Terminal Schwann cell structure is altered in diaphragm of mdx mice

The diaphragm muscle of the mdx mouse is a model system of Duchenne muscular dystrophy, since it completely lacks dystrophin and shows severe fiber necrosis and loss of specific muscle force by 4-6 weeks of age. Changes in neuromuscular junction structure also become apparent around 4 weeks including postsynaptic acetylcholine receptor declustering, loss of postsynaptic junctional folds, abnormally complex presynaptic nerve terminals, and muscle fiber denervation. Normally, terminal Schwann cells (TSCs) cap both nerve terminals and acetylcholine receptors at the neuromuscular junction, and play a crucial role in regeneration of motor axons following muscle denervation by guiding axons to grow from innervated junctions to nearby denervated junctions. However, their role in restoring innervation in dystrophic muscle is unknown. We now show that TSCs fail to cap fully the neuromuscular junction in dystrophic muscle; TSCs extend processes, but the organization of these extensions is abnormal. TSC processes of dystrophic muscle do not form bridges from denervated fibers to nearby innervated endplates, but appear to be directed away from these endplates. Adequate signaling for TSC reactivity is present, since significant muscle fiber denervation and acetylcholine receptor declustering are present. Thus, significant structural denervation is present in the diaphragm of mdx mice and the ability of TSCs to form bridges between adjacent endplates to guide reinnervation of muscle fibers is impaired, possibly attenuating the ability of dystrophic muscle to recover from denervation and ultimately leading to muscle weakness.     

Schwann cell guidance of nerve growth between synaptic sites explains changes in the pattern of muscle innervation and remodeling of synaptic sites following peripheral nerve injuries

Terminal Schwann cells (SCs) are nonmyelinating glia that are a prominent component of the neuromuscular junction (NMJ) where motor neurons form synapses onto muscle fibers. These cells play important roles not only in development and maintenance of the neuromuscular synapse but also restoring synaptic function after nerve damage. In response to muscle denervation, terminal SCs undergo dramatic changes in their gene expression patterns as well as in their morphology, such as extending elaborate processes into inter-junctional space. These SC processes serve as a path to guide axon terminal sprouts from nearby innervated junctions, promoting rapid reinnervation of denervated fibers. We studied the role of terminal SCs in synapse reformation by using two different fluorescent proteins to simultaneously label motor axons and SCs; we examined these junctions repeatedly in living animals using a fluorescence microscope. Here, we show that alterations in the patterns of muscle innervation following recovery from nerve injury can be explained by SC guidance of regenerating axons. In turn, this guidance leads to remodeling of the NMJ itself.     

Leukemia inhibitory factor enhances the regeneration of transected rat sciatic nerve and the function of reinnervated muscle

The cytokine leukemia inhibitory factor (LIF) favors the survival and growth of axons in vitro and in vivo. Fibronectin has been shown to enhance nerve regeneration when added in combination with various growth factors including LIF. The goal of this study was to evaluate the effect of LIF plus fibronectin on the regeneration of transected nerve and functional recovery of reinnervated skeletal muscle, in one experimental model of peripheral nerve repair, at two recovery times. The rat sciatic nerve was cut at mid-thigh level and a silicone cuff containing either saline (control), LIF, or LIF plus fibronectin (L + F) was used to bridge the proximal and distal nerve stumps leaving a 1 cm gap between them. Rats were then explored at 6 or 12 weeks following the initial surgery. Regenerating nerves were assessed by measuring the diameter of myelinated axons, conduction velocity, and number of myelinated fibers. Muscle reinnervation was assessed by measuring muscle mass, force of contraction, and histologically for changes in muscle fiber type (type I and type II). In this report we demonstrate that at 6 weeks there were significant increases in 1) nerve conduction velocity, 2) myelinated axon diameter, and 3) number of myelinated axons over that of control (saline-treated) animals. Both LIF groups demonstrated a shift in type II muscle fiber area compared to saline-treated controls, with the L + F group having a significant increase in muscle mass. At 12 weeks there was an improved recovery over and above that demonstrated at 6 weeks. Muscle mass was 65% and 42% greater than control for LIF and L + F, respectively. Force of contraction, conduction velocity, myelinated fiber number, and diameter were also significantly greater for both LIF- and L + F- treated rats than saline-treated rats. These results demonstrate that LIF significantly improves the regeneration of damaged peripheral nerves and the preservation of muscle viability, resulting in greatly enhanced recovery of skeletal muscle function. J. Neurosci. Res. 47:208–215, 1997. © 1997 Wiley-Liss, Inc.     

