Study on influence of external factors on the electrical excitability of PC12 quasi-neuronal networks through Voltage Threshold Measurement Method.

The aim of this paper was to investigate the influence of four different external factors (acetylcholine, ethanol, temperature and lidocaine hydrochloride) on PC12 quasi-neuronal networks by multielectrode-array-based Voltage Threshold Measurement Method (VTMM). At first, VTMM was employed to measure the lowest amplitude of the voltage stimulating pulses that could just trigger the action potential from PC12 quasi-neuronal networks under normal conditions, and the amplitude was defined as the normal voltage threshold (VTh). Then the changes of the VTh of PC12 quasi-neuronal networks treated by the four external factors were tested respectively. The results showed the normal VTh of PC12 quasi-neuronal networks was 36 mV. The VTh has negative correlation with the concentration of acetylcholine and has positive correlation with the concentration of ethanol. The curves of the correlation of the VTh with temperature and the concentration of lidocaine hydrochloride were U-shaped and Λ-shaped respectively. Comparing with our earlier studies on hippocampal neuronal networks and hippocampal slices, PC12 quasi-neuronal networks not only had the same typical voltage threshold characteristic, but also had similar changes on electrical excitability when treated by the four external factors mentioned above. Therefore, the rapid-formed PC12 quasi-neuronal networks could replace neuronal networks in proper conditions, and VTMM could be used to analyze the influence of external factors on the electrical excitability of PC12 quasi-neuronal networks.

A flexible electrode array for determining regions of motor function activated by epidural spinal cord stimulation in rats with spinal cord injury.

Epidural stimulation of the spinal cord is a promising technique for the recovery of motor function after spinal cord injury. The key challenges within the reconstruction of motor function for paralyzed limbs are the precise control of sites and parameters of stimulation. To activate lower-limb muscles precisely by epidural spinal cord stimulation, we proposed a high-density, flexible electrode array. We determined the regions of motor function that were activated upon epidural stimulation of the spinal cord in a rat model with complete spinal cord, which was established by a transection method. For evaluating the effect of stimulation, the evoked potentials were recorded from bilateral lower-limb muscles, including the vastus lateralis, semitendinosus, tibialis anterior, and medial gastrocnemius. To determine the appropriate stimulation sites and parameters of the lower muscles, the stimulation characteristics were studied within the regions in which motor function was activated upon spinal cord stimulation. In the vastus lateralis and medial gastrocnemius, these regions were symmetrically located at the lateral site of L1 and the medial site of L2 vertebrae segment, respectively. The tibialis anterior and semitendinosus only responded to stimulation simultaneously with other muscles. The minimum and maximum stimulation threshold currents of the vastus lateralis were higher than those of the medial gastrocnemius. Our results demonstrate the ability to identify specific stimulation sites of lower muscles using a high-density and flexible array. They also provide a reference for selecting the appropriate conditions for implantable stimulation for animal models of spinal cord injury. This study was approved by the Animal Research Committee of Southeast University, China (approval No. 20190720001) on July 20, 2019.

A hybrid method for real-time stimulation artefact removal during functional electrical stimulation with time-variant parameters.

Objective. In this study, a hybrid method combining hardware and software architecture is proposed to remove stimulation artefacts (SAs) and extract the volitional surface electromyography (sEMG) in real time during functional electrical stimulations (FES) with time-variant parameters.Approach. First, an sEMG detection front-end (DFE) combining fast recovery, detector and stimulator isolation and blanking is developed and is capable of preventing DFE saturation with a blanking time of 7.6 ms. The fragment between the present stimulus and previous stimulus is set as an SA fragment. Second, an SA database is established to provide six high-similarity templates with the current SA fragment. The SA fragment will be de-artefacted by a 6th-order Gram-Schmidt (GS) algorithm, a template-subtracting method, using the provided templates, and this database-based GS algorithm is called DBGS. The provided templates are previously collected SA fragments with the same or a similar evoking FES intensity to that of the current SA fragment, and the lengths of the templates are longer than that of the current SA fragment. After denoising, the sEMG will be extracted, and the current SA fragment will be added to the SA database. The prototype system based on DBGS was tested on eight able-bodied volunteers and three individuals with stroke to verify its capacity for stimulation removal and sEMG extraction.Results.The average stimulus artefact attenuation factor, SA index and correlation coefficient between clean sEMG and extracted sEMG for 6th-order DBGS were 12.77 ± 0.85 dB, 1.82 ± 0.37 dB and 0.84 ± 0.33 dB, respectively, which were significantly higher than those for empirical mode decomposition combined with notch filters, pulse-triggered GS algorithm, 1st-order and 3rd-order DBGS. The sEMG-torque correlation coefficients were 0.78 ± 0.05 and 0.48 ± 0.11 for able-bodied volunteers and individuals with stroke, respectively.Significance.The proposed hybrid method can extract sEMG during dynamic FES in real time.

