Whisker Movements Research Articles

A new experimental model for studying peripheral nerve regeneration in dual innervated facial reanimation.

Donor nerve selection is a crucial factor in determining clinical outcomes of facial reanimation. Although dual innervation approaches using two neurotizers have shown promise, there is a lack of evidence-based comparison in the literature. Furthermore, no animal model of dual reinnervation has yet been published. This study aimed to establish such a model and verify its technical and anatomical feasibility by performing dual-innervated reanimation approaches in Wistar rats. Fifteen Wistar rats were divided into four experimental groups and one control group. The sural nerve was exposed and used as a cross-face nerve graft (CFNG), which was then anastomosed to the contralateral buccal branch of the facial nerve through a subcutaneous tunnel on the forehead. The CFNG, the masseteric nerve (MN), and the recipient nerve were coapted in one or two stages. The length and width of the utilized structures were measured under an operating microscope. Return of whisker motion was visually confirmed. Nine out of the eleven rats that underwent surgery survived the procedure. Whisker motion was observed in all experimental animals, indicating successful reinnervation. The mean duration of the surgical procedures did not differ significantly between the experimental groups, ensuring similar conditions for all groups. Our experimental study confirmed that the proposed reanimation model in Wistar rats is anatomically and technically feasible, with a high success rate, and shows good prospects for future experiments.

Flexible electronic brush: Real-time multimodal sensing powered by reservoir computing through whisker dynamics.

Multimodal sensing using soft body dynamics plays a crucial role in controlling soft robotic motions. An intriguing application of such soft robot control is to mimic whiskers and digitize soft body motion through whisker dynamics. The challenge herein is to simultaneously monitor the directions, speed, force, and slip information of the whisker motion. The existing whisker-like sensors cannot detect slip information effectively. To address this challenge, this study develops a multitasking electronic brush (e-brush) composed of bundle of whiskers powered by reservoir computing (RC). Four pressure sensors are integrated into the brush to monitor its motion, speed, force, slip, and target surface. These sensors can provide long-term, low-pressure detection as low as 50 pascals, allowing for the precise monitoring of brush movements. A RC algorithm is developed to extract multiple brush motion parameters, including the slip. As a proof of concept for multitasking e-brush, the motion trajectory of handwriting is successfully detected.

The potential role of SCF combined with DPCs in facial nerve repair.

Facial nerve injuries lead to significant functional impairments and psychological distress for affected patients. Effective repair of these injuries remains a challenge. For longer nerve gaps, the regeneration outcomes after nerve grafting remain suboptimal due to limited sources and postoperative immune responses. Tissue engineering techniques are conventional methods for repairing peripheral nerve defects. This study explores the potential of dental pulp cells (DPCs) combined with stem cell factor (SCF) to enhance neurogenic differentiation and improve facial nerve regeneration. DPCs were isolated from rabbit dental pulp, the pluripotency of the cells was identified from three perspectives: osteogenic differentiation, adipogenic differentiation, and neurogenic differentiation. In vivo experiments involved injuring the buccal branch of the facial nerve in New Zealand white rabbits, followed by treatment with PBS, DPCs, SCF, or SCF + DPCs. Functional recovery was assessed over 12weeks, with SCF + DPCs demonstrating the most significant improvement in whisker movement scores. Histomorphological evaluations revealed enhanced myelinated fiber density and axonal morphology in the SCF + DPCs group. RNA sequencing identified 608 differentially expressed genes, with enrichment in the TGF-β signaling pathway. In in vitro experiments, we demonstrated from multiple angles using Western blot analysis, Real-time quantitative polymerase chain reaction (QPCR) analysis, and immunofluorescence staining that SCF can promote the neurogenic differentiation of DPCs through the TGF-β1 signaling pathway. Our findings indicate that the combination of SCF and DPCs offers a promising strategy for enhancing facial nerve repair.

GlyT2-Positive Interneurons Regulate Timing and Variability of Information Transfer in a Cerebellar-Behavioral Loop.

