Misdirection of regenerating axons and functional recovery following sciatic nerve injury in rats

Poor functional recovery found after peripheral nerve injury has been attributed to the misdirection of regenerating axons to reinnervate functionally inappropriate muscles. We applied brief electrical stimulation (ES) to the common fibular (CF) but not the tibial (Tib) nerve just prior to transection and repair of the entire rat sciatic nerve, to attempt to influence the misdirection of its regenerating axons. The specificity with which regenerating axons reinnervated appropriate targets was evaluated physiologically using compound muscle action potentials (M responses) evoked from stimulation of the two nerve branches above the injury site. Functional recovery was assayed using the timing of electromyography (EMG) activity recorded from the tibialis anterior (TA) and soleus (Sol) muscles during treadmill locomotion and kinematic analysis of hindlimb locomotor movements. Selective ES of the CF nerve resulted in restored M-responses at earlier times than in unstimulated controls in both TA and Sol muscles. Stimulated CF axons reinnervated inappropriate targets to a greater extent than unstimulated Tib axons. During locomotion, functional antagonist muscles, TA and Sol, were coactivated both in stimulated rats and in unstimulated but injured rats. Hindlimb kinematics in stimulated rats were comparable to untreated rats, but significantly different from intact controls. Selective ES promotes enhanced axon regeneration but does so with decreased fidelity of muscle reinnervation. Functional recovery is neither improved nor degraded, suggesting that compensatory changes in the outputs of the spinal circuits driving locomotion may occur irrespective of the extent of misdirection of regenerating axons in the periphery.     

Immediate electrical stimulation enhances regeneration and reinnervation and modulates spinal plastic changes after sciatic nerve injury and repair

We have studied whether electrical stimulation immediately after nerve injury may enhance axonal regeneration and modulate plastic changes at the spinal cord level underlying the appearance of hyperreflexia. Two groups of adult rats were subjected to sciatic nerve section followed by suture repair. One group (ES) received electrical stimulation (3 V, 0.1 ms at 20 Hz) for 1 h after injury. A second group served as control (C). Nerve conduction, H reflex, motor evoked potentials, and algesimetry tests were performed at 1, 3, 5, 7 and 9 weeks after surgery, to assess muscle reinnervation and changes in excitability of spinal cord circuitry. The electrophysiological results showed higher levels of reinnervation, and histological results a significantly higher number of regenerated myelinated fibers in the distal tibial nerve in group ES in comparison with group C. The monosynaptic H reflex was facilitated in the injured limb, to a higher degree in group C than in group ES. The amplitudes of motor evoked potentials were similar in both groups, although the MEP/M ratio was increased in group C compared to group ES, indicating mild central motor hyperexcitability. Immunohistochemical labeling of sensory afferents in the spinal cord dorsal horn showed prevention of the reduction in expression of substance P at one month postlesion in group ES. In conclusion, brief electrical stimulation applied after sciatic nerve injury promotes axonal regeneration over a long distance and reduces facilitation of spinal motor responses.     

Electrical stimulation restores the specificity of sensory axon regeneration

Electrical stimulation at the time of nerve repair promotes motoneurons to reinnervate appropriate pathways leading to muscle and stimulates sensory neurons to regenerate. The present experiments examine the effects of electrical stimulation on the specificity of sensory axon regeneration. The unoperated rat femoral cutaneous branch is served by 2-3 times more DRG neurons than is the muscle branch. After transection and repair of the femoral trunk, equal numbers of DRG neurons project to both branches. However, 1 h of electrical stimulation restores the normal proportion of DRG neurons reinnervating skin and muscle. To ask if the redistribution of stimulated neurons results from enhanced specificity of target reinnervation, we developed a new technique of sequential double labeling. DRG neurons projecting to the femoral muscle branch were prelabeled with Fluoro Gold 2 weeks before the nerve was transected proximally and repaired with or without 1 h of 20-Hz electrical stimulation. Three weeks after repair, the muscle nerve was labeled a second time with Fluororuby. The percentage of regenerating neurons that both originally served muscle and returned to muscle after nerve repair increased from 40% without stimulation to 75% with stimulation. Electrical stimulation thus dramatically alters the distribution of regenerating sensory axons, replacing normally random behavior with selective reinnervation of tissue-specific targets. If the enhanced regeneration specificity resulting from electrical stimulation is found to improve function in a large animal model, this convenient and safe technique may be a useful adjunct to clinical nerve repair.     

