension is 30 to 40 mN / m.

[0015] Preferably, the raw material components of the hydrogel include sodium alginate and polyacrylamide. Further preferably, the raw material components of the hydrogel are the peelable polyacrylamide / sodium alginate ultrathin hydrogel disclosed in the invention patent application "A peelable ultrathin hydrogel, preparation method and application" (CN113754897A).

[0016] Preferably, the insulating polymer is selected from at least one of poly(p-xylene), polyethylene terephthalate, polyimide, and polydimethylsiloxane. Further preferably, the insulating polymer is selected from at least one of poly(p-xylene), polyethylene terephthalate, and polyimide. Even more preferably, the insulating polymer is selected from poly(p-xylene).

[0017] Preferably, the diameter of the electrode site is 50 μm to 200 μm. Further preferably, the diameter of the electrode site is 80 μm to 120 μm.

[0018] Preferably, the line width of the connecting line is 20 μm to 100 μm. Further preferably, the line width of the connecting line is 20 μm to 80 μm. Even further preferably, the line width of the connecting line is 30 μm to 50 μm.

[0019] Preferably, the diameter of the pad is 100 μm to 2000 μm. Further preferably, the diameter of the pad is 200 μm to 2000 μm. Even further preferably, the diameter of the pad is 500 μm to 1000 μm.

[0020] The electrode sites can perform real-time signal monitoring and stimulation of the implanted brain area, and the pads can transmit the recorded electrophysiological signals to electronic equipment for decoding and analysis.

[0021] The second aspect of the present invention provides a method for preparing the neural electrode according to the first aspect of the present invention, comprising the following steps:

[0022] (1) Using a surface-modified insulating polymer and combining it with a hydrogel to prepare a hydrogel insulating encapsulation layer;

[0023] (2) using aerosol printing technology to print on the hydrogel surface of the hydrogel insulating packaging layer to obtain the electrode sites, connecting wires, and pads;

[0024] (3) Covering the upper surfaces of the electrode sites, connecting wires, and pads with a hydrogel insulating packaging layer, etching to obtain electrode sites and pad openings, thereby obtaining the neural electrode.

[0025] Preferably, in step (1), the surface modification of the insulating polymer is carried out by O2 plasma. After O2 plasma modification, many oxygen-containing functional groups are generated on the surface of the insulating polymer, and these functional groups can form hydrogen bonds with the hydrogel to increase the bonding strength between the two.

[0026] Preferably, in step (1), the surface modification time is 2 min to 10 min.

[0027] Preferably, in step (1), the method for preparing the hydrogel comprises the following steps:

[0028] S1: peeling off one of the two hydrophobically modified release films attached to the two side surfaces of the peelable hydrogel to expose one side of the hydrogel base, and the other hydrophobically modified release film that has not been peeled off plays a role in fixing the shape of the hydrogel;

[0029] S2: placing the hydrogel with one side exposed on a hot stage to heat and dry it, preferably, the heating temperature is 50°C to 90°C; more preferably, the heating temperature is 60°C to 70°C; the purpose of heating and drying is to increase the Young's modulus of the hydrogel and separate it from another unpeeled hydrophobic modified release film;

[0030] S3: placing the dried hydrogel separated from another unpeeled hydrophobically modified release film into deionized water to completely swell the hydrogel; preferably, the swelling time is 1 min to 40 min; more preferably, the swelling time is 5 min to 20 min;

[0031] S4: The fully swollen hydrogel is taken out from the deionized water in a conformal manner, and the taking-out method can be with the aid of a substrate material; preferably, the substrate material can be polyethylene terephthalate, polyimide, polydimethylsiloxane and a sieve; further preferably, the taking-out method is with the aid of a pre-soaked sieve, so that when the fully swollen hydrogel is taken out from the deionized water in a conformal manner, excess deionized water can be leaked out.