GlyT2-positive interneurons, Golgi and Lugaro cells, reside in the input layer of the cerebellar cortex in a key position to influence information processing. Here, we examine the contribution of GlyT2-positive interneurons to network dynamics in Crus 1 of mouse lateral cerebellar cortex during free whisking. We recorded neuronal population activity using Neuropixels probes before and after chemogenetic downregulation of GlyT2-positive interneurons in male and female mice. Under resting conditions, cerebellar population activity reliably encoded whisker movements. Reductions in the activity of GlyT2-positive cells produced mild increases in neural activity which did not significantly impair these sensorimotor representations. However, reduced Golgi and Lugaro cell inhibition did increase the temporal alignment of local population network activity at the initiation of movement. These network alterations had variable impacts on behavior, producing both increases and decreases in whisking velocity. Our results suggest that inhibition mediated by GlyT2-positive interneurons primarily governs the temporal patterning of population activity, which in turn is required to support downstream cerebellar dynamics and behavioral coordination.

Enhancing facial nerve regeneration with scaffold-free conduits engineered using dental pulp stem cells and their endogenous, aligned extracellular matrix.

Objective. Engineered nerve conduits must simultaneously enhance axon regeneration and orient axon extension to effectively restore function of severely injured peripheral nerves. The dental pulp contains a population of stem/progenitor cells that endogenously express neurotrophic factors (NTFs), growth factors known to induce axon repair. We have previously generated scaffold-free dental pulp stem/progenitor cell (DPSC) sheets comprising an aligned extracellular matrix (ECM). Through the intrinsic NTF expression of DPSCs and the topography of the aligned ECM, these sheets both induce and guide axon regeneration. Here, the capacity of bioactive conduits generated using these aligned DPSC sheets to restore function in critical-sized nerve injuries in rodents was evaluated.Approach. Scaffold-free nerve conduits were formed by culturing DPSCs on a substrate with aligned microgrooves, inducing the cells to align and deposit an aligned ECM. The sheets were then detached from the substrate and assembled into scaffold-free cylindrical tissues.Main results. In vitroanalyses confirmed that scaffold-free DPSC conduits maintained an aligned ECM and had uniformly distributed NTF expression. Implanting the aligned DPSC conduits across critical-sized defects in the buccal branch of rat facial nerves resulted in the regeneration of a fascicular nerve-like structure and myelinated axon extension across the injury site. Furthermore, compound muscle action potential and stimulated whisker movement measurements revealed that the DPSC conduit treatment promoted similar functional recovery compared to the clinical standard of care, autografts. Significance. This study demonstrates that scaffold-free aligned DPSC conduits supply trophic and guidance cues, key design elements needed to successfully promote and orient axon regeneration. Consequently, these conduits restore function in nerve injuries to similar levels as autograft treatments. These conduits offer a novel bioactive approach to nerve repair capable of improving clinical outcomes and patient quality of life.

The effectiveness of concentrated growth factor in facial nerve crush injury

AimTo evaluate the effect of concentrated growth factor (CGF) on regeneration of facial nerve after crush injury. Materials and MethodsFourteen rats were randomized into two groups. The control group (CG) (n = 7) received a crush injury to the right facial nerve. The CGF group (CGFG) (n = 7) received a crush injury to the right facial nerve and concentrated growth factor prepared from their own blood thereafter. Left facial nerves were used for functional comparison. Nerve function was evaluated using whisker movements and electromyography. Histologic properties were evaluated using hematoxylin and eosin and Masson-trichrome staining, and immunohistochemical properties were evaluated using Neurofilament-H and Anti-Tau degeneration markers. ResultsIn the CGFG, whisker functions began to recover earlier and recovered more quickly compared with the CG. The CG showed significantly prolonged latency and reduced amplitudes in the first week compared with the CGFG (p < 0.05). Recordings of 4th-week latency and amplitudes were similar to the preoperative period in the CGFG (p > 0.05), whereas recordings of the same week were significantly worse in the CG (p < 0.05). Edema and fibrosis were also more pronounced in the CG compared with the CGFG. Neurofilament-H and Anti-Tau were at significantly high levels in the CG (p < 0.05). ConclusionConcentrated growth factor promotes recovery in facial crush injury and may prove a cost-effective, practical, and effective treatment choice in peripheral nerve injury.

Biomimetic Hydrodynamic Sensor with Whisker Array Architecture and Multidirectional Perception Ability.