A spatio-temporal analysis of motoneuron survival, axonal regeneration and neurotrophic factor expression after lumbar ventral root avulsion and implantation

Reimplantation of avulsed rat lumbar spinal ventral roots results in poor recovery of function of the denervated hind limb muscles. In contrast, reimplantation of cervical or sacral ventral roots is a successful repair strategy that results in a significant degree of regeneration. A possible explanation for this difference could be that following lumbar root avulsion, axons have to travel longer distances towards their target muscles, resulting in prolonged denervation of the distal nerve and a diminished capacity to support regeneration. Here we present a detailed spatio-temporal analysis of motoneuron survival, axonal regeneration and neurotrophic factor expression following unilateral avulsion and implantation of lumbar ventral roots L3, L4, and L5. Reimplantation prolongs the survival of motoneurons up to one month post-lesion. The first regenerating motor axons entered the reimplanted ventral roots during the first week and large numbers of fibers gradually enter the lumbar plexus between 2 and 4 weeks, indicating that axons enter the reimplanted roots and plexus over an extended period of time. However, motor axon counts show that relatively few axons reach the distal sciatic nerve in the 16 week post-lesion period. The observed initial increase and subsequent decline in expression of glial cell line-derived neurotrophic factor and brain-derived neurotrophic factor correlate with the apparent spatio-temporal decline in the regenerative capacity of motor axons, indicating that the distal nerve is losing its capacity to support regenerating motor axons following prolonged denervation. These findings have important implications for future strategies to promote long-distance regeneration through distal, chronically denervated peripheral nerves.     

Regenerated EDL muscle of rats requires innervation to maintain AChE molecular forms

Extensores digitorum longi of rats, infarcted and denervated by different surgical procedures, were used to analyze by biochemical and cytochemical methods the acetylcholinesterase (AChE) changes during muscle degeneration, regeneration, and early or delayed reinnervation. Biochemical tests showed that the regenerating muscle produces globular AChE forms (36% of controls) and small amounts of A12 (16S) asymmetric form (5% of controls); at the end of the regeneration, innervation and electromechanical function are required for the complete recovery of globular forms, and are absolutely critical to prevent A12 (16S) disappearance. Cytochemical observations showed that, unlike nicotinic receptor, AChE deposited at the neuromuscular junction before ischemic necrosis is protected from breakdown, as is the basal lamina of muscle fibers. Taken together, these observations contribute to the understanding of the factors that play a critical role in muscle repair and are, therefore, of clinical relevance.     

Long-term effects of muscle-derived protein with molecular mass of 77 kDa (MDP77) on nerve regeneration

The long-term effects of the 77-kDa muscle-derived protein (MDP77) on motor and sensory nerve regeneration were examined in vivo. Fourteen-millimeter bridge grafts of the right sciatic nerve of SD rats were carried out with silicone tubes containing a solution of type I collagen together with 0, 5, 10, or 20 microg/ml recombinant human MDP77 (N = 10 in each group). Recovery of motor and sensory function was evaluated monthly by the maximal toe-spread index (TSI) and hot-plate test, respectively, for 6 months after the operation. Electrophysiology (nerve conduction velocity), histology (diameter and total number of the regenerated myelinated axons in the tube), and immunohistochemistry (total number of Schwann cells in the tube), as well as measurement of soleus muscle weight, were also performed at this time. Motor, but not sensory, function recovered rapidly in the MDP77-treated groups in a dose-dependent manner. Electrophysiological measurements and the ratio of soleus muscle weight corroborated the positive effects of MDP77 on motor nerve regeneration, but no facilitation of sensory nerve recovery was observed. Furthermore, histological and immunohistochemical evaluations suggested that MDP77 treatment accelerates Schwann cell migration, followed by enhanced maturation of regenerating axons, resulting in functional recovery of both the nerves and the atrophied, denervated muscle.     

