Vagus nerve activation system, construction method, conductive hydrogel and application

Abstract

The present application relates to the technical field of vagus nerve activation, and particularly relates to a vagus nerve activation system, a construction method, a conductive hydrogel and application. The vagus nerve activation system comprises: a high-frequency electric stimulation generation module, which is used for generating a high-frequency biphasic square wave pulse electric stimulation signal; an injectable conductive hydrogel module, which is used for forming a local conductive interface around a cervical vagus nerve, and the injectable conductive hydrogel module contains PEDOT:PSS, N,O-carboxymethyl chitosan (NOCC) and aldehyde-based hyaluronic acid (HA-CHO); and an ultrasound-guided positioning module, which is used for accurately delivering the injectable conductive hydrogel module to a target site around the cervical vagus nerve; wherein the electric stimulation signal generated by the high-frequency electric stimulation generation module is transmitted to the vagus nerve through the local conductive interface, so as to realize selective activation of afferent fibers of the vagus nerve.

Description

Technical Field

[0001] This invention relates to the field of vagus nerve activation technology, and in particular to a vagus nerve activation system, its construction method, a conductive hydrogel, and its applications. Background Technology

[0002] Chronic consciousness disorder refers to a state in which the recovery of consciousness is impaired for more than 28 days after severe brain injury, including vegetative state (VS) and minimally conscious state (MCS). Epidemiological data shows that there are approximately 70,000 to 100,000 patients with chronic consciousness disorder in my country, placing a heavy burden on families and society. Current clinical methods for promoting awakening mainly include hyperbaric oxygen therapy, drug therapy, and traditional rehabilitation therapy, but their overall efficacy is limited. Finding new and effective strategies for promoting awakening is an important research topic in the field of rehabilitation medicine.

[0003] Vagus nerve stimulation (VNS) is a neuromodulation technique that uses electrical stimulation of the vagus nerve to regulate the function of the central nervous system. The vagus nerve contains approximately 80% afferent fibers and is a crucial bridge connecting the periphery and the central nervous system. Numerous studies have shown that it can activate the nucleus tractus solitarius (NTS) and modulate cortical excitability through multi-level neural circuits, demonstrating its potential application in promoting wakefulness.

[0004] Current VNS technologies mainly include three types: traditional invasive vagus nerve stimulation (cVNS) involves surgically exposing the vagus nerve in the neck and wrapping electrodes around the nerve trunk. Although the stimulation is precise, it has problems such as large surgical trauma, many complications (such as hoarseness, cough, arrhythmia, etc.), long-term implantation of foreign bodies leading to tissue fibrosis and infection risks, and low patient acceptance; percutaneous auricular vagus nerve stimulation (taVNS) achieves non-invasive modulation by stimulating the auricular branch of the vagus nerve on the surface of the auricle. It has advantages such as being non-invasive, portable, and safe, but the stimulation parameters are mostly biphasic square wave -30 Hz -1.0 mA - pulse width 0.5. The limitations of traditional percutaneous vagus nerve stimulation (cVNS) include limited overall efficacy, a small effective stimulation point due to the limited area of ​​the auricle, inability to precisely control stimulation intensity and range, skin irritation discomfort, and significant individual variability. Cervical percutaneous vagus nerve stimulation (cVNS), as a technique between invasive and non-invasive methods, has gained attention in recent years. However, a key problem with traditional percutaneous stimulation is that electrical stimulation inevitably activates efferent fibers along with afferent fibers, leading to peripheral side effects such as decreased heart rate and intestinal motility inhibition, limiting the safety and effectiveness of clinical applications. Furthermore, conductive hydrogels, as a novel biofunctional material combining the excellent biocompatibility of hydrogels and the electrical properties of conductive polymers, show broad application prospects in tissue engineering, wearable electronics, and neural interfaces. However, existing conductive hydrogels still face challenges in minimally invasive nerve stimulation applications, including insufficient injection performance, poor shear thinning and self-repair properties, the need for systematic verification of biocompatibility and degradation behavior, and insufficient electrical compatibility with nerve tissue. Therefore, how to achieve selective activation of vagus nerve afferent fibers without increasing surgical trauma, while simultaneously constructing a stable and reliable stimulation-nerve interface, has become a pressing technical problem to be solved in this field. Summary of the Invention

[0005] This invention provides a vagus nerve activation system, a construction method, a conductive hydrogel, and its applications to solve the above-mentioned problems.