[0032] Preferably, in step (1), the method for preparing the hydrogel insulating encapsulation layer comprises the following steps:

[0033] The fully swollen hydrogel is transferred conformally to the insulating polymer surface modified by O2 plasma, and then placed on a hot plate for heating to promote the evaporation of water and the fixation of the hydrogel. Preferably, the heating temperature is 50°C to 90°C; more preferably, the heating temperature is 60°C to 70°C.

[0034] Preferably, in step (3), the etching is performed by mask plasma etching, and the mask is made of polydimethylsiloxane / polyimide.

[0035] Preferably, in step (3), the process flow of opening the holes includes:

[0036] S1: patterning the mask through a laser lithography process according to the electrode sites and pads of the neural electrode;

[0037] S2: The upper and lower surfaces of the electrode sites, connecting wires and pads are respectively covered with a hydrogel insulating packaging layer, and after aligning the patterned mask with the electrode sites and pads, holes are opened in the electrode sites and pads, and the hole opening process is plasma etching.

[0038] The third aspect of the present invention provides an electrophysiological signal monitoring device, comprising the neural electrode described in the first aspect of the present invention or a neural electrode prepared by the method for preparing the neural electrode described in the second aspect of the present invention.

[0039] Compared with the prior art, the present invention has the following beneficial effects:

[0040] The present invention provides an implantable neural electrode for long-term electrophysiological signal monitoring. Specifically, the implantable neural electrode for long-term electrophysiological signal monitoring prepared by the above method has good electrical properties and excellent flexibility, and has good conformal adhesion after being implanted in the cerebral cortex of mice, and can monitor the normal physiological activities of mice; in addition, with the help of the volume capacitance of the conductive hydrogel, electrical stimulation is successfully implemented on specific brain areas.

[0041] The present invention provides an implantable neural electrode for long-term electrophysiological signal monitoring. No biotoxic materials are introduced during the preparation process. With the help of the high biocompatibility and biofriendliness of the materials, the neural electrode of the present invention has good biocompatibility and can continuously monitor in vivo for up to one year. Therefore, the implantable neural electrode for long-term electrophysiological signal monitoring of the present invention has important application value for studying the signal response and neural circuits of various brain regions, and provides a favorable tool for long-term in vivo monitoring. BRIEF DESCRIPTION OF THE DRAWINGS

[0042] The present invention will be further described below with reference to the accompanying drawings and embodiments, wherein:

[0043] Figure 1 The flexibility and mechanical compliance of the neural electrode prepared in Example 1 are demonstrated.

[0044] Figure 2 1 is the stress-strain curve of the nerve electrode in Example 1 in the dry and swollen states.

[0045] Figure 3 These are cell fluorescence images of the neural electrodes and the blank control group in Example 1 cultured in the extract solution for 1 day, 3 days, 5 days and 7 days.

[0046] Figure 4 This is the in vitro cytotoxicity test of the neural electrode in Example 1.

[0047] Figure 5 This is a fluorescent image of the in vivo biocompatibility of the neural electrode in Example 1.

[0048] Figure 6 This is a comparison diagram of the fluorescence intensity between the implanted area and the non-implanted area of ​​the neural electrode in Example 1.

[0049] Figure 7 This is a schematic diagram of the neural electrodes covering the left hemisphere of the mouse brain in Example 1.

[0050] Figure 8 The neural electrode in Example 1 captures the neural response of mice during spontaneous whisker movements.

[0051] Fig. 9 This is the neural response of the neural electrode in Example 1 when applying electrical stimulation to the mouse.

[0052] Fig.10 This is a waveform diagram of electrophysiological signals captured by all channels of the neural electrode in Example 1 one day after implantation in a New Zealand rabbit.

[0053] Fig.11 The purpose is to decode signals of different wavelength bands for the electrophysiological signals captured by all channels of the neural electrodes in Example 1 after being implanted in New Zealand rabbits on the first day.