Treadmill training promotes axon regeneration in injured peripheral nerves

Physical activity after spinal cord injury promotes improvements in motor function, but its effects following peripheral nerve injury are less clear. Although axons in peripheral nerves are known to regenerate better than those in the CNS, methods of accelerating regeneration are needed due to the slow overall rate of growth. Therefore we studied the effect of two weeks of treadmill locomotion on the growth of regenerating axons in peripheral nerves following injury. The common fibular nerves of thy-1-YFP-H mice, in which a subset of axons in peripheral nerves express yellow fluorescent protein (YFP), were cut and repaired with allografts from non-fluorescent littermates, and then harvested two weeks later. Mice were divided into groups of low-intensity continuous training (CT, 60 min), low-intensity interval training (IT; one group, 10 reps, 20 min total), and high-intensity IT (three groups, 2, 4, and 10 reps). One repetition consisted of 2 min of running and 5 min of rest. Sixty minutes of CT resulted in the highest exercise volume, whereas 2 reps of IT resulted in the lowest volume of exercise. The lengths of regenerating YFP⁺ axons were measured in images of longitudinal optical sections of nerves. Axon profiles were significantly longer than control in all exercise groups except the low-intensity IT group. In the CT group and the high-intensity IT groups that trained with 4 or 10 repetitions axons were more than twice as long as unexercised controls. The number of intervals did not impact axon elongation. Axon sprouting was enhanced in IT groups but not the CT group. Thus exercise, even in very small quantities, increases axon elongation in injured peripheral nerves whereas continuous exercise resulting in higher volume (total steps) may have no net impact on axon sprouting.     

Rate and extent of functional reinnervation in fast-twitch and slow-twitch muscles of the dystrophic mouse (C57Bl6Jdy2jdy2j)

The extent of functional reinnervation of fast-twitch extensor digitorum longus muscle in dystrophic and normal mice was determined at various times after nerve transection. Functional reinnervation was assessed by measuring the twitch tension evoked by stimulation of the nerve central to the site of transection. In control mice aged 4 to 6 weeks at the time of denervation, complete reinnervation was observed after 6 weeks. In dystrophic mice of the same age reinnervation was clearly impaired. The ratio of functional innervation of the operated leg to that of the contralateral unoperated leg was only 0.62 after 6 weeks. In older dystrophic mice (4 to 6 months at the time of nerve transection) the reinnervation ratio was even lower, 0.43 after 12 weeks. Reinnervation of slow-twitch soleus muscle was assessed 8 weeks after denervation and was also found to be reduced in the older dystrophic animals. The extent of reinnervation was reflected in the measured values of muscle weight, twitch tension per unit wet weight, and twitch time course. The impairment of reinnervation of dystrophic muscle is consistent with, but not proof of, a neurogenic defect in murine muscular dystrophy.     

Thyroid hormone enhances transected axonal regeneration and muscle reinnervation following rat sciatic nerve injury

Improvement of nerve regeneration and functional recovery following nerve injury is a challenging problem in clinical research. We have already shown that following rat sciatic nerve transection, the local administration of triiodothyronine (T3) significantly increased the number and the myelination of regenerated axons. Functional recovery is a sum of the number of regenerated axons and reinnervation of denervated peripheral targets. In the present study, we investigated whether the increased number of regenerated axons by T3‐treatment is linked to improved reinnervation of hind limb muscles. After transection of rat sciatic nerves, silicone or biodegradable nerve guides were implanted and filled with either T3 or phosphate buffer solution (PBS). Neuromuscular junctions (NMJs) were analyzed on gastrocnemius and plantar muscle sections stained with rhodamine α‐bungarotoxin and neurofilament antibody. Four weeks after surgery, most end‐plates (EPs) of operated limbs were still denervated and no effect of T3 on muscle reinnervation was detected at this stage of nerve repair. In contrast, after 14 weeks of nerve regeneration, T3 clearly enhanced the reinnervation of gastrocnemius and plantar EPs, demonstrated by significantly higher recovery of size and shape complexity of reinnervated EPs and also by increased acetylcholine receptor (AChRs) density on post synaptic membranes compared to PBS‐treated EPs. The stimulating effect of T3 on EP reinnervation is confirmed by a higher index of compound muscle action potentials recorded in gastrocnemius muscles. In conclusion, our results provide for the first time strong evidence that T3 enhances the restoration of NMJ structure and improves synaptic transmission. © 2010 Wiley‐Liss, Inc.     