Regeneration of muscle axons in the frog is directed by diffusible factors from denervated muscle and nerve tubes

In the frog, peripheral muscle axons regenerate after a lesion to reinnervate the original synaptic sites on muscle fibers. Previous experiments in the frog have shown that satellite cells of the nerve tube direct the outgrowth of regenerating muscle axons over distances of many millimeters. In the present experiments, denervated muscle was used as a target for regenerating muscle axons. Muscle and satellite cells of the nerve tube also were placed in filters to determine if their influence on axonal outgrowth was exerted by diffusible factors. Filters were used with a pore size of 0.22 micron. With this pore size, target cells were isolated from physical contact with the surrounding cells; yet an exchange of fluids--and therefore of molecules released by the target cells--could occur across the filter. In the presence of denervated muscle or satellite cells of the nerve tube in filters, regenerating axons turn and grow toward the target cells. This influence on the direction of axonal outgrowth was produced over distances of 6 mm by muscles and 4 mm by cells of the nerve tubes. This directed outgrowth is in marked contrast to the random pattern of outgrowth in the absence of the targets. The present findings set the stage for tissue culture experiments in which the phenomena observed in vivo can be analyzed in terms of mechanisms. The present finding that denervated muscle attracts regenerating axons means that sufficient material may be available for the characterization and isolation of the relevant molecules.     

Recovery of muscle after different periods of denervation and treatments

Three aspects of reinnervation and recovery of skeletal muscle following various periods of denervation were investigated: (1) the effect of duration of denervation; (2) the effect of hyperthyroidism on recovery; and (3) whether the muscle or the nerve limits recovery. The rat medial gastrocnemius (MG) nerve was cut and then resutured after 0, 3, 7, 21, or 56 days. In a second group of animals, the MG muscle was denervated and, in addition, the animal received triiodothyronine (T₃) supplementation during reinnervation. The third group of animals had the denervated MG muscle reinnervated by a larger number of newly transected foreign axons. The force produced by the reinnervated muscle depends on the period that the muscle was denervated. Recovery was impaired when the period of denervation exceeded 7 days. T₃ treatment did not benefit the return of force production, nor did providing the muscle with a larger number of newly transected axons.     

The effect of duration of muscle denervation on functional recovery in the rat model

The effect of long-term denervation on neuromuscular recovery was studied in a rat hind limb model. The posterior tibial nerve was transected and repaired immediately or after denervation periods of 2 weeks, or 1, 3, 6, 9, or 12 months. Six months following reconstruction excellent axonal regeneration was seen across all nerve repairs irrespective of periods of denervation. However, there was a precipitous and profound decrease in the recovery of both muscle mass and integrated motor function if the reconstruction was delayed for longer than 1 month. Rather than a progressive change proportional to the length of the denervation period, significant, more discrete changes occurred sometime after 1 month of denervation that precluded a full recovery of muscle mass. Integrated motor function quantified using walking track analysis was impaired even after immediate nerve repair. © 1997 John Wiley & Sons, Inc. Muscle Nerve 20: 858–866, 1997     

Long-term delivery of FGF-6 changes the fiber type and fatigability of muscle reinnervated from embryonic neurons transplanted into adult rat peripheral nerve