[0054] Fig.12 The graphs are for the changes in signal intensity versus frequency of the nerve electrode in Example 1 under three different physiological conditions on the 1st day and the 365th day after implantation in New Zealand rabbits. DETAILED DESCRIPTION

[0055] The present invention is further described in detail below through specific embodiments and test examples, but is not limited to all discussions and data.

[0056] The conductive ink is the EG and LiTFSI modified PEDOT:PSS conductive ink disclosed in the invention patent application "PEDOT:PSS / EG / LiTFSI conductive ink and ultra-flexible electrode and monitoring method of electrophysiological signals" (CN114574040A). The preparation method of the PEDOT:PSS / EG / LiTFSI conductive ink includes: modifying the PEDOT:PSS solution with ethylene glycol and lithium bistrifluoromethanesulfonyl imide, wherein the viscosity of the PEDOT:PSS / EG / LiTFSI conductive ink is 1 to 1000 cP and the surface tension is 30 to 40 mN / m.

[0057] The raw material components of the hydrogel are the peelable polyacrylamide / sodium alginate ultrathin hydrogel disclosed in the invention patent application "A peelable ultrathin hydrogel, preparation method and application" (CN113754897A), and the preparation method is prepared according to the preparation process of Example 1 thereof.

[0058] The insulating polymer is polyparaxylene.

[0059] It is particularly important to emphasize that, unless otherwise specified, the raw materials, reagents or devices in the present invention can be obtained from conventional commercial channels.

[0060] The term "about" used in the present application indicates that the error range of a numerical value is within 2%.

[0061] Embodiment 1:

[0062] A method for preparing a neural electrode comprises the following steps:

[0063] S1: peel off one of the two hydrophobic modified release films attached to the two sides of the peelable hydrogel prepared in Example 1 of CN113754897A, so that one side of the hydrogel base is exposed, and the other hydrophobic modified release film that is not peeled off plays a role in fixing the shape of the hydrogel;

[0064] S2: placing the hydrogel with one side exposed on a hot stage and heating it at 60°C to dry it, so as to increase the Young's modulus of the hydrogel and facilitate its separation from the other unpeeled hydrophobic modified release film;

[0065] S3: placing the dried hydrogel separated from another unpeeled hydrophobic modified release film into deionized water and soaking it for 20 min to allow it to swell completely;

[0066] S4: With the aid of a sieve, the fully swollen hydrogel is conformally removed from the deionized water, and the excess deionized water is drained through the sieve;

[0067] S5: Using O2 plasma to modify the surface of poly(p-xylene) to generate oxygen-containing functional groups on its surface;

[0068] S6: Conformally transferring the hydrogel to the poly(p-xylene) surface modified by O2 plasma, and then placing it on a hot stage and heating it at 60°C to promote the evaporation of water and the fixation of the hydrogel, thereby obtaining a hydrogel insulating encapsulation layer;

[0069] S7: Place the PEDOT:PSS / EG / LiTFSI conductive ink in Example 1 of CN114574040A in the ink cartridge of an aerosol printer, and print it on the hydrogel surface of the hydrogel insulating packaging layer according to the designed electrode pattern to obtain electrode sites, connecting wires, and solder pads. The electrode sites and solder pads are connected by connecting wires, wherein the diameter of the electrode site in contact with the cerebral cortex is 100μm; the diameter of the solder pad connected to the circuit board is 700μm; and the width of the connecting line is 40μm.

[0070] S8: Patterning the PDMS / PI mask by laser lithography according to the electrode sites and pads of the neural electrodes;

[0071] S9: Cover the upper surfaces of the electrode sites, connecting wires, and pads with a hydrogel insulating packaging layer, align the patterned mask with the electrode sites and pads, and use plasma etching to open holes in the electrode sites and pads to obtain neural electrodes.

[0072] Material performance test:

[0073] The neural electrode obtained in Test Example 1 was tested for mechanical properties, electrical properties and biocompatibility, and the specific results are as follows:

[0074] (1) Figure 1 The flexibility and mechanical compliance of the neural electrode prepared in Example 1 are demonstrated. Figure 1 It can be seen that the neural electrodes can be bent into any shape and fit perfectly with irregular surfaces, thanks to the good flexibility and mechanical compliance of the hydrogel insulating packaging layer.