One hour electrical stimulation accelerates functional recovery after femoral nerve repair

The clinical outcome of peripheral nerve injuries requiring surgical repair is usually poor and efficient therapies do not exist. Recent work has suggested that low-frequency electrical stimulation of the severed nerve which produces repeated discharges of the parent motoneuron perikarya positively influences axonal regeneration, even if applied once for a period of only 1 h. Here we provide the first evidence for locomotor functional benefits of such stimulation. We transected the femoral nerve of adult C57BL/6J mice proximal to the bifurcation of the quadriceps and saphenous branches and electrically stimulated the proximal nerve stump for 1 h at 20-Hz frequency prior to nerve repair with a silicone cuff. Three months later, the ability of the quadriceps muscle to extend the knee in sham-stimulated mice had recovered to 63% of the preoperative values as estimated by single-frame motion analysis. After electrical stimulation, the outcome was only slightly better (73%) but the rate of functional recovery was considerably accelerated. Near-maximum recovery was achieved 6 weeks earlier than in the control group. The beneficial effects were associated with larger motoneuron cell bodies and increased diameters of regenerated axons in the quadriceps nerve branch, but not with enhanced preferential reinnervation by motoneurons of muscle as opposed to skin. The observed acceleration of functional restoration and the positive effects on motoneurons and regenerated axons indicate the potential of a clinically feasible approach for improvement of nerve repair outcome in human patients in which delayed target reinnervation is a factor limiting recovery.     

Functional electrical stimulation following nerve injury in a large animal model

Introduction: Controversy exists over the effects of functional electrical stimulation (FES) on reinnervation. We hypothesized that intramuscular FES would not delay reinnervation after recurrent laryngeal nerve (RLn) axonotmesis. Methods: RLn cryo-injury and electrode implantation in ipsilateral posterior cricoarytenoid muscle (PCA) were performed in horses. PCA was stimulated for 20 weeks in eight animals; seven served as controls. Reinnervation was monitored through muscle response to hypercapnia, electrical stimulation and exercise. Ultimately, muscle fiber type proportions and minimum fiber diameters, and RLn axon number and degree of myelination were determined. Results: Laryngeal function returned to normal in both groups within 22 weeks. FES improved muscle strength and geometry, and induced increased type I:II fiber proportion (p = 0.038) in the stimulated PCA. FES showed no deleterious effects on reinnervation. Discussion: Intramuscular electrical stimulation did not delay PCA reinnervation after axonotmesis. FES can represent a supportive treatment to promote laryngeal functional recovery after RLn injury. Muscle Nerve 59:717–725, 2019     

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.     

Electrophysiologic assessment of sciatic nerve regeneration in the rat: surrounding limb muscles feature strongly in recordings from the gastrocnemius muscle

Striking inconsistencies between the results of morphometric and electrophysiologic examinations of the regenerating nerve were observed in a previous study featuring the bridging of a 14 mm gap in the rat sciatic nerve. To shed light on this dichotomy, seven further rats were subjected to permanent sciatic nerve transection and assessed electrophysiologically, histologically and by retrograde axonal tracing at various postoperative intervals (1 h to 8 weeks). The results of the histological examinations and retrograde tracing revealed that in spite of the fact that compound muscle action potentials could be recorded in the gastrocnemius muscle, no reinnervation of the gastrocnemius muscle, either physiological or aberrant, had actually taken place. Furthermore, it was established that the electrical activity recorded in the gastrocnemius muscle after stimulation of the proximal or distal stump is generated by surrounding hind limb muscles unaffected by denervation. These are stimulated either directly, or indirectly due to spreading of the impulse. It is therefore strongly recommended that caution should be exercised when interpreting recordings from the gastrocnemius muscle after stimulation of a regenerating sciatic nerve in laboratory rodents.     

Electrophysiological function during voiding after simulated childbirth injuries

During vaginal delivery dual injuries of the pudendal nerve and the external urethral sphincter (EUS), along with other injuries, are correlated with later development of stress urinary incontinence. It is not known how combinations of these injuries affect neuromuscular recovery of the micturition reflex. We investigated the EUS electromyogram (EMG) and the pudendal nerve motor branch potentials (PNMBP) during voiding 4 days, 3 weeks or 6 weeks after injury; including vaginal distension (VD), pudendal nerve crush (PNC), both PNC and VD (PNC + VD), and pudendal nerve transection (PNT); and in controls. Pudendal nerve and urethral specimens were excised and studied histologically. No bursting activity was recorded in the EUS EMG during voiding 4 days after all injuries, as well as 3 weeks after PNC + VD. Bursting activity demonstrated recovery 3 weeks after either VD or PNC and 6 weeks after PNC + VD, but the recovered intraburst frequency remained significantly decreased compared to controls. Bursting results of PNMBP were similar to the EMG, except bursting in PNMBP 4 days after VD and the recovered intraburst frequency was significantly increased compared to controls after PNC and PNC + VD. After PNT, neither the EUS nor the pudendal nerve recovered by 6 weeks after injury. Our findings indicate bursting discharge during voiding recovers more slowly after PNC + VD than after either PNC or VD alone. This was confirmed histologically in the urethra and the pudendal nerve and may explain why pudendal nerve dysfunction has been observed years after vaginal delivery.     