Conjoint analysis of influence of LC-HCL and Mor-HCL on Vth and neurite length in hippocampal neuronal network.

Lidocaine hydrochloride (LC-HCl) and morphine hydrochloride (Mor-HCl) are two kinds of most prevalently used anesthetics. However, their influences on electrical excitability of hippocampal neuronal networks and hippocampal brain slices were rarely studied. Previously, our group have assessed the influence of acetylcholine, alcohol and temperature change on the excitability of neural networks with the so-called Voltage Threshold Measurement Method (VTMM) based on microelectrode array (MEA). In this paper, we will study the influence of LC-HCl and Mor-HCl on the electrical excitability of neural networks and the morphological features of neurons, and discuss the relations between the changes of electrical excitability of neural networks and the morphological changes of neurons. The results of VTMM showed: The voltage threshold (VTh) of hippocampal neuronal networks and hippocampal brain slices first increased and then decreased as the LC-HCl concentration increased. The VTh of hippocampal neuronal networks and hippocampal brain slices increased as the Mor-HCl concentration increased. The results of HCS experiments showed: The neurite length change of cultured hippocampal neuronal networks increased first and then decreased with increased LC-HCl concentration, but decreased as the Mor-HCl concentration increased. The combined analysis of VTMM and HCS experiments showed that under effects of the two drugs, the VTh and the hippocampal neurite length were strongly negatively correlated.

Influence of spinal cord injury on core regions of motor function.

Functional electrical stimulation is an effective way to rebuild hindlimb motor function after spinal cord injury. However, no site map exists to serve as a reference for implanting stimulator electrodes. In this study, rat models of thoracic spinal nerve 9 contusion were established by a heavy-impact method and rat models of T6/8/9 spinal cord injury were established by a transection method. Intraspinal microstimulation was performed to record motion types, site coordinates, and threshold currents induced by stimulation. After transection (complete injury), the core region of hip flexion migrated from the T13 to T12 vertebral segment, and the core region of hip extension migrated from the L1 to T13 vertebral segment. Migration was affected by post-transection time, but not transection segment. Moreover, the longer the post-transection time, the longer the distance of migration. This study provides a reference for spinal electrode implantation after spinal cord injury. This study was approved by the Institutional Animal Care and Use Committee of Nantong University, China (approval No. 20190225-008) on February 26, 2019.

System Design and Experimental Research of Neural Signal Blocking based on Spike Trapping Principle.

Different neural signal blocking technologies provide potential therapy for various diseases caused by pathological or involuntary neural activities. In this paper, a novel neural signal blocking concept based on spike-trapping principle is introduced. The blocking system was designed and implemented. In the system, a compound action potential was induced by a proximal stimulus, and a blocking signal was generated after a neural signal was detected, which was applied to the distal end of the nerve to block the nerve conduction. The reliability of the system was verified on bullfrog sciatic nerve gastrocnemius muscle specimens. The optimal blocking signal parameters, the relationship between nerve diameter and blocking threshold voltage, and the nerve safety of blocking signals were explored. The experimental results show that the system is possible to block a pathological or involuntary neural signal automatically. The average voltage of block threshold and the minimum pulse width are -1.66 V and 120 ms, respectively. The blocking threshold decreases as the diameter of the nerve increases. The blocking signals at the threshold level are safe for the stimulated nerve.

An Anti Stimulation Artifacts and M-waves Surface Electromyography Detector with a Short Blanking Time.