[0075] (2) Figure 2 is the stress-strain curve of the nerve electrode in Example 1 in the dry and swollen states, Figure 2 a is dry state, Figure 2 b is the swelling state, Figure 2 From a, we can see that the elongation at break of the dry nerve electrode is 13.57% and the Young's modulus is 245MPa. Figure 2 b It can be seen that since the hydrogel absorbs water during the implantation process, thereby reducing its own Young's modulus, the elongation at break of the fully swollen implantable electrode has not changed significantly (10.35%), but its Young's modulus has decreased by an order of magnitude to 61.3MPa. Therefore, whether in a dry state or a fully swollen state, the elongation at break of the neural electrode meets the range of deformation caused by daily brain activities (10%), and the introduction of the hydrogel can act as a dissipative layer during the stretching process, effectively reducing the overall Young's modulus of the neural electrode.

[0076] (3) Figure 3 These are cell fluorescence images of the neural electrodes and blank control group in Example 1 cultured in the extract for 1 day, 3 days, 5 days and 7 days. The neural electrode prepared in Example 1 was tested for in vitro cytotoxicity using rat cortical neuron cells, rat cortical neuron cell complete culture medium and DAPI fluorescence staining. The experimental process is as follows: After adding 0.25% trypsin containing EDTA (Gibco) to rat cortical neuron cells (Wuhan Procell Life Science Co., Ltd.) stored in rat cortical neuron cell culture medium, the culture flask was placed in a 37°C, 5% CO2 incubator and cultured for 3-5 minutes to digest the cells and detach the cells; in order to obtain the neural electrode culture medium extract, the flask was incubated at 3 cm 2 / mL ratio, the nerve electrode is precisely cut and immersed in the complete medium of rat cortical neuron cells, and placed in a 37°C, 5% CO2 incubator for at least 24 hours; during the extraction process, the rat cortical neuron cells that have been detached from the wall are completely broken up to make them evenly dispersed in the culture medium, and then inoculated on a 96-well plate, and placed in a 37°C, 5% CO2 incubator for 24 hours to make them adhere to the wall; after obtaining the extract, the rat cortical neuron cell complete culture medium of the rat cortical neuron cells on the 96-well plate is replaced with the extract, while the rat cortical neuron complete culture medium of the control group is kept unchanged; after being placed in a 37°C, 5% CO2 incubator for 1 day, 3 days, 5 days and 7 days, the cells are stained and characterized by DAPI fluorescent staining. Figure 3 As shown, rat cortical neuron cells survived well, and no cell reduction was observed on the 7th day compared with the control group. In addition, the cell counting kit-8 (CCK-8) test showed that after 7 days of culture, the cell viability of the six different concentrations (4%, 7%, 13%, 25%, 50%, 100%) of the extraction medium did not decrease significantly ( Figure 4 ). Therefore, it is proved that the neural electrode of the present invention has low in vitro cytotoxicity.