[0006] This invention discloses a vagus nerve activation system, characterized in that it comprises:

[0007] A high-frequency electrical stimulation generation module is used to generate high-frequency biphasic square wave pulse electrical stimulation signals; An injectable conductive hydrogel module for forming a local conductive interface around the vagus nerve in the neck, the injectable conductive hydrogel module containing PEDOT:PSS, N,O-carboxymethyl chitosan (NOCC), and aldehyde-modified hyaluronic acid (HA-CHO); and An ultrasound-guided positioning module is used to precisely deliver the injectable conductive hydrogel module to a target site around the vagus nerve in the neck. The electrical stimulation signal generated by the high-frequency electrical stimulation generation module is transmitted to the vagus nerve through the local conductive interface, thereby achieving selective activation of the afferent fibers of the vagus nerve.

[0008] Optionally, the high-frequency electrical stimulation signal parameters are: frequency 5 kHz, intensity 0.5-2.0 mA, pulse width 0.05-0.2 ms, and stimulation mode of cyclic stimulation of 30 s ON-30 s OFF. The mass fraction of PEDOT:PSS in the injectable conductive hydrogel module is 15-35%, and the mass ratio of NOCC to HA-CHO is 1:0.5 to 1:2.

[0009] This invention also discloses a conductive hydrogel for vagal nerve electrical stimulation, comprising: Conductive component: PEDOT:PSS, mass fraction 25%; Biocompatible backbone component: N,O-carboxymethyl chitosan (NOCC), concentration 1.2% wt / v; Functional modification component: Aldehyde-modified hyaluronic acid (HA-CHO), concentration 3%wt / v; In this process, the amino group on NOCC and the aldehyde group on HA-CHO form an imine bond cross-linking network through a Schiff base reaction, and PEDOT:PSS is uniformly distributed in the cross-linking network to form nanoscale conductive channels.

[0010] Optionally, the hydrogel has a three-dimensional porous network structure with an average pore size of 50-200 μm, a porosity of 85-92%, an electrical conductivity of 0.5-8 S / m, and a gelation time of 8-12 seconds.

[0011] This invention also discloses a method for constructing a selective activation system for vagal nerve afferent fibers, comprising the following steps: (a) Preparation of an injectable conductive hydrogel precursor solution containing PEDOT:PSS, N,O-carboxymethyl chitosan (NOCC) and aldehyde-modified hyaluronic acid (HA-CHO). (b) Under ultrasound guidance, the precursor solution is delivered via an injection needle to a target site around the vagus nerve in the neck; (c) The precursor solution gels in situ in the tissue environment through Schiff base reaction, forming a local conductive interface that surrounds the vagus nerve; (d) The percutaneous electrode of the high-frequency electrical stimulation generation module is placed on the body surface corresponding to the local conductive interface, and a biphasic square wave pulse electrical stimulation signal with a frequency of 5 kHz is applied. The electrical stimulation signal is transmitted to the vagus nerve through the local conductive interface.

[0012] Optionally, the final mass fraction of PEDOT:PSS in the precursor solution in step (a) is 25%, and the volume ratio of NOCC to HA-CHO is 1:1; the electrical stimulation parameters in step (d) are: frequency 5kHz, intensity 0.5-2.0 mA, pulse width 0.05-0.2 ms, and stimulation mode of cyclic stimulation of 30 s ON-30 s OFF.

[0013] The present invention also discloses the application of the system described above in the fabrication of a neuromodulation device.

[0014] The present invention also discloses the application of the composite conductive hydrogel as described above in the preparation of conductive interface materials for nerve electrical stimulation.

[0015] The present invention also discloses the application of the method described above in the preparation of a minimally invasive neurostimulation system.

[0016] The present invention also discloses the application of the above-described composite conductive hydrogel in the preparation of injectable biomedical materials.