A surface electromyography (sEMG) detector, that not only removes stimulation artifacts entirely but also increases the recording time, has been developed in this paper. The sEMG detector consists of an sEMG detection circuit and a stimulation isolator. The sEMG detection circuit employs a stimulus isolate switch (SIS), a blanking (BLK) and non-linear feed-back (NFB) circuit to remove the artifacts and to increase the recording time. In the SIS, the connection between stimulator and stimulation electrodes, along with the stimulation electrodes and the ground are controlled by an opto-isolator, and the connection of instrument amplifier and the recording electrodes are controlled by CMOS-based switches. The mode switches of the BLK and the NFB circuit also employs CMOS-based switches. By an accurate timing adjustment, the voluntary EMG can be recorded during electrical stimulation. Two 6 able-bodied experiments have been performed to test the three anti-artifact sEMG detector: BLK, BLK&SIS, BLK&SIS&NFB. The results indicate that the BLK&SIS&NFB proposed in this work effectively removes stimulus artifacts and M-waves, and has a longer recording time compared with BLK and BLK&SIS circuits.

Design and Experimental Research of Implantable Lower Esophageal Sphincter Stimulator Based on Android Bluetooth.

The aim of this study is to design an implantable Lower Esophageal Sphincter (LES) stimulator connected and controlled by an Android Bluetooth for the treatment of the gastroesophageal reflux disease (GERD). Then the animal experiments are carried out to evaluate the function of the system. The LES stimulator is composed of an external controller, an Android application (APP) via a smart phone and an implantable electronic device (IED). The external controller is designed to receive the settings parameters information sent by the Android APP via a Bluetooth module, and then is programmed to generate specific electrical stimulation pulses to the LES. The Android APP controls the start and stop of stimulation and the settings of stimulation parameters. The in vivo IED consists of a bipolar stimulating lead, a bipolar head connector and a receiving module. The bipolar stimulating lead is constructed of biocompatible materials: platinum-iridium electrodes which are coated with parylene and an outer silicone rubber sheathing. The size of the receiving module has been significantly decreased to 20×20×2 mm3, which is packaged by polydimethylsiloxane (PDMS) and proposed to deliver stimulation pulses from the external controller to the implantable lead. The one-month implantation experiment on rabbits has been performed to evaluate the LES stimulator. The results indicate that the proposed LES stimulator meets the requirements of the functions, effectiveness and safety.

Experimental Study on Nerve Signals Block for Spasticity Based on Antimissile Strategy.

A method of blocking neural signal for spasticity which is based on the antimissile strategy was proposed. When the pathological nerve action potential signal is detected at the proximal end of the nerve, such a potential signal that is opposite to the signal of the primary neural activity is applied at the distal end of the nerve at a proper delay so as to block the pathological nerve signal. Preliminary tests were performed on toad sciatic nerve-gastrocnemius specimens. Firstly, the effect of the distance between blocking electrodes on the blocking pulse voltage threshold was studied based on the electrical tension induced by the nerve signal on the controlled muscle. Then, the effective parameters of the blocking waveform were studied. Finally, the delay range of the blocking pulse compared to the pathogen action potential was studied. The results showed that in the sciatic nerve-gastrocnemius specimens, the most effective distance between the blocking electrode pairs was 5 mm and the anodic block required an inverted triangle waveform. The voltage threshold of an effective anodic blocking pulse was 1 V and the minimum pulse width was 90 ms. Under the condition of voltage threshold and minimum pulse width, the time shifting value of blocking pulse was greater than 1ms. It is concluded from the study that the spastic action potential caused by the disease can be effectively blocked, and limb muscle spasms can be eliminated under the action of appropriate electrode configuration and blocking signal waveforms.

Quantitative evaluation of extrinsic factors influencing electrical excitability in neuronal networks: Voltage Threshold Measurement Method (VTMM).