Collateral sprouting of sensory axons after end-to-side nerve coaptation--a longitudinal study in the rat

The end-to-side nerve coaptation is able to induce collateral sprouting of axons from the donor nerve and to provide functional reinnervation of the target tissue. Sensory axon sprouting and its effects on the donor nerve up to 9 months after the end-to-side nerve coaptation were studied in the rat. Peroneal, tibial and saphenous nerves were transected and ligated, and the distal stump of the transected peroneal nerve was sutured to the side of the uninjured sural nerve. The average skin area of the residual sensitivity to pinch due to the axons sprouting through the recipient peroneal nerve did not change statistically significantly between 4 and 9 months after surgery. Axon counting, measurements of compound action potentials and retrograde neuron labeling indicate that the sprouting of the myelinated sensory axons and unmyelinated axons through the recipient nerve was largely completed by 2 months and 4 months after the end-to-side nerve coaptation, respectively, and remained stable thereafter for at least 9 months. A decrease in the amplitude and area of the CAP of myelinated fibers, observed in the donor nerve up to 4 months after surgery, was probably due to mild degeneration of nerve fibers and a tendency of the diameter of myelinated axons to decline. However, no significant changes in functional, electrophysiological or morphological properties of the donor nerve could be observed at the end of the observational period, indicating that end-to-side nerve coaptation has no detrimental effect on the donor nerve on a long-term scale.     

Effect of locomotor training on muscle performance in the context of nerve-muscle communication dysfunction

Introduction: The effects of locomotor training (LT) on skeletal muscle after peripheral nerve injury and acetylcholinesterase deficiency are not well documented. Methods: We determined the effects of LT on mouse soleus muscle performance after sciatic nerve transection with excision (full and permanent denervation), nerve transection (partial functional reinnervation), nerve crush (full denervation with full functional reinnervation), and acetylcholinesterase deficiency (alteration in neuromuscular junction functioning). Results: We found no significant effect of LT on the recovery of soleus muscle weight, maximal force in response to muscle stimulation, and fatigue resistance after nerve transection with or without excision. However, LT significantly increased soleus muscle fatigue resistance after nerve crush and acetylcholinesterase deficiency. Moreover, hindlimb immobilization significantly aggravated the deficit in soleus muscle maximal force production and atrophy after nerve crush. Conclusions: LT is beneficial, and reduced muscle use is detrimental for intrinsic muscle performance in the context of disturbed nerve–muscle communication. Muscle Nerve, 2012     

Neurotrophic factors improve muscle reinnervation from embryonic neurons

Motoneurons die in diseases like amyotrophic lateral sclerosis and after spinal cord trauma, inducing muscle denervation. We tested whether transplantation of embryonic cells with neurotrophic factors into peripheral nerve of adult rats improves muscle reinnervation and motor unit function more than cells alone. One week after sciatic nerve section, embryonic ventral spinal cord cells were transplanted into the tibial nerve with or without glial cell line-derived neurotrophic factor, hepatocyte growth factor, and insulin-like growth factor-1. These cells represented the only neuron source for muscle reinnervation. Ten weeks after transplantation, all medial gastrocnemius muscles contracted in response to electrical stimulation of cell transplants with factors. Only 80% of muscles responded with cells alone. Factors and cells resulted in survival of more motoneurons and reinnervation of more muscle fibers for a given axon (motor unit) number. Greater reinnervation from embryonic cells may enhance muscle excitation by patterned electrical stimulation.     