[0077] (4) In vivo immunofluorescence analysis was performed in a rat model as follows: For in vivo chronic biocompatibility assessment, rats were anesthetized with an intraperitoneal injection of sodium pentobarbital solution (40-50 mg / kg). Before the start of surgery, it was necessary to confirm that the animals had reached a state of analgesia (surgical anesthesia period), which is characterized by the gradual inhibition of respiration, circulation, muscle tone, and protective reflexes. Anesthetized animals were perfused and fixed with phosphate-buffered saline (PBS, Beingmate, China) and 4% paraformaldehyde (PFA, Beingmate, China) at 4, 8, 12, and 16 weeks after neural electrode implantation. The whole brain was dissected and stored in paraformaldehyde at 4°C for 24 h, and then fixed in 30% sucrose solution (1×PBS) at 4°C for 48 h. The dehydrated brain was then frozen in optimal cutting temperature (OCT, Sakura, USA), and brain slices (40 μm) containing the motor cortex were cut from the coronal direction using a cryostat (Leica, CM1950, USA). The brain slices were rinsed three times with PBS, blocked for 2 h at room temperature (5% BSA and 0.3% Triton X-100), and then incubated with primary antibodies (anti-Iba1, 1:1000, rabbit, Invitrogen; anti-GFAP, 1:300, IgG1, Cell Signaling) diluted in PBST (0.3% Triton X-100) at 4°C overnight to label microglia and astrocytes. Figure 5 As shown in the figure, during the whole implantation period, astrocytes and microglia showed no abnormal cell morphology compared with the non-implanted sites. Figure 6 The fluorescence intensity comparison diagram of the implanted area and the non-implanted area of ​​the neural electrode in Example 1 shows that during the entire observation period of up to 16 weeks, the fluorescence intensity expression levels of the three markers in the implanted area are comparable to those in the area without the electrode array implanted. This indicates that the neural electrode of the present invention has excellent electrical stability, good in vivo biocompatibility, low in vitro cytotoxicity, and can be used for in vivo signal monitoring of organisms.

[0078] (5) The neural electrode prepared in Example 1 was implanted into the brain of a mouse to verify whether the neural electrode could be used to record and stimulate the normal electrophysiological activities of the mouse. The specific operation was as follows:

[0079] Before the operation, the mouse was anesthetized by intraperitoneal injection of 1% sodium pentobarbital solution (60-80 mg / kg). At the beginning of the operation, the mouse's head was fixed and the hair on the surface of the skull was shaved. After disinfecting with iodine and cutting the skin on the surface of the skull, the exposed area was carefully wiped with hydrogen peroxide to remove the soft tissue on the surface of the skull. After the skull was completely exposed, wipe the surface of the skull with a clean cotton ball, and then wipe the surface of the skull dry with an ear wash ball so that the skull sutures were clearly visible. 1 to 2 skull nails were implanted above the cerebellum as a ground reference. Before implantation, the brain was adjusted to ensure that the animal's brain was of the same height and flat on both sides. A skull screw was implanted above the cerebellum as a ground wire. Then, the left brain of the rat was opened with a dental drill, and the exposed size should be slightly larger than the implanted electrode. The craniotomy process should minimize damage to brain tissue. The electrode array implantation area was then exposed by craniotomy and removal of the dura mater. Figure 7 As shown, the neural electrode of the present invention can effectively cover the key areas of the left hemisphere of the brain of C57BL / 6 mice, including the motor cortex, somatosensory cortex, visual cortex, etc. The neural electrode for long-term electrophysiological signal monitoring has a high electrode density, so it can fully capture brain activity and provide a valuable tool for large-scale understanding and regulation of brain function. Figure 8 a is a schematic diagram of the neural response of a C57BL / 6 mouse during spontaneous whisker movement captured by the neural electrode in Example 1. Figure 8 As shown in b, the small white circles represent the distribution of 128 electrode sites in the cerebral cortex. It can be seen that whisker movements evoke different response characteristics in multiple cortical areas. It is worth noting that the primary motor cortex (M1), secondary motor cortex (M2) and secondary visual cortex (V2) all show signal responses when the whiskers move, indicating that these cortices are widely involved in these sensory motor functions. In addition, the primary somatosensory cortex (S1), which plays a key role in whisker movement control, also shows significant activity. Every time a whisker movement occurs, Figure 8 The spectrograms shown in c all produce obvious peaks and corresponding amplitude changes.