The electrical excitability of neural networks is influenced by different environmental factors. Effective and simple methods are required to objectively and quantitatively evaluate the influence of such factors, including variations in temperature and pharmaceutical dosage. The aim of this paper was to introduce 'the voltage threshold measurement method', which is a new method using microelectrode arrays that can quantitatively evaluate the influence of different factors on the electrical excitability of neural networks. We sought to verify the feasibility and efficacy of the method by studying the effects of acetylcholine, ethanol, and temperature on hippocampal neuronal networks and hippocampal brain slices. First, we determined the voltage of the stimulation pulse signal that elicited action potentials in the two types of neural networks under normal conditions. Second, we obtained the voltage thresholds for the two types of neural networks under different concentrations of acetylcholine, ethanol, and different temperatures. Finally, we obtained the relationship between voltage threshold and the three influential factors. Our results indicated that the normal voltage thresholds of the hippocampal neuronal network and hippocampal slice preparation were 56 and 31 mV, respectively. The voltage thresholds of the two types of neural networks were inversely proportional to acetylcholine concentration, and had an exponential dependency on ethanol concentration. The curves of the voltage threshold and the temperature of the medium for the two types of neural networks were U-shaped. The hippocampal neuronal network and hippocampal slice preparations lost their excitability when the temperature of the medium decreased below 34 and 33°C or increased above 42 and 43°C, respectively. These results demonstrate that the voltage threshold measurement method is effective and simple for examining the performance/excitability of neuronal networks.

Electromyographic bridge for promoting the recovery of hand movements in subacute stroke patients: A randomized controlled trial.

OBJECTIVE: The electromyographic bridge (EMGB) detects surface electromyographic signals from a non-paretic limb. It then generates electric pulse trains according to the electromyographic time domain features, which can be used to stimulate a paralysed or paretic limb in real time. This strategy can be used for the contralateral control of neuromuscular electrical stimulation (NMES) to improve motor function after stroke. The aim of this study was to compare the treat-ment effects of EMGB vs cyclic NMES on wrist and finger impairments in subacute stroke patients. METHODS: A total of 42 hemiplegic patients within 6 months of their cerebrovascular accidents were randomly assigned to 4-week treatments with EMGB or cyclic NMES. Each group underwent a standard rehabilitation programme and 10 sessions per week of hand training with EMGB or cyclic NMES. Outcome measures were: Brunnstrom stage, upper extremity components of the Fugl-Meyer Assessment, Motor Status Scale, voluntary surface electromyographic ratio and active range of motion of the wrist and finger joints. RESULTS: The EMGB group showed significantly greater improvements than the cyclic NMES group on the following measures: Brunnstrom stages for the hand, upper extremity - Fugl-Meyer Assessment, Motor Status Scale, and the voluntary surface electromyographic ratio of wrist and finger extensors. Eleven and 4 participants of the EMGB group who had no active wrist and finger movements, respectively, at the start of the treatment could perform measurable wrist and finger extensions after EMGB training. The corresponding numbers in the cyclic NMES group were only 4 and 1. CONCLUSION: In the present group of subacute stroke patients, the results favour EMGB over cyclic NMES for augmenting the recovery of volitional wrist and finger motion.

Real-time and wearable functional electrical stimulation system for volitional hand motor function control using the electromyography bridge method.

Voluntary participation of hemiplegic patients is crucial for functional electrical stimulation therapy. A wearable functional electrical stimulation system has been proposed for real-time volitional hand motor function control using the electromyography bridge method. Through a series of novel design concepts, including the integration of a detecting circuit and an analog-to-digital converter, a miniaturized functional electrical stimulation circuit technique, a low-power super-regeneration chip for wireless receiving, and two wearable armbands, a prototype system has been established with reduced size, power, and overall cost. Based on wrist joint torque reproduction and classification experiments performed on six healthy subjects, the optimized surface electromyography thresholds and trained logistic regression classifier parameters were statistically chosen to establish wrist and hand motion control with high accuracy. Test results showed that wrist flexion/extension, hand grasp, and finger extension could be reproduced with high accuracy and low latency. This system can build a bridge of information transmission between healthy limbs and paralyzed limbs, effectively improve voluntary participation of hemiplegic patients, and elevate efficiency of rehabilitation training.

VANET Clustering Based Routing Protocol Suitable for Deserts.

In recent years, there has emerged applications of vehicular ad hoc networks (VANETs) towards security, safety, rescue, exploration, military and communication redundancy systems in non-populated areas, besides its ordinary use in urban environments as an essential part of intelligent transportation systems (ITS). This paper proposes a novel algorithm for the process of organizing a cluster structure and cluster head election (CHE) suitable for VANETs. Moreover, it presents a robust clustering-based routing protocol, which is appropriate for deserts and can achieve high communication efficiency, ensuring reliable information delivery and optimal exploitation of the equipment on each vehicle. A comprehensive simulation is conducted to evaluate the performance of the proposed CHE and routing algorithms.