Electrical Stimulation to Enhance Axon Regeneration After Peripheral Nerve Injuries in Animal Models and Humans

Injured peripheral nerves regenerate their lost axons but functional recovery in humans is frequently disappointing. This is so particularly when injuries require regeneration over long distances and/or over long time periods. Fat replacement of chronically denervated muscles, a commonly accepted explanation, does not account for poor functional recovery. Rather, the basis for the poor nerve regeneration is the transient expression of growth-associated genes that accounts for declining regenerative capacity of neurons and the regenerative support of Schwann cells over time. Brief low-frequency electrical stimulation accelerates motor and sensory axon outgrowth across injury sites that, even after delayed surgical repair of injured nerves in animal models and patients, enhances nerve regeneration and target reinnervation. The stimulation elevates neuronal cyclic adenosine monophosphate and, in turn, the expression of neurotrophic factors and other growth-associated genes, including cytoskeletal proteins. Electrical stimulation of denervated muscles immediately after nerve transection and surgical repair also accelerates muscle reinnervation but, at this time, how the daily requirement of long-duration electrical pulses can be delivered to muscles remains a practical issue prior to translation to patients. Finally, the technique of inserting autologous nerve grafts that bridge between a donor nerve and an adjacent recipient denervated nerve stump significantly improves nerve regeneration after delayed nerve repair, the donor nerves sustaining the capacity of the denervated Schwann cells to support nerve regeneration. These reviewed methods to promote nerve regeneration and, in turn, to enhance functional recovery after nerve injury and surgical repair are sufficiently promising for early translation to the clinic.     

Electrical stimulation promotes sensory neuron regeneration and growth-associated gene expression

Brief electrical stimulation enhances the regenerative ability of axotomized motor [Nix, W.A., Hopf, H.C., 1983. Electrical stimulation of regenerating nerve and its effect on motor recovery. Brain Res. 272, 21-25; Al-Majed, A.A., Neumann, C.M., Brushart, T.M., Gordon, T., 2000. Brief electrical stimulation promotes the speed and accuracy of motor axonal regeneration. J. Neurosci. 20, 2602-2608] and sensory [Brushart, T.M., Jari, R., Verge, V., Rohde, C., Gordon, T., 2005. Electrical stimulation restores the specificity of sensory axon regeneration. Exp. Neurol. 194, 221-229] neurons. Here we examined the parameter of duration of stimulation on regenerative capacity, including the intrinsic growth programs, of sensory neurons. The effect of 20 Hz continuous electrical stimulation on the number of DRG sensory neurons that regenerate their axons was evaluated following transection and surgical repair of the femoral nerve trunk. Stimulation was applied proximal to the repair site for 1 h, 3 h, 1 day, 7 days or 14 days at the time of nerve repair. Following a 21-day regeneration period, DRG neurons that regenerated axons into the muscle and cutaneous sensory nerve branches were retrogradely identified. Stimulation of 1 h led to a significant increase in DRG neurons regenerating into cutaneous and muscle branches when compared to 0 h (sham) stimulation or longer periods of stimulation. Stimulation for 1 h also significantly increased the numbers of neurons that regenerated axons beyond the repair site 4 days after lesion and was correlated with a significant increase in expression of growth-associated protein 43 (GAP-43) mRNA in the regenerating neurons at 2 days post-repair. An additional indicator of heightened plasticity following 1 h stimulation was elevated expression of brain-derived neurotrophic factor (BDNF). The effect of brief stimulation on enhancing sensory and motoneuron regeneration holds promise for inducing improved peripheral nerve repair in the clinical setting.     

Treadmill training enhances axon regeneration in injured mouse peripheral nerves without increased loss of topographic specificity

We investigated the extent of misdirection of regenerating axons when that regeneration was enhanced by using treadmill training. Retrograde fluorescent tracers were applied to the cut proximal stumps of the tibial and common fibular nerves 2 or 4 weeks after transection and surgical repair of the mouse sciatic nerve. The spatial locations of retrogradely labeled motoneurons were studied in untreated control mice and in mice receiving 2 weeks of treadmill training, according to either a continuous protocol (10 m/minute, 1 hour/day, 5 days/week) or an interval protocol (20 m/minute for 2 minutes, followed by a 5‐minute rest, repeated four times, 5 days/week). More retrogradely labeled motoneurons were found in both treadmill‐trained groups. The magnitude of this increase was as great as or greater than that found after using other enhancement strategies. In both treadmill‐trained groups, the proportions of motoneurons labeled from tracer applied to the common fibular nerve that were found in spinal cord locations reserved for tibial motoneurons in intact mice were no greater than in untreated control mice and significantly less than those found after electrical stimulation or chondroitinase treatment. Treadmill training in the first 2 weeks following peripheral nerve injury produces a marked enhancement of motor axon regeneration without increasing the propensity of those axons to choose pathways leading to functionally inappropriate targets. J. Comp. Neurol. 517:