[0017] The beneficial effects of the vagus nerve activation system provided in this invention are as follows: In terms of stimulation strategy, the invention achieves preferential activation of vagus nerve afferent fibers through 5kHz high-frequency electrical stimulation, keeping the heart rate variability rate within 5% and significantly improving safety; in terms of material system, it adopts a PEDOT:PSS / NOCC / HA-CHO ternary composite conductive hydrogel, which combines injectability, in-situ gelation properties, and good biocompatibility; in terms of system construction, it achieves minimally invasive operation through ultrasound-guided injection combined with percutaneous electrodes, avoiding surgical trauma and improving patient compliance; in terms of synergistic effect, the high-frequency stimulation strategy and the conductive hydrogel interface work together to reduce side effects while retaining the awakening effect. Attached Figure Description

[0018] The technical solution of the present invention will be further described in detail below with reference to the accompanying drawings and embodiments. In the accompanying drawings: Figure 1 Figures A and B are schematic diagrams illustrating the preparation of PEDOT:PSS / NOCC / HA-CHO according to the present invention; Figure 2 This is a schematic diagram of the microstructure of PEDOT:PSS / NOCC / HA-CHO of the present invention; Figure 3A -H is a schematic diagram of the physical and chemical characterization of PEDOT:PSS / NOCC / HA-CHO of the present invention; Figure 4A -D is a schematic diagram of the shear properties (A), adhesion (B), and injectability (C / D) of PEDOT:PSS / NOCC / HA-CHO of this invention; Figure 5 AF is a schematic diagram of the biosafety verification of PEDOT:PSS / NOCC / HA-CHO of this invention; Figure 6 AB is a schematic diagram of the c-Fos expression characteristics of NTS neurons after 5 kHz-cVNS intervention in Experiment 1 of this invention; Figure 7A -D is a safety assessment of peripheral autonomic nerve function in mice after 5 kHz-cVNS intervention; Figure 8A -B represents the dynamic changes in calcium signaling in NTS neurons after 5 kHz-cVNS intervention; Figure 9A -B is an in vivo biocompatibility evaluation of PNH conductive hydrogel. Figure 10A -C represents the effect of different stimulation parameters on changes in mouse heart rate. Detailed Implementation

[0019] It should be noted that, unless otherwise specified, the embodiments and features described in this application can be combined with each other. The preferred embodiments of the present invention will now be described in detail with reference to the accompanying drawings.

[0020] This invention provides a vagus nerve activation system, comprising a high-frequency electrical stimulation generation module, an injectable conductive hydrogel module, and an ultrasound-guided positioning module. The high-frequency electrical stimulation generation module generates a high-frequency (5 kHz) biphasic square wave pulse electrical stimulation signal. The injectable conductive hydrogel module forms a local conductive interface around the cervical vagus nerve, and the injectable conductive hydrogel module contains PEDOT:PSS, N,O-carboxymethyl chitosan (NOCC), and aldehyde-modified hyaluronic acid (HA-CHO). The ultrasound-guided positioning module precisely delivers the injectable conductive hydrogel module to the target site around the cervical vagus nerve. The electrical stimulation signal generated by the high-frequency electrical stimulation generation module is transmitted to the vagus nerve through the local conductive interface, thereby achieving selective activation of the afferent fibers of the vagus nerve.

[0021] This invention achieves selective activation of vagal nerve afferent fibers through high-frequency (5 kHz) electrical stimulation, while effectively avoiding activation of efferent fibers. Screening criteria (heart rate variability <5%) ensure no significant cardiac side effects, thus significantly reducing peripheral side effects such as decreased heart rate. The mechanism involves the different activation thresholds of different types of nerve fibers under high-frequency stimulation. Vagal nerve afferent fibers are mainly distributed in the superficial layer of the epineurium and are more easily activated by high-frequency electric fields, while efferent fibers are mainly concentrated in the central region of the nerve and respond less strongly to high-frequency stimulation. Therefore, 5 kHz electrical stimulation can achieve selective and preferential activation of afferent fibers while effectively avoiding activation of efferent fibers.

[0022] To verify the selective activation effect and ensure safety, this invention uses a heart rate change rate of less than 5% as a screening criterion to exclude significant cardiac effects. Multidimensional verification is performed using three methods: vagus nerve transection experiment, solitary tract nucleus calcium imaging, and c-Fos immunofluorescence. The vagus nerve transection experiment distinguishes the independent roles of afferent and efferent fibers, solitary tract nucleus calcium imaging monitors central neuron activation in real time, and c-Fos immunofluorescence labels immediate early gene expression in neurons after stimulation. This comprehensive approach confirms the selective activation mechanism and safety of 5kHz high-frequency stimulation.

[0023] Compared with existing technologies, this invention increases the stimulation frequency from the traditional 30 Hz to 5 kHz, maintains the same and effective afferent fiber activation effect as traditional technologies, significantly inhibits efferent fiber activation instead of the significant activation of traditional technologies, reduces the decrease in heart rate from 10% to 15% to less than 5% in traditional technologies, eliminates the significant effect on intestinal motility inhibition instead of the significant effect of traditional technologies, maintains the same and effective wakefulness effect as traditional technologies, and significantly improves overall safety from the general level of traditional technologies.