The rapid development of ocean exploration and underwater robot technology has put forward new requirements for underwater sensing methods, which can be used for hydrodynamic characteristics perception, underwater target tracking, and even underwater cluster communication. Here, inspired by the specialized undulated surface structure of the seal whisker and its ability to suppress vortex-induced vibration, a multidirectional hydrodynamic sensor based on biomimetic whisker array structure and magnetic 3D self-decoupling theory is introduced. The magnetic-based sensing method enables wireless connectivity between the magnetic functional structures and electronics, simplifying device design and endowing complete watertightness. The 3D self-decoupling capability enables the sensor, like a seal or other organisms, to perceive arbitrary whisker motions caused by the action of water flow without complex calibration and additional sensing units. The whisker sensor is capable of detecting a variety of hydrodynamic information, including the velocity (RMSE<0.061ms-1) and direction of the steady flow field, the frequency (error<0.05Hz) of the dynamic vortex wake, and the orientation (error<7°) of the vortex wake source, demonstrating its extensive potential for underwater environmental perception and communication, especially in deep sea conditions.

Co-coding of head and whisker movements by both VPM and POm thalamic neurons

Rodents continuously move their heads and whiskers in a coordinated manner while perceiving objects through whisker-touch. Studies in head-fixed rodents showed that the ventroposterior medial (VPM) and posterior medial (POm) thalamic nuclei code for whisker kinematics, with POm involvement reduced in awake animals. To examine VPM and POm involvement in coding head and whisker kinematics in awake, head-free conditions, we recorded thalamic neuronal activity and tracked head and whisker movements in male mice exploring an open arena. Using optogenetic tagging, we found that in freely moving mice, both nuclei equally coded whisker kinematics and robustly coded head kinematics. The fraction of neurons coding head kinematics increased after whisker trimming, ruling out whisker-mediated coding. Optogenetic activation of thalamic neurons evoked overt kinematic changes and increased the fraction of neurons leading changes in head kinematics. Our data suggest that VPM and POm integrate head and whisker information and can influence head kinematics during tactile perception.

Maternal immune activation affects female offspring whisker movements during object exploration in a rat model of neurodevelopmental disorders

Poly I:C rat offspring are used to investigate the effects of in utero exposure to maternal immune activation (MIA) and have been suggested as a model of neurodevelopmental disorders (NDD). The behavioural symptoms of this model are diverse and can vary with external factors, including the choice of background strain and husbandry practices. Measuring whisker movements provides quantitative, robust measurements of sensory, motor and cognitive behaviours in rodents. In this study, whisker movements were investigated in 50-day-old male and female offspring of MIA-exposed rat dams and compared to age-matched offspring of control (vehicle) dams. Rat offspring were filmed using high-speed videography in a sequential object exploration task with smooth and textured objects. Poly I:C treatment effects were found in female offspring that did not increase whisker mean angular position during object exploration, especially for the smooth object, indicating an attentional deficit. Whisker tracking during object exploration is demonstrated here, for the first time, as a useful, quick and non-invasive tool to identify both treatment effects and sex differences in a model of MIA-induced NDDs.

Thalamocortical Dynamics during Rapid Eye Movement Sleep in the Mouse Somatosensory Pathway.

Rapid eye movement (REM) sleep, also referred to as paradoxical sleep for the striking resemblance of its electroencephalogram (EEG) to the one observed in wakefulness, is characterized by the occurrence of transient events such as limb twitches or facial and rapid eye movements. Here, we investigated the local activity of the primary somatosensory or barrel cortex (S1) in naturally sleeping head-fixed male mice during REM. Through local field potential recordings, we uncovered local appearances of spindle waves in the barrel cortex during REM concomitant with strong delta power, challenging the view of a wakefulness-like activity in REM. We further performed extra- and intracellular recordings of thalamic cells in head-fixed mice. Our data show high-frequency thalamic bursts of spikes and subthreshold spindle oscillations in approximately half of the neurons of the ventral posterior medial nucleus which further confirmed the thalamic origin of local cortical spindles in S1 in REM. Cortical spindle oscillations were suppressed, while thalamus spike firing increased, associated with rapid mouse whisker movements and S1 cortical activity transitioned to an activated state. During REM, the sensory thalamus and barrel cortex therefore alternate between high (wake-like) and low (non-REM sleep-like) activation states, potentially providing a neuronal substrate for mnemonic processes occurring during this paradoxical sleep stage.