A frequency and pulse-width co-modulation strategy for transcutaneous neuromuscular electrical stimulation based on sEMG time-domain features.

OBJECTIVE: Surface electromyography (sEMG) is often used as a control signal in neuromuscular electrical stimulation (NMES) systems to enhance the voluntary control and proprioceptive sensory feedback of paralyzed patients. Most sEMG-controlled NMES systems use the envelope of the sEMG signal to modulate the stimulation intensity (current amplitude or pulse width) with a constant frequency. The aims of this study were to develop a strategy that co-modulates frequency and pulse width based on features of the sEMG signal and to investigate the torque-reproduction performance and the level of fatigue resistance achieved with our strategy. APPROACH: We examined the relationships between wrist torque and two stimulation parameters (frequency and pulse width) and between wrist torque and two sEMG time-domain features (mean absolute value (MAV) and number of slope sign changes (NSS)) in eight healthy volunteers. By using wrist torque as an intermediate variable, customized and generalized transfer functions were constructed to convert the two features of the sEMG signal into the two stimulation parameters, thereby establishing a MAV/NSS dual-coding (MNDC) algorithm. Wrist torque reproduction performance was assessed by comparing the torque generated by the algorithms with that originally recorded during voluntary contractions. Muscle fatigue was assessed by measuring the decline percentage of the peak torque and by comparing the torque time integral of the response to test stimulation trains before and after fatigue sessions. MAIN RESULTS: The MNDC approach could produce a wrist torque that closely matched the voluntary wrist torque. In addition, a smaller decay in the wrist torque was observed after the MNDC-coded fatigue stimulation was applied than after stimulation using pulse-width modulation alone. SIGNIFICANCE: Compared with pulse-width modulation stimulation strategies that are based on sEMG detection, the MNDC strategy is more effective for both voluntary muscle force reproduction and muscle fatigue reduction.

The Principle of the Micro-Electronic Neural Bridge and a Prototype System Design.

The micro-electronic neural bridge (MENB) aims to rebuild lost motor function of paralyzed humans by routing movement-related signals from the brain, around the damage part in the spinal cord, to the external effectors. This study focused on the prototype system design of the MENB, including the principle of the MENB, the neural signal detecting circuit and the functional electrical stimulation (FES) circuit design, and the spike detecting and sorting algorithm. In this study, we developed a novel improved amplitude threshold spike detecting method based on variable forward difference threshold for both training and bridging phase. The discrete wavelet transform (DWT), a new level feature coefficient selection method based on Lilliefors test, and the k-means clustering method based on Mahalanobis distance were used for spike sorting. A real-time online spike detecting and sorting algorithm based on DWT and Euclidean distance was also implemented for the bridging phase. Tested by the data sets available at Caltech, in the training phase, the average sensitivity, specificity, and clustering accuracies are 99.43%, 97.83%, and 95.45%, respectively. Validated by the three-fold cross-validation method, the average sensitivity, specificity, and classification accuracy are 99.43%, 97.70%, and 96.46%, respectively.

Design of a pulse-triggered four-channel functional electrical stimulator using complementary current source and time division multiplexing output method.

In this paper, a four-channel pulse-triggered functional electrical stimulator using complementary current source and time division output method is proposed for the research and application of functional electrical stimulation (FES). The high-voltage compliance and output impedance is increased by adopting the complementary current source, which can also realize the linear voltage-to-current conversion and high channel isolation. A high-voltage analog switch chip MAX14803, combined with a FIFO queue algorithm in the microprocessor, is used to setup the H-bridge and multiplexers for the four-channel time division multiplexing output. With this method, the size and cost of the key components are reduced greatly. The stimulating core circuit area is 30 × 50 mm(2). According to the experiments, the stimulator can achieve the four-channel charge-balanced biphasic stimulation with a current range between 0 and 60 mA and a single-phase pulse amplitude up to 60 V.

[Study on display of acupuncture meridian based on electric signal transfer characteristics].