Motoneuron death leads to muscle denervation and atrophy. Transplantation of embryonic neurons into peripheral nerves results in reinnervation and provides a strategy to rescue muscles from atrophy independent of neuron replacement in a damaged or diseased spinal cord. But the count of regenerating axons always exceeds the number of motor units in this model, so target-derived trophic factor levels may limit reinnervation. Our aim was to examine whether long-term infusion of fibroblast growth factor-6 (FGF-6) into denervated medial gastrocnemius muscles improved the function of muscles reinnervated from neurons transplanted into nerve of adult Fischer rats. Factor delivery (10 microg, 4 weeks) began after sciatic nerve transection. After a week of nerve degeneration, 1 million embryonic day 14-15 ventral spinal cord cells were transplanted into the distal tibial stump as a neuron source. Ten weeks later, neurons that expressed motoneuron markers survived in the nerves. More myelinated axons were in nerves to saline-treated muscles than in FGF-6-treated muscles. However, each group showed comparable reductions in muscle fiber atrophy because of reinnervation. Mean reinnervated fiber area was 43%-51% of non-denervated fibers. Denervated fiber area averaged 11%. FGF-6-treated muscles were more fatigable than other reinnervated muscles but had stronger motor units and fewer type I fibers than did saline-treated muscles. FGF-6 thus influenced function by changing the type of fiber reinnervated by transplanted neurons. Deficits in FGF-6 may also contribute to the increase in type I fibers in muscles reinnervated from peripheral axons, suggesting that the effects of FGF-6 on fiber type are independent of the neuron source used for reinnervation.     

Acrylamide inhibits nerve sprouting induced by botulinum toxin type A

Botulinum toxin type A is a potent muscle relaxant that blocks the transmission and release of acetylcholine at the neuromuscular junction. Intramuscular injection of botulinum toxin type A has served as an effective and safe therapy for strabismus and focal dystonia. However, muscular weakness is temporary and after 3–4 months, muscle strength usually recovers because functional recovery is mediated by nerve sprouting and reconstruction of the neuromuscular junction. Acrylamide may produce neurotoxic substances that cause retrograde necrotizing neuropathy and inhibit nerve sprouting caused by botulinum toxin type A. This study investigated whether acrylamide inhibits nerve sprouting after intramuscular injection of botulinum toxin type A. A tibial nerve sprouting model was established through local injection of botulinum toxin type A into the right gastrocnemius muscle of Sprague-Dawley rats. Following intramuscular injection, rats were given intraperitoneal injection of 3% acrylamide every 3 days for 21 days. Nerve sprouting appeared 2 weeks after intramuscular injection of botulinum toxin type A and single-fiber electromyography revealed abnormal conduction at the neuromuscular junction 1 week after intramuscular injection of botulinum toxin type A. Following intraperitoneal injection of acrylamide, the peak muscle fiber density decreased. Electromyography jitter value were restored to normal levels 6 weeks after injection. This indicates that the maximal decrease in fiber density and the time at which functional conduction of neuromuscular junction was restored were delayed. Additionally, the increase in tibial nerve fibers was reduced. Acrylamide inhibits nerve sprouting caused by botulinum toxin type A and may be used to prolong the clinical dosage of botulinum toxin type A.     

Nerve conduction and electromyography studies

Nerve conduction studies (NCS) and electromyography (EMG), often shortened to 'EMGs', are a useful adjunct to clinical examination of the peripheral nervous system and striated skeletal muscle. NCS provide an efficient and rapid method of quantifying nerve conduction velocity (CV) and the amplitude of both sensory nerve action potentials (SNAPs) and compound motor action potentials (cMAPs). The CV reflects speed of propagation of action potentials, by saltatory conduction, along large myelinated axons in a peripheral nerve. The amplitude of SNAPs is in part determined by the number of axons in a sensory nerve, whilst amplitude of cMAPs reflects integrated function of the motor axons, neuromuscular junction and striated muscle. Repetitive nerve stimulation (RNS) can identify defects of neuromuscular junction (NMJ) transmission, pre- or post-synaptic. Needle EMG examination can detect myopathic changes in muscle and signs of denervation. Combinations of these procedures can establish if motor and/or sensory nerve cell bodies or peripheral nerves are damaged (e.g. motor neuronopathy, sensory ganglionopathy or neuropathy), and also indicate if the primary target is the axon or the myelin sheath (i.e. axonal or demyelinating neuropathies). The distribution of nerve damage can be determined as either generalised, multifocal (mononeuropathy multiplex) or focal. The latter often due to compression at the common entrapment sites (such as the carpal tunnel, Guyon's canal, cubital tunnel, radial groove, fibular head and tarsal tunnel, to name but a few of the reported hundred or so 'entrapment neuropathies').     