Implications of tactile enrichment on the behaviour and whisker movements of four species of carnivorans

Caniformia include a range of aquatic, semi-aquatic and terrestrial species, which reveal diversity in their whisker arrangements and shape. Whisker movements play a crucial role in the perception of tactile information, allowing whiskered mammals to distinguish between shapes, sizes, and textures. Despite the significance of whisker movements in sensory perception, few studies have focussed on measuring whisker movements during tactile sensing. Whisker enrichment tasks have the potential to expand behavioural repertoires of animals in captivity and reduce stereotypical behaviours. However, despite whiskers being essential in guiding foraging and exploration in many mammalian species, tactile whisker enrichment tasks are rare. Here, we utilised a novel tactile enrichment device to investigate the whisker movements in four Caniform species in captivity, including two pinnipeds- South African fur seals (Arctocephalus pussilus) and harbour seals (Phoca vitulina), a mustelid – Eurasian otter (Lutra lutra), and a canid – red fox (Vulpes vulpes) during a texture discrimination task. This study is the first to explore the impact of tactile enrichment on the behavioural repertoire of caniforms in captivity and provides the first insight of whisker movements in South African fur seals. The introduction of the tactile enrichment device did not increase the behavioural repertoire, nor did it lead to an increase in stereotypical behaviours or aggression in any of the species. However, it did successfully encourage natural whisker movements in the pinnipeds and otter. The whisker amplitude measure was especially high in the South African fur seals. We suggest that such a complex, discrimination-based enrichment task might only be feasible for more trainable caniforms, such as pinnipeds, rather than more neophobic species, such as the red fox, which did not interact with the enrichment device throughout the study. Therefore, while our enrichment device increased natural whisker movements, an even simpler foraging task might encourage tactile sensing without the requirement for training, making tactile whisker enrichment available to a wider group of species.

Simple spike patterns and synaptic mechanisms encoding sensory and motor signals in Purkinje cells and the cerebellar nuclei

Whisker stimulation in awake mice evokes transient suppression of simple spike probability in crus I/II Purkinje cells. Here, we investigated how simple spike suppression arises synaptically, what it encodes, and how it affects cerebellar output. Invitro, monosynaptic parallel fiber (PF)-excitatory postsynaptic currents (EPSCs) facilitated strongly, whereas disynaptic inhibitory postsynaptic currents (IPSCs) remained stable, maximizing relative inhibitory strength at the onset of PF activity. Short-term plasticity thus favors the inhibition of Purkinje spikes before PFs facilitate. Invivo, whisker stimulation evoked a 2-6ms synchronous spike suppression, just 6-8ms (∼4 synaptic delays) after sensory onset, whereas active whisker movements elicited broadly timed spike rate increases that did not modulate sensory-evoked suppression. Firing in the cerebellar nuclei (CbN) inversely correlated with disinhibition from sensory-evoked simple spike suppressions but was decoupled from slow, non-synchronous movement-associated elevations of Purkinje firing rates. Synchrony thus allows the CbN to high-pass filter Purkinje inputs, facilitating sensory-evoked cerebellar outputs that can drive movements.

Global and local neuronal coding of tactile information in the barrel cortex.

During tactile sensation in rodents, the whisker movements across surfaces give rise to intricate whisker motions that encompass discrete and transient stick-slip events, effectively conveying valuable information regarding surface properties. These surface characteristics are transformed into cortical neuronal responses. This study examined the coding strategies underlying these transformations in rat whiskers. We found that changes in surface coarseness modified the number and magnitude of stick-slip events, which in turn both modulated properties of neuronal responses. Global changes in the number of stick-slip events primarily affected neuronal discharge rates and the degree of neuronal synchronization. In contrast, local changes in the magnitude of stick-slip events affected the transformation of these kinematic and kinetic characteristics into neuronal discharges. Most cortical neurons exhibited surface coarseness selectivity through global and local stick-slip event properties. However, this selectivity varied across coding strategies in the same neurons, given that each coding strategy reflected different aspects of changes in whisker-surface interactions. The degree of spatial similarity in surface coarseness preference in adjacently recorded neurons differed among these coding strategies. Adjacently recorded neurons exhibited the same surface coarseness preference in their firing rates but not through other coding strategies. Through these results, we were able to show that local stick-slip event properties contribute to texture discrimination, complementing and surpassing global coding in this context. These findings suggest that the representation of surface coarseness in the cortex may rely on concurrent coding strategies that integrate tactile information across different spatiotemporal scales.