[0024] The optimal parameters for the high-frequency electrical stimulation signal are a biphasic square wave pulse with a frequency of 5 kHz, an intensity of 1.0 mA, a pulse width of 0.1 ms, and a stimulation mode of cyclic stimulation consisting of continuous stimulation for 30 s followed by a 30 s interval.

[0025] This invention employs a PEDOT:PSS / NOCC / HA-CHO (hereinafter referred to as PNH) ternary composite conductive hydrogel as an injectable conductive hydrogel module. In the PNH hydrogel, PEDOT:PSS serves as the conductive component, providing electrical conduction; N,O-carboxymethyl chitosan (NOCC) serves as the biocompatible framework component, providing a cross-linking network and biocompatibility; and aldehyde-modified hyaluronic acid (HA-CHO) serves as the functional modification component, participating in the Schiff base reaction and enhancing material properties. The PNH hydrogel constructs a locally conductive interface through the following method: under ultrasound guidance, the PNH hydrogel precursor solution is delivered via an injection needle to a target site around the vagus nerve in the neck; combined with… Figure 1 and Figure 2 As shown, the amino groups in NOCC and the aldehyde groups in HA-CHO in the precursor solution undergo a Schiff base reaction in the tissue environment to generate imine bonds and form a three-dimensional network structure, which crosslinks in situ into a gel within 5 to 20 seconds. The gelled PNH hydrogel wraps around the vagus nerve, forming a local conductive interface that is closely attached to the nerve tissue. The electrical stimulation signal generated by the high-frequency electrical stimulation generation module is transmitted to the vagus nerve through this local conductive interface to achieve long-term stable nerve stimulation.

[0026] PEDOT:PSS is a composite conductive polymer of poly(3,4-ethylenedioxythiophene) and polystyrene sulfonic acid. PEDOT serves as the conductive polymer backbone, providing charge transport channels, while PSS acts as a water-soluble anionic polymer dopant and dispersant, ensuring stable dispersion of PEDOT in aqueous solution. This composite material possesses high conductivity, optical transparency, aqueous solution processability, and good biocompatibility. In this invention, a commercially available PH1000 type PEDOT:PSS dispersion is used as the conductive component of the PNH hydrogel, uniformly distributed at a mass fraction of 15% to 35% within the three-dimensional network of the hydrogel to form nanoscale conductive channels. This results in an overall conductivity of 0.5 to 8 S / m for the hydrogel, thereby enabling efficient transmission of electrical stimulation signals from transcutaneous electrodes to the vagus nerve.

[0027] Specifically, in the injectable conductive hydrogel module, the mass fraction of PEDOT:PSS is 15-35%, and the mass ratio of NOCC to HA-CHO is 1:0.5 to 1:2. The mass fraction of PEDOT:PSS can be 15%, 20%, 25%, 30%, or 35%, and the mass ratio of NOCC to HA-CHO can be 1:0.5, 1:0.8, 1:1, 1:1.2, 1:1.5, 1:1.8, or 1:2.

[0028] Furthermore, this invention also discloses a conductive hydrogel for vagal nerve electrical stimulation, comprising a conductive component, a biocompatible framework component, and a functional modification component. The conductive component is PEDOT:PSS, with a mass fraction of 15-35%; the biocompatible framework component is N,O-carboxymethyl chitosan (NOCC), with a concentration of 1-3% wt / v; and the functional modification component is aldehyde-modified hyaluronic acid (HA-CHO), with a concentration of 1-3% wt / v. The amino groups on NOCC and the aldehyde groups on HA-CHO form an imine bond crosslinking network through a Schiff base reaction, and PEDOT:PSS is uniformly distributed within this crosslinking network to form nanoscale conductive channels.

[0029] The PEDOT:PSS conductive component used in this invention is derived from commercial PH1000 type PEDOT:PSS dispersion, with a solid content of 1.0%-1.3%. The optimal addition ratio in the hydrogel is 25% by mass. This component is uniformly distributed in the three-dimensional network of the hydrogel to form nanoscale conductive channels, thereby providing electrical conduction function for the transmission of electrical stimulation signals from the transcutaneous electrode to the vagus nerve.