[0080] (6) Direct stimulation of specific brain regions can clarify the functional connections and information flow within the neural network. Therefore, using the same mouse model, the present invention uses two electrodes located in the S1 region to provide bidirectional square wave currents in opposite phases for directional stimulation, such as Fig. 9 When stimulation is applied, the remaining electrodes can simultaneously record the response of the cerebral cortex electrophysiological signals generated by the stimulation ( Fig. 9 b), reflecting the multifunctionality of the neural electrode of Example 1 in that it can both apply stimulation and record signals at the same time. In addition, different waveforms can be recorded in the M1, S1 and V2 cortices ( Fig. 9c). Among them, the channel located in S1 closest to the stimulation site showed the most obvious response, characterized by an initial sharp drop in potential, followed by an increase, and then a gradual return to baseline. The electrode located at M1 showed the opposite trend, which was due to its proximity to the antiphase stimulation, so the amplitude was lower and the time to return to the baseline was longer. The electrode placed in the V2 cortex responded to the bidirectional current from both stimulation points. The potential initially rose sharply with the stimulation pulse, then dropped rapidly with the negative stimulation, and then gradually returned to the baseline. The results show that the neural electrode can accurately record the electrophysiological signals generated by the normal physiological activities of mice, and the high electrode density can fully capture brain activity. In addition, this neural electrode can successfully apply electrical stimulation to the implanted area and record the electrophysiological signal response while the electrical stimulation is being performed. Therefore, this neural electrode provides a valuable tool for understanding and regulating brain function on a large scale.

[0081] (7) The neural electrode of Example 1 was implanted into the rabbit brain to verify whether the neural electrode could be used to monitor the signals generated by the normal electrophysiological activities of the rabbit for a long time. The specific operation was as follows:

[0082] Before the start of the operation, the New Zealand rabbit was anesthetized by intravenous injection of 10mg / ml propofol (10mg / kg), and was placed on a heating pad after the breathing was stable and the muscles were relaxed. During the operation, a nose cone was used to maintain anesthesia by adding 3% isoflurane to oxygen. Sterile eye ointment was applied to the rabbit's eyeballs to reduce the risk of corneal irritation and dehydration during the operation. At the beginning of the operation, the New Zealand rabbit's head was fixed and the hair on the surface of the skull was shaved. After disinfecting the skin on the surface of the skull with iodine and cutting, the exposed area was carefully wiped with hydrogen peroxide, and the soft tissue on the surface of the skull was removed. After the skull was completely exposed, the skull surface was wiped with a clean cotton ball, and then the skull surface was wiped dry with an ear wash bulb so that the skull sutures were clearly visible. 6 to 7 skull pins were implanted in the left hemisphere of the rabbit to fix the dental adhesive more firmly, and 4 additional ground wires were inserted to prevent the ground wire from being damaged during long-term implantation. The right brain of the rabbit was opened with a dental drill, and the size of the exposure should be slightly larger than the implanted electrode. The craniotomy process should minimize damage to brain tissue. After opening the skull with sharp tweezers and removing the meninges, the sterilized neural electrodes were implanted on the surface of the exposed area. The removed skull was sterilized and covered back on the craniotomy site and sealed with biological glue. After the biological glue dried, the self-designed shielded electrode box was fixed to the rabbit brain with dental cement. After the operation, the rabbit was given a subcutaneous injection of meloxicam (sc, 0.2mg / kg) for pain relief, and a subcutaneous injection of enrofloxacin (sc, 2.5mg / kg). The rabbit was placed on a warming blanket and returned to the rabbit cage after full recovery.