Experimental strategies to promote functional recovery after peripheral nerve injuries

The capacity of Schwann cells (SCs) in the peripheral nervous system to support axonal regeneration, in contrast to the oligodendrocytes in the central nervous system, has led to the misconception that peripheral nerve regeneration always restores function. Here, we consider how prolonged periods of time that injured neurons remain without targets during axonal regeneration (chronic axotomy) and that SCs in the distal nerve stumps remain chronically denervated (chronic denervation) progressively reduce the number of motoneurons that regenerate their axons. We demonstrate the effectiveness of low-dose, brain-derived neurotrophic and glial-derived neurotrophic factors to counteract the effects of chronic axotomy in promoting axonal regeneration. High-dose brain-derived neurotrophic factor (BDNF) on the other hand, acting through the p75 receptor, inhibits axonal regeneration and may be a factor in stopping regenerating axons from forming neuromuscular connections in skeletal muscle. The immunophilin, FK506, is also effective in promoting axonal regeneration after chronic axotomy. Chronic denervation of SCs (>1 month) severely deters axonal regeneration, although the few motor axons that do regenerate to reinnervate muscles become myelinated and form enlarged motor units in the reinnervated muscles. We found that in vitro incubation of chronically denervated SCs with transforming growth factor-beta re-established their growth-supportive phenotype in vivo, consistent with the idea that the interaction between invading macrophages and denervated SCs during Wallerian degeneration is essential to sustain axonal regeneration by promoting the growth-supportive SC phenotype. Finally, we consider the effectiveness of a brief period of 20 Hz electrical stimulation in promoting the regeneration of axons across the surgical gap after nerve repair.     

Heat shock protein is a key therapeutic target for nerve repair in autoimmune peripheral neuropathy and severe peripheral nerve injury

Guillain-Barré syndrome (GBS) is an autoimmune peripheral neuropathy and a common cause of neuromuscular paralysis. Preceding infection induces the production of anti-ganglioside (GD) antibodies attacking its own peripheral nerves. In severe proximal peripheral nerve injuries that require long-distance axon regeneration, motor functional recovery is virtually nonexistent. Damaged axons fail to regrow and reinnervate target muscles. In mice, regenerating axons must reach the target muscle within 35 days (critical period) to reform functional neuromuscular junctions and regain motor function. Successful functional recovery depends on the rate of axon regeneration and debris removal (Wallerian degeneration) after nerve injury. The innate-immune response of the peripheral nervous system to nerve injury such as timing and magnitude of cytokine production is crucial for Wallerian degeneration. In the current study, forced expression of human heat shock protein (hHsp) 27 completely reversed anti-GD-induced inhibitory effects on nerve repair assessed by animal behavioral assays, electrophysiology and histology studies, and the beneficial effect was validated in a second mouse line of hHsp27. The protective effect of hHsp27 on prolonged muscle denervation was examined by performing repeated sciatic nerve crushes to delay regenerating axons from reaching distal muscle from 37 days up to 55 days. Strikingly, hHsp27 was able to extend the critical period of motor functional recovery for up to 55 days and preserve the integrity of axons and mitochondria in distal nerves. Cytokine array analysis demonstrated that a number of key cytokines which are heavily involved in the early phase of innate-immune response of Wallerian degeneration, were found to be upregulated in the sciatic nerve lysates of hHsp27 Tg mice at 1 day postinjury. However, persistent hyperinflammatory mediator changes were found after chronic denervation in sciatic nerves of littermate mice, but remained unchanged in hHsp27 Tg mice. Taken together, the current study provides insight into the development of therapeutic strategies to enhance muscle receptiveness (reinnervation) by accelerating axon regeneration and Wallerian degeneration.     