[0030] The NOCC (N,O-carboxymethyl chitosan) used in this invention is a natural polysaccharide derivative with non-immunogenicity, good biocompatibility and biodegradability. In the hydrogel system, it serves as a biocompatible backbone component, providing a cross-linked network structure and biocompatibility.

[0031] The HA-CHO (aldehyde-modified hyaluronic acid) used in this invention is a chemically modified product of hyaluronic acid. It has good biocompatibility and can participate in Schiff base reactions. In the hydrogel system, it acts as a functional modifying component and works synergistically with NOCC to form a three-dimensional cross-linked network and improve the overall performance of the material.

[0032] This invention innovatively achieves in-situ spontaneous cross-linking of the conductive hydrogel through a Schiff base reaction. Specifically, the amino groups on NOCC react with the aldehyde groups on HA-CHO to generate imine bonds and form a three-dimensional network structure. The gelation time is approximately 10 seconds, thus achieving rapid in-situ fixation. The conductive hydrogel has the following performance characteristics: a three-dimensional porous network structure with uniform pore size distribution; rheological properties including injectability, shear thinning, and self-healing; good biocompatibility, with no significant damage to vagus nerve function after 4 weeks of implantation; excellent electrical properties, with an adjustable conductivity range of 0.1 to 10 S / m; and controllable degradation performance, capable of in vivo degradation with an adjustable degradation cycle.

[0033] Specifically, the conductive hydrogel has a three-dimensional porous network structure with an average pore size of 50-200 μm and a porosity of 85-92%, making it suitable for cell infiltration and nutrient transport; its electrical conductivity is 0.5-8 S / m, and its gelation time is 8-12 seconds. PEDOT:PSS is uniformly distributed in the hydrogel network, forming nanoscale conductive channels.

[0034] In practical applications, the conductive hydrogel is used to precisely locate the anatomical position of the vagus nerve in the neck under ultrasound guidance. The conductive hydrogel precursor solution is injected into the target site around the vagus nerve through an injection needle. The precursor solution undergoes a Schiff base reaction in the tissue environment and gels in situ within 10 seconds, forming a local conductive interface that surrounds the vagus nerve. The external percutaneous electrode efficiently transmits the electrical stimulation signal to the vagus nerve through this local conductive interface, achieving selective activation of the afferent fibers.

[0035] Compared with existing technologies, this invention employs a strategy of combining 5kHz high-frequency electrical stimulation with the conductive hydrogel of this application. In terms of the arousal effect, traditional 30Hz-cVNS can shorten the righting reflex recovery time, increase the motor score AUC, and adjust arousal-related EEG activity. This invention also achieves and retains the shortened righting reflex recovery time, increased motor score AUC, and adjusted arousal-related EEG activity. In terms of safety, traditional 30Hz-cVNS has a significant decrease in heart rate (10% to 15%) and significant intestinal motility inhibition. This invention reduces the decrease in heart rate to less than 5% and improves intestinal motility inhibition to no significant effect. The overall safety is significantly improved from the general level of traditional technologies.

[0036] Therefore, this invention has the following technical effects: It achieves selective activation of vagal nerve afferent fibers by employing a 5kHz high-frequency electrical stimulation strategy, significantly reducing side effects such as decreased heart rate and intestinal motility inhibition caused by efferent fiber activation; it achieves minimally invasive operation through ultrasound-guided injection and in-situ gelation technology, avoiding the trauma and complications of traditional cervical incision surgery; it improves stimulation efficiency and long-term stability by forming a uniform conductive interface around the vagus nerve using PNH conductive hydrogel; it achieves good biocompatibility by using a PEDOT:PSS / NOCC / HA-CHO ternary composite conductive hydrogel system, with no significant damage to nerve function within 4 weeks of implantation; it achieves adjustable performance of parameters such as conductivity and degradation cycle by controlling the proportion of each component to adapt to different clinical needs; and it achieves a balance between safety and effectiveness by reducing side effects while retaining the arousal effect of vagal nerve stimulation.

[0037] This invention also discloses a method for constructing a selective activation system for vagal nerve afferent fibers, comprising the following steps: (a) Preparation of an injectable conductive hydrogel precursor solution containing PEDOT:PSS, N,O-carboxymethyl chitosan (NOCC) and aldehyde-modified hyaluronic acid (HA-CHO). (b) Under ultrasound guidance, the precursor solution is delivered via an injection needle to a target site around the vagus nerve in the neck; (c) The precursor solution gels in situ in the tissue environment through Schiff base reaction, forming a local conductive interface that surrounds the vagus nerve; (d) The percutaneous electrode of the high-frequency electrical stimulation generation module is placed on the body surface corresponding to the local conductive interface, and a biphasic square wave pulse electrical stimulation signal with a frequency of 5 kHz is applied. The electrical stimulation signal is transmitted to the vagus nerve through the local conductive interface.