OBJECTIVE: To analyze the real-time effects of electroacupuncture (EA) stimulation by means of multiport network theory and measurement of the electric signals transfer coefficient, so as to explore a way for determining the running course of acupuncture meridian and for characterizing its physiological activities in the living body. METHODS: The body was modeled as a complex inhomogeneous 3-dimensions multi-port network, and the meridians were assumed to be "the most smoothly channels for signal transmission". Experiments were performed in 12 beheaded toads whose forelimbs and hindlimbs were divided into proportional coordinates. A concentric electrode with a conical tip was inserted into the toad leg for electrical stimulation of the local muscle, and another electrode alike was inserted into different spots of the limbs to detect the spreading signals in a 3-dimensional scanning mode. Following detecting the response electric voltage values of various spots and calculating their transfer coefficients, the spots which acquired a maximum signal value were considered to be the "acupoints". The imaginary connective lines passing the "acupoints" were regarded as the running courses of acupuncture meridians. RESULTS: A total of twelve 3-dimensional curves were detected based on the connected lines of electric signal transfer function extremum spots 2 mm beneath the skin of the ipsilateral fore- and hind-limbs of 12 spinal toads. CONCLUSION: The present study initially validates the feasibility of electric signal transfer coefficient measurement for displaying the running course of acupuncture meridian in the toad fore- and hindlimbs.

Motor function rebuilding of limbs based on communication principle and electronic system.

In this paper, we report our novel idea on the function rebuilding for hemiplegic limbs and the primary experiments. The main concept is to connect the control-lost nerves or neuromuscular junctions by using a multi-channel micro- electronic neural bridge (MENB), regenerate the nervous signal, and rebuild the motor functions of the related limb. Since the injured nervous system in stroke-related hemiplegia is located in the brain and difficult to be identified and operate on, we use another nervous system functioning as a new signal source to supply similar neural signals. In these cases, that means, two independent nervous systems are connected by a MENB. As preclinical experiments, we have made a series of tests on bodies of animals and healthy human. The principle, the system construction and the experimental results will be given.

[Study on signal transmission characteristics of meridian based on electrical network theory and experiments].

Study on features of acupoints with resistance test in the past half century is reviewed in this article. Mechanism and technology of the method are introduced as well as its shortcomings. The determination method of signal transmission along meridians with the combination of electrical network theories and practice is advanced. And the result of a series experiments on one meridian at the superficial part of the body are given as well. Thus, it is concluded that the signals of the point-in/point-out and the signals along a non-meridian path with the same distance are significantly different, which gives a verification of the feasibility of the method by using electrical network theories to set out characteristics of signal transmission along meridians dynamically.

Glutathione S-transferase interacting with far-red insensitive 219 is involved in phytochrome A-mediated signaling in Arabidopsis.

Far-red (FR) insensitive 219 (FIN219) was previously shown to be involved in phytochrome A-mediated FR light signaling. To further understand its function and regulatory relation with other light-signaling components, a yeast two-hybrid approach was used to isolate FIN219-interacting partners. Here, we demonstrate that FIN219-interacting protein 1 (FIP1) interacts with FIN219 in vitro and in vivo and is composed of 217 amino acids that belong to the tau class of the large glutathione S-transferase gene family. FIP1 was further shown to have glutathione S-transferase activity. The gain of function and partial loss of function of FIP1 resulted in a hyposensitive hypocotyl phenotype under continuous FR (cFR) light and a delayed flowering phenotype under long-day conditions, which suggests that FIP1 may exist in a complex to function in the regulation of Arabidopsis (Arabidopsis thaliana) development. In addition, FIP1 mRNA was down-regulated in the suppressor of phytochrome A-105 1 mutant and differentially expressed in constitutive photomorphogenic 1-4 (cop1-4) and cop1-5 mutants under cFR. Intriguingly, FIP1 expression was up-regulated in the fin219 mutant under all light conditions, except cFR. Furthermore, promoter activity assays revealed that FIP1 expression was light dependent, mainly associated with vascular tissues, and developmentally regulated. Subcellular localization studies revealed that the beta-glucuronidase-FIP1 fusion protein was localized in the nucleus and cytoplasm. Taken together, these data indicate that FIP1 may interact with FIN219 to regulate cell elongation and flowering in response to light.