[0083] like Fig.10As shown in the waveform diagram of the electrophysiological signals captured by all channels of the neural electrode in Example 1 on the first day after implantation in the New Zealand rabbit, on the first day after implantation, all channels displayed the electrophysiological signals generated by the freely moving New Zealand rabbit during normal physiological activities, and there were slight differences in the signals recorded between each channel. The various behavioral states of the freely moving New Zealand rabbit were divided into exploratory behavior, stereotyped behavior, and static behavior to study the response of the cortical electrical signals of the New Zealand rabbit during different physiological activities. Fig.11 As shown, the signals generated when New Zealand rabbits undergo different physiological activities have obvious differences in response in different bands. For example, in the resting behavior state, the signal response in the δ band (1-4 Hz) is the strongest, and as the frequency increases, the signal changes in the exploratory behavior and stereotyped behavior states are also strengthened. It is worth noting that in the stereotyped behavior state, the reaction is mainly concentrated in the β band (13-30 Hz) and weakens with the increase of frequency. On the contrary, in the exploratory behavior state, the signal power increases with the increase of frequency and reaches a peak in the γ band (30-120 Hz). This observation shows that this neural electrode can accurately capture neural activities in a wide frequency range associated with different states. Subsequently, the present invention recorded the long-term electrophysiological activities of a freely moving rabbit. As shown Fig.12 As shown, the neural electrode successfully recorded the cortical electrical signals generated by the New Zealand rabbit during three different physiological activities on the 1st day and the 365th day, thus demonstrating that this electrode has the ability to stably record electrophysiological signals during the one-year study period.

[0084] Obviously, the above embodiments of the present invention are merely examples for clearly illustrating the present invention, and are not limitations on the embodiments of the present invention. For ordinary technicians in the relevant field, other different forms of changes or modifications can be made on the basis of the above description. All obvious changes or modifications derived from the technical scheme of the present invention, such as changing the diameter of the electrode site, the line width of the connecting line, or the diameter of the pad, the raw material components of the hydrogel, etc., are still within the scope of protection of the present invention.

Claims

1. A neural electrode, characterized in that: The neural electrode comprises two layers of hydrogel insulation packaging layers, an electrode site, a connecting wire, and a welding pad between the two layers of hydrogel insulation packaging layers, and the electrode site and the welding pad are connected by the connecting wire; The electrode sites, connecting wires and pads are all made of conductive hydrogel; The upper and lower hydrogel insulation packaging layers have the same material composition; The material composition of the upper and lower hydrogel insulation packaging layers includes insulating polymer and hydrogel; The insulating polymer is an insulating polymer surface-modified by plasma.

2. The neural electrode according to claim 1, characterized in that: The raw material composition of the conductive hydrogel includes conductive ink and hydrogel; The conductive ink is modified poly(3,4-ethylenedioxythiophene):poly(styrene sulfonate).

3. The neural electrode according to claim 1, characterized in that: The raw material components of the hydrogel include sodium alginate and polyacrylamide.

4. The neural electrode according to claim 1, characterized in that: The insulating polymer is selected from at least one of polyparaxylene, polyethylene terephthalate, polyimide and polydimethylsiloxane.

5. The neural electrode according to claim 1, characterized in that: The diameter of the electrode site is 50 μm to 200 μm; And / or, the line width of the connecting line is 20 μm to 100 μm; And / or, the diameter of the pad is 100 μm to 2000 μm.

6. The method for preparing a neural electrode according to any one of claims 1 to 5, characterized in that: The following steps are involved: (1) Using a surface-modified insulating polymer and combining it with a hydrogel to prepare a hydrogel insulating encapsulation layer; (2) using aerosol printing technology to print on the hydrogel surface of the hydrogel insulating packaging layer to obtain the electrode sites, connecting wires, and pads; (3) Covering the upper surfaces of the electrode sites, connecting wires, and pads with a hydrogel insulating packaging layer, etching to obtain electrode sites and pad openings, thereby obtaining the neural electrode.

7. The method for preparing a neural electrode according to claim 6, characterized in that: In step (1), the surface modification is carried out using O2 plasma.

8. The method for preparing a neural electrode according to claim 6, characterized in that: In step (1), the surface modification time is 2 min to 10 min.

9. The method for preparing a neural electrode according to claim 6, characterized in that: In step (3), the etching is performed by mask plasma etching, and the mask is made of polydimethylsiloxane / polyimide.

10. An electrophysiological signal monitoring device, comprising the neural electrode according to any one of claims 1 to 5 or the neural electrode prepared by the method for preparing the neural electrode according to claims 6 to 9.

Images