[0038] Specifically, in step (a), the final mass fraction of PEDOT:PSS in the precursor solution is 25%, and the volume ratio of NOCC to HA-CHO is 1:1; the electrical stimulation parameters in step (d) are: frequency 5kHz, intensity 0.5-2.0 mA, pulse width 0.05-0.2 ms, and stimulation mode of cyclic stimulation of 30 s ON-30 s OFF.

[0039] The present invention also discloses the application of the system described above in the fabrication of a neuromodulation device.

[0040] The present invention also discloses the application of the composite conductive hydrogel as described above in the preparation of conductive interface materials for nerve electrical stimulation.

[0041] The present invention also discloses the application of the method described above in the preparation of a minimally invasive neurostimulation system.

[0042] The present invention also discloses the application of the above-described composite conductive hydrogel in the preparation of injectable biomedical materials.

[0043] The technical solution of the present invention will be further described in detail below through embodiments.

[0044] Example 1: Preparation of PNH conductive hydrogel The source and specifications of the materials are shown in Table 1: Table 1

[0045] Preparation steps Step 1: NOCC solution preparation. Dissolve an appropriate amount of NOCC powder in deionized water and stir magnetically for 2 hours (500 rpm) to prepare a homogeneous solution of 1.2% (wt / v). Sterilize by filtration through a 0.22 μm filter membrane.

[0046] Step 2: Preparation of HA-CHO solution. Dissolve an appropriate amount of HA-CHO powder in PBS buffer (pH 7.4), stir magnetically for 1 hour (400 rpm), and prepare a 3% (wt / v) homogeneous solution. Filter and sterilize using a 0.22 μm filter membrane.

[0047] Step 3: Pretreatment of PEDOT:PSS solution. Take a commercially available pH 1000 dispersion and sonicate it for 10 minutes (100W power, 40 kHz frequency) to remove air bubbles.

[0048] Step 4: Preparation of ternary mixed solution. Measure NOCC solution and HA-CHO solution (volume ratio 1:1) according to the ratio, mix and stir magnetically for 10 minutes, add PEDOT:PSS solution (final PEDOT:PSS content 25%), continue stirring for 5 minutes, vortex mix for 2 minutes, and centrifuge to remove bubbles (3000 rpm, 3 minutes).

[0049] Step 5: Gel formation verification. Take a small amount of the mixture and drop it onto a glass plate, observe the gel formation time (about 10 seconds), and perform rheological property testing.

[0050] The performance test results are shown in Table 2 below.

[0051] Table 2

[0052] Example 2: Optimization of High-Frequency Vagus Nerve Stimulation Parameters Determining the optimal parameters: Figure 10A -C represents the effect of different stimulation parameters on changes in mouse heart rate. Figure 10A The effect of different stimulation frequencies on the rate of change of heart rate in mice under the conditions of fixed square wave and 1.0 mA; Figure 10B The effect of different stimulation waveforms on the rate of change of heart rate in mice under the conditions of fixed 5 kHz and 1.0 mA; Figure 10C The effect of different stimulus intensities on the rate of change of heart rate in mice was investigated under the conditions of a fixed 5 kHz and square wave. The vertical axis represents the rate of change of heart rate, and the red dashed line represents the screening threshold of ±5% heart rate change. The results showed that under the conditions of frequency ≥ 5 kHz, square wave, and intensity ≤ 1.0 mA, the rate of change of heart rate in mice was basically maintained within ±5%. Based on the above results, square wave - 5 kHz - 1.0 mA was determined as the candidate stimulus parameters for subsequent validation experiments. The specific stimulus parameters are shown in Table 3. Table 3

[0053] Selective verification experiments: Experiment 1: Activation of the NTS in the afferent nuclei of the vagus nerve. The left cervical vagus nerve of mice was exposed under anesthesia and randomly divided into three groups: sham operation group, afferent fiber transection group, and efferent fiber transection group. Each group was given electrical stimulation at 5 kHz and 30 Hz, respectively, and the c-Fos expression level in the NTS region was recorded.