Treatment of denervated muscle by gangliosides

Short-duration cooling of the nerve to the extensor digitorum longus muscle of the rat in vivo induced partially reversible denervation of the muscle and atrophy in the type 2 muscle fibers. Increases in cyclic adenosine monophosphate, cyclic guanosine monophosphate phosphodiesterase, adenylate cyclase, and guanlate cyclase were observed in the denervated muscle. Treatment with gangliosides of the bovine brain cortex seemed to improve the excitability of the surviving motor units and to encourage recovery of neuromuscular trophic control, but it did not affect the nerve conduction velocity or the contractile properties of the denervated muscle.     

Gpr126/Adgrg6 contributes to the terminal Schwann cell response at the neuromuscular junction following peripheral nerve injury

Gpr126/Adgrg6 is an adhesion G protein-coupled receptor essential for Schwann cell (SC) myelination with important contributions to repair after nerve crush injury. Despite critical functions in myelinating SCs, the role of Gpr126 within nonmyelinating terminal Schwann cells (tSCs) at the neuromuscular junction (NMJ), is not known. tSCs have important functions in synaptic maintenance and reinnervation, and after injury tSCs extend cytoplasmic processes to guide regenerating axons to the denervated NMJ. In this study, we show that Gpr126 is expressed in tSCs, and that absence of Gpr126 in SCs (SC-specific Gpr126 knockout, cGpr126) results in a NMJ maintenance defect in the hindlimbs of aged mice, but not in young adult mice. After nerve transection and repair, cGpr126 mice display delayed NMJ reinnervation, altered tSC morphology with decreased S100β expression, and reduced tSC cytoplasmic process extensions. The immune response promoting reinnervation at the NMJ following nerve injury is also altered with decreased macrophage infiltration, Tnfα, and anomalous cytokine expression compared to NMJs of control mice. In addition, Vegfa expression is decreased in muscle, suggesting that cGpr126 non-cell autonomously modulates angiogenesis after nerve injury. In sum, cGpr126 mice demonstrated delayed NMJ reinnervation and decreased muscle mass following nerve transection and repair compared to control littermates. The integral function of Gpr126 in tSCs at the NMJ provides the framework for new therapeutic targets for neuromuscular disease.     

Chronic Schwann cell denervation and the presence of a sensory nerve reduce motor axonal regeneration

Motor axonal regeneration is compromised by chronic distal nerve stump denervation, induced by delayed repair or prolonged regeneration distance, suggesting that the pathway for regeneration is progressively impaired with time and/or distance. In the present experiments, we tested the impacts of (i) chronic distal sensory nerve stump denervation on axonal regeneration and (ii) sensory or motor innervation of a nerve graft on the ability of motoneurons to regenerate their axons from the opposite end of the graft. Using the motor and sensory branches of rat femoral nerve and application of neuroanatomical tracers, we evaluated the numbers of regenerated femoral motoneurons and nerve fibers when motoneurons regenerated (i) into freshly cut and 2-month chronically denervated distal sensory nerve stump, (ii) alone into a 4-cm-long distally ligated sensory autograft (MGL) and, (iii) concurrently as sensory (MGS) or motor (MGM) nerves regenerated into the same autograft from the opposite end. We found that all (315 +/- 24: mean +/- SE) the femoral motoneurons regenerated into a freshly cut distal sensory nerve stump as compared to 254 +/- 20 after 2 months of chronic denervation. Under the MGL condition, 151 +/- 5 motoneurons regenerated, which was not significantly different from the MGM group (134 +/- 13) but was significantly reduced to 99 +/- 2 in the MGS group (P < 0.05). The number of regenerated nerve fibers was 1522 +/- 81 in the MGL group, 888 +/- 18 in the MGM group, and 516 +/- 44 in the MGS group, although the high number of nerve fibers in the MGL group was due partly to the elaboration of multiple sprouts. Nerve fiber number and myelination were reduced in the MGS group and increased in the MGM group. These results demonstrate that both chronic denervation and the presence of sensory nerve axons reduced desired motor axonal regeneration into sensory pathways. A common mechanism may involve reduced responsiveness of sensory Schwann cells within the nerve graft or chronically denervated distal nerve stump to regenerating motor axons. The findings confirm that motor regeneration is optimized by avoiding even short-term denervation. They also imply that repairing pure motor nerves (without their cutaneous sensory components) to distal nerve stumps should be considered clinically when motor recovery is the main desired outcome.     