Reliability and stability of tactile perception in the whisker somatosensory system.

Rodents rely on their whiskers as vital sensory tools for tactile perception, enabling them to distinguish textures and shapes. Ensuring the reliability and constancy of tactile perception under varying stimulus conditions remains a fascinating and fundamental inquiry. This study explores the impact of stimulus configurations, including whisker movement velocity and object spatial proximity, on texture discrimination and stability in rats. To address this issue, we employed three distinct approaches for our investigation. Stimulus configurations notably affected tactile inputs, altering whisker vibration's kinetic and kinematic aspects with consistent effects across various textures. Through a texture discrimination task, rats exhibited consistent discrimination performance irrespective of changes in stimulus configuration. However, alterations in stimulus configuration significantly affected the rats' ability to maintain stability in texture perception. Additionally, we investigated the influence of stimulus configurations on cortical neuronal responses by manipulating them experimentally. Notably, cortical neurons demonstrated substantial and intricate changes in firing rates without compromising the ability to discriminate between textures. Nevertheless, these changes resulted in a reduction in texture neuronal response stability. Stimulating multiple whiskers led to improved neuronal texture discrimination and maintained coding stability. These findings emphasize the importance of considering numerous factors and their interactions when studying the impact of stimulus configuration on neuronal responses and behavior.

Whisker kinematics in the cerebellum.

The whisker system is widely used as a model system for understanding sensorimotor integration. Purkinje cells in the crus regions of the cerebellum have been reported to linearly encode whisker midpoint, but it is unknown whether the paramedian and simplex lobules as well as their target neurons in the cerebellar nuclei also encode whisker kinematics and if so which ones. Elucidating how these kinematics are represented throughout the cerebellar hemisphere is essential for understanding how the cerebellum coordinates multiple sensorimotor modalities. Exploring the cerebellar hemisphere of mice using optogenetic stimulation, we found that whisker movements can be elicited by stimulation of Purkinje cells in not only crus1 and crus2, but also in the paramedian lobule and lobule simplex; activation of cells in the medial paramedian lobule had on average the shortest latency, whereas that of cells in lobule simplex elicited similar kinematics as those in crus1 and crus2. During spontaneous whisking behaviour, simple spike activity correlated in general better with velocity than position of the whiskers, but it varied between protraction and retraction as well as per lobule. The cerebellar nuclei neurons targeted by the Purkinje cells showed similar activity patterns characterized by a wide variety of kinematic signals, yet with a dominance for velocity. Taken together, our data indicate that whisker movements are much more prominently and diversely represented in the cerebellar cortex and nuclei than assumed, highlighting the rich repertoire of cerebellar control in the kinematics of movements that can be engaged during coordination. KEY POINTS: Excitation of Purkinje cells throughout the cerebellar hemispheres induces whisker movement, with the shortest latency and longest duration within the paramedian lobe. Purkinje cells have differential encoding for the fast and slow components of whisking. Purkinje cells encode not only the position but also the velocity of whiskers. Purkinje cells with high sensitivity for whisker velocity are preferentially located in the medial part of lobule simplex, crus1 and lateral paramedian. In the downstream cerebellar nuclei, neurons with high sensitivity for whisker velocity are located at the intersection between the medial and interposed nucleus.

Neural mechanisms underlying uninstructed orofacial movements during reward-based learning behaviors

During reward-based learning tasks, animals make orofacial movements that globally influence brain activity at the timings of reward expectation and acquisition. These orofacial movements are not explicitly instructed and typically appear along with goal-directed behaviors. Here, we show that reinforcing optogenetic stimulation of dopamine neurons in the ventral tegmental area (oDAS) in mice is sufficient to induce orofacial movements in the whiskers and nose without accompanying goal-directed behaviors. Pavlovian conditioning with a sensory cue and oDAS elicited cue-locked and oDAS-aligned orofacial movements, which were distinguishable by a machine-learning model. Inhibition or knockout of dopamine D1 receptors in the nucleus accumbens inhibited oDAS-induced motion but spared cue-locked motion, suggesting differential regulation of these two types of orofacial motions. In contrast, inactivation of the whisker primary motor cortex (wM1) abolished both types of orofacial movements. We found specific neuronal populations in wM1 representing either oDAS-aligned or cue-locked whisker movements. Notably, optogenetic stimulation of wM1 neurons successfully replicated these two types of movements. Our results thus suggest that accumbal D1-receptor-dependent and -independent neuronal signals converge in the wM1 for facilitating distinct uninstructed orofacial movements during a reward-based learning task.