[0054] Figure 6 The c-Fos expression characteristics of NTS neurons after 5 kHz-cVNS intervention. A: IF staining images of NTS c-Fos in mice in the control and 5 kHz-cVNS groups (scale bar: 100 μm for low power field, 25 μm for high power field); B: Statistical count of total c-Fos positive neurons in NTS in the control and 5 kHz-cVNS groups (n=6, P<0.001).

[0055] Figure 7A -D is a safety assessment of the peripheral autonomic nervous system in mice after 5 kHz-cVNS intervention. Figure 7A Raw echocardiographic recordings of mice before (top) and during (bottom) 5 kHz-cVNS stimulation; Figure 7B Comparison of heart rate fluctuation amplitude during intervention between Control and 5 kHz-cVNS group mice (n=6, ns, P>0.05). Figure 7C Comparison of steel ball expulsion time between Control and 5 kHz-cVNS mice (n=6, ns, P>0.05); Figure 7D The time to first defecation was compared between the Control group and the 5 kHz-cVNS group (n=6, ns, P>0.05).

[0056] Results: During the 5 kHz-cVNS intervention, there were no statistically significant differences in heart rate fluctuation amplitude, steel ball expulsion time, and first defecation time between the mice and the sham stimulation group (P>0.05, marked as "ns"). No adverse reactions such as sudden drop in heart rate or intestinal motility disorder occurred. However, after the conventional 30 Hz-cVNS intervention, the mice showed a significant decrease in heart rate and abnormal activation of intestinal motility, with obvious side effects mediated by non-selective activation of vagal nerve efferent fibers.

[0057] Experiment 2: NTS calcium imaging verification. Cre-dependent GCaMP6s transgenic rats were constructed, and AAV-hSyn-Cre was stereotactically injected into the NTS. In vivo calcium imaging was used to record NTS neuronal activity during 5 kHz stimulation.

[0058] Figure 8A -B represents the dynamic changes in calcium signaling in NTS neurons after 5 kHz-cVNS intervention. Figure 8A The curves show the average calcium signal (ΔF / F) over time during the 5 kHz stimulation period and the stimulation of NTS neurons. The red shaded area represents the signal fluctuation range (Stimulating: 0-30 s; Post-stimulation: 30-60 s). Figure 8B This corresponds to a calcium imaging thermogram, with colors ranging from blue to red representing increases in neuronal calcium signal intensity, visually demonstrating the spatiotemporal characteristics of NTS neuron activation induced by 5 kHz stimulation.

[0059] Results: 5 kHz stimulation can effectively activate neurons in the NTS area.

[0060] Safety assessment: Four weeks after the hydrogel was injected into the area around the vagus nerve in the neck, samples were taken. The assessment included: evaluating the tissue response of major organs such as the heart, liver, spleen, lungs and kidneys by hematoxylin-eosin staining; evaluating the integrity of vagus nerve fibers by neurofilament protein immunofluorescence; and observing the interface between the PNH hydrogel and nerve tissue by electron microscopy.

[0061] Figure 9A -B represents the in vivo biocompatibility evaluation of the PNH conductive hydrogel. Figure 9A Comparison of heart rate variability between the WEI control group and the PNH hydrogel group; Figure 9B HE staining results of major organs in two groups of mice (scale bar: 200 μm).

[0062] Results: HE staining showed that the heart, liver, spleen, lungs and kidneys of mice in the PNH hydrogel group had intact structures and no obvious toxic damage such as cell degeneration, necrosis or inflammatory infiltration. There was no significant difference compared with the control group, suggesting that PNH hydrogel did not show obvious systemic toxicity under the conditions of this study and has good in vivo biocompatibility.

[0063] In summary, this invention achieves preferential activation of vagal nerve afferent fibers through 5 kHz high-frequency electrical stimulation, keeping the heart rate variability within 5% and significantly improving safety. Regarding the material system, it employs a PEDOT:PSS / NOCC / HA-CHO ternary composite conductive hydrogel, which combines injectability, in-situ gelation properties, and good biocompatibility. In terms of system construction, ultrasound-guided injection combined with percutaneous electrodes enables minimally invasive procedures, avoiding surgical trauma and improving patient compliance. Regarding synergistic effects, the high-frequency stimulation strategy and the conductive hydrogel interface work together to retain the arousal effect while reducing side effects.