Fibrillation potential amplitude to quantitatively assess denervation muscle atrophy

Denervated muscle fibers exhibit spontaneous, repetitive single muscle fiber discharges and display fibrillation potentials detectable by electromyography. To explore the changing pattern of fibrillation potential amplitude after peripheral nerve injury and its relationship to the degree of muscle atrophy, fibrillation potential amplitudes were recorded on completely denervated biceps brachii of 173 patients with brachial plexus injury. Biceps brachii biopsies were taken at the same sites as the electromyogram recordings in 63 patients. The biopsies were analyzed by ATPase staining and the cross-sectional areas of fast and slow-twitch fibers were calculated. We found that the fibrillation potential amplitude and the cross-sectional areas of denervated muscle decay over time (P < 0.05), and both correlate negatively with denervation time (P < 0.01-0.05) within the first 15 months. The fibrillation potential amplitude correlates positively with both type I and II fiber cross-sectional areas (P < 0.0005-0.01). Our results show that fibrillation potential amplitude is closely correlated with muscle fiber size during the first 15 months after nerve injury, and it may therefore serve as a convenient index to evaluate quantitatively the degree of atrophy of denervated muscles. Electromyographic studies thus may help in designing treatment strategies.     

Electrical stimulation accelerates nerve regeneration and functional recovery in delayed peripheral nerve injury in rats

The present study aims to investigate the potential of brief electrical stimulation (ES; 3 V, 20 Hz, 20 min) in improving functional recovery in delayed nerve injury repair (DNIR). The sciatic nerve of Sprague Dawley rats was transected, and the repair of nerve injury was delayed for different time durations (2, 4, 12 and 24 weeks). Brief depolarizing ES was applied to the proximal nerve stump when the transected nerve stumps were bridged with a hollow nerve conduit (5 mm in length) after delayed periods. We found that the diameter and number of regenerated axons, the thickness of myelin sheath, as well as the number of Fluoro‐Gold retrograde‐labeled motoneurons and sensory neurons were significantly increased by ES, suggesting that brief ES to proximal nerve stumps is capable of promoting nerve regeneration in DNIR with different delayed durations, with the longest duration of 24 weeks. In addition, the amplitude of compound muscle action potential (gastrocnemius muscle) and nerve conduction velocity were also enhanced, and gastrocnemius muscle atrophy was partially reversed by brief ES, indicating that brief ES to proximal nerve stump was able to improve functional recovery in DNIR. Furthermore, brief ES was capable of increasing brain‐derived neurotrophic factor (BDNF) expression in the spinal cord in DNIR, suggesting that BDNF‐mediated neurotrophin signaling might be one of the contributing factors to the beneficial effect of brief ES on DNIR. In conclusion, the present findings indicate the potential of using brief ES as a useful method to improve functional recovery for delayed repair of peripheral nerve lesions.     

Differential effects of nerve, muscle, and fat tissue on regenerating nerve fibers in vivo

Axonal regeneration through silicone tubes was studied using distal nerve stumps, denervated, preatrophied muscle tissue, as well as fat tissue as a target. During the first stage of regeneration, i.e., within 2-3 weeks after surgery, a thin, filamentous structure consisting of fibrin and connective tissue was seen bridging the gap in all systems. Thereafter, this cord obviously served as a guideline for the outgrowth of increasing numbers of axons into distal nerve stumps as well as into muscle tissue, but not into fat tissue. These findings confirm that preatrophied muscle tissue has a similar "neurotrophic" effect on regenerating nerve fibers as distal nerve stumps. The ineffectivity of fat tissue in promoting nerve fiber regeneration could be attributed either to the absence of "neurotrophic factors" or even to an inhibitory effect.