Effects of platelet-rich fibrin and dexamethasone on nerve regeneration after acute facial-nerve injury

Background Acute facial-nerve injury. Objective To investigate the effects of platelet-rich fibrin (PRF) and dexamethasone on nerve regeneration. Materials and methods Thirty-six rats were randomly divided into six groups. Facial-nerve injury was created using a full-thickness incision in all groups except Group E. Next, primary anastomosis, PRF application, topical dexamethasone application, primary anastomosis with topical PRF and dexamethasone application, and no facial-nerve repair were performed in Groups A, B, C, D, and F, respectively. Clinical, functional, and structural improvements were evaluated at eight weeks. Results The mean eye-closure movement score in Group B was significantly higher than that in Group F (p < .001). The mean whisker-movement score in Group B was significantly higher than that in Group F (p = .001). The mean amplitude of whisker movement in Group F was significantly lower than those in Groups A, B, C, and E, and the mean amplitude in Group D was significantly lower than that in Group E (p < .001). Furthermore, an improvement in nerve ultrastructure was observed in Group B. Conclusion PRF application has a positive effect on nerve recovery after anastomosis. Significance Contribute to the literature to improve nerve regeneration.

The interconnection and function of associative memory neurons are upregulated for memory strengthening.

Memories associated to signals have been proven to rely on the recruitment of associative memory neurons that are featured by mutual synapse innervations among cross-modal cortices. Whether the consolidation of associative memory is endorsed by the upregulation of associative memory neurons in an intramodal cortex remains to be examined. The function and interconnection of associative memory neurons were investigated by in vivo electrophysiology and adeno-associated virus-mediated neural tracing in those mice that experienced associative learning by pairing the whisker tactile signal and the olfactory signal. Our results show that odorant-induced whisker motion as a type of associative memory is coupled with the enhancement of whisking-induced whisker motion. In addition to some barrel cortical neurons encoding both whisker and olfactory signals, i.e., their recruitment as associative memory neurons, the synapse interconnection and spike-encoding capacity of associative memory neurons within the barrel cortex are upregulated. These upregulated alternations were partially observed in the activity-induced sensitization. In summary, associative memory is mechanistically based on the recruitment of associative memory neurons and the upregulation of their interactions in intramodal cortices.

Microstimulation of sensory cortex engages natural sensory representations

Cortical activity patterns occupy a small subset of possible network states. If this is due to intrinsic network properties, microstimulation of sensory cortex should evoke activity patterns resembling those observed during natural sensory input. Here, we use optical microstimulation of virally transfected layer 2/3 pyramidal neurons in the mouse primary vibrissal somatosensory cortex to compare artificially evoked activity with natural activity evoked by whisker touch and movement ("whisking"). We find that photostimulation engages touch- but not whisking-responsive neurons more than expected by chance. Neurons that respond to photostimulation and touch or to touch alone exhibit higher spontaneous pairwise correlations than purely photoresponsive neurons. Exposure to several days of simultaneous touch and optogenetic stimulation raises both overlap and spontaneous activity correlations among touch and photoresponsive neurons. We thus find that cortical microstimulation engages existing cortical representations and that repeated co-presentationof natural and artificial stimulation enhances this effect.

Effect of Electrical Stimulation on Nerve-Guided Facial Nerve Regeneration.

This study aimed to investigate the effect of electrical stimulation on poly(d,l-lactide-co-ε-caprolactone) nerve guidance conduits (NGCs) in promoting the recovery of facial function and nerve regeneration after facial nerve (FN) injury in a rat model. In the experimental group, both the NGC and transcutaneous electrical nerve stimulation (ES) were used simultaneously; in the control group, only NGC was used. ES groups were divided into two groups, and direct current (DC) and charge-balanced pulse stimulation (Pulse) were applied. The ES groups showed significantly improved whisker movement than the NGC-only group. The number of myelinated neurons was higher in ES groups, and the myelin sheath was also thicker and more uniform. In addition, the expression of neurostructural proteins was also higher in ES groups than in the NGC-only group. This study revealed that FN regeneration and functional recovery occurred more efficiently when ES was applied in combination with NGCs.