[0064] Compared with existing traditional invasive cVNS techniques, the system of this invention significantly reduces surgical trauma from cervical incision to puncture with a 22-25G injection needle; reduces heart rate decline from 10-15% to less than 5%, an improvement of over 70%; eliminates intestinal inhibition from significant improvement to no significant impact; shortens operation time from 1-2 hours of surgery to less than 30 minutes of injection, a reduction of 80%; significantly reduces infection risk from medium-high risk to extremely low risk; significantly improves patient acceptance from low to high level; and retains the same awakening effect as traditional invasive cVNS.

[0065] It should be understood that the above embodiments are only used to illustrate the technical solutions of the present invention, and are not intended to limit them. Those skilled in the art can modify the technical solutions described in the above embodiments, or make equivalent substitutions for some of the technical features; and all such modifications and substitutions should fall within the protection scope of the appended claims of the present invention.

Claims

1. A vagus nerve activation system, characterized in that, include: A high-frequency electrical stimulation generation module is used to generate high-frequency biphasic square wave pulse electrical stimulation signals; An injectable conductive hydrogel module for forming a local conductive interface around the vagus nerve in the neck, the injectable conductive hydrogel module containing PEDOT:PSS, N,O-carboxymethyl chitosan (NOCC), and aldehyde-modified hyaluronic acid (HA-CHO); and An ultrasound-guided positioning module is used to precisely deliver the injectable conductive hydrogel module to a target site around the vagus nerve in the neck. The electrical stimulation signal generated by the high-frequency electrical stimulation generation module is transmitted to the vagus nerve through the local conductive interface, thereby achieving selective activation of the afferent fibers of the vagus nerve.

2. The system according to claim 1, characterized in that, The high-frequency electrical stimulation signal parameters are: frequency 5 kHz, intensity 0.5-2.0 mA, pulse width 0.05-0.2 ms, and stimulation mode of cyclic stimulation of 30 s ON-30 s OFF. The injectable conductive hydrogel module contains 15-35% PEDOT:PSS by mass, and the mass ratio of NOCC to HA-CHO is 1:0.5 to 1:

2.

3. A conductive hydrogel for vagal nerve electrical stimulation, characterized in that, include: Conductive component: PEDOT:PSS, mass fraction 25%; Biocompatible backbone component: N,O-carboxymethyl chitosan (NOCC), concentration 1.2% wt / v; Functional modification component: Aldehyde-modified hyaluronic acid (HA-CHO), concentration 3%wt / v; In this process, the amino group on NOCC and the aldehyde group on HA-CHO form an imine bond cross-linking network through a Schiff base reaction, and PEDOT:PSS is uniformly distributed in the cross-linking network to form nanoscale conductive channels.

4. The conductive hydrogel according to claim 3, characterized in that, The hydrogel has a three-dimensional porous network structure with an average pore size of 100 μm, a porosity of 90%, an electrical conductivity of 4 S / m, and a gelation time of 10 seconds.

5. A method for constructing a selective activation system for vagal nerve afferent fibers, characterized in that, Includes the following steps: (a) Preparation of an injectable conductive hydrogel precursor solution containing PEDOT:PSS, N,O-carboxymethyl chitosan (NOCC) and aldehyde-modified hyaluronic acid (HA-CHO). (b) Under ultrasound guidance, the precursor solution is delivered via an injection needle to a target site around the vagus nerve in the neck; (c) The precursor solution gels in situ in the tissue environment through Schiff base reaction, forming a local conductive interface that surrounds the vagus nerve; (d) The percutaneous electrode of the high-frequency electrical stimulation generation module is placed on the body surface corresponding to the local conductive interface, and a biphasic square wave pulse electrical stimulation signal with a frequency of 5 kHz is applied. The electrical stimulation signal is transmitted to the vagus nerve through the local conductive interface.

6. The method according to claim 5, characterized in that, In step (a), the final mass fraction of PEDOT:PSS in the precursor solution is 25%, and the volume ratio of NOCC to HA-CHO is 1:1; the electrical stimulation parameters in step (d) are: frequency 5kHz, intensity 0.5-2.0 mA, pulse width 0.05-0.2ms, and stimulation mode of cyclic stimulation of 30 s ON-30 s OFF.

7. The application of the system as described in claim 1 or 2 in the fabrication of a neuromodulation device.

8. The application of a composite conductive hydrogel as described in claim 3 or 4 in the preparation of conductive interface materials for nerve electrical stimulation.

9. The application of the method as described in claim 5 or 6 in the preparation of a minimally invasive neurostimulation system.

10. The application of a composite conductive hydrogel as described in claim 3 or 4 in the preparation of injectable biomedical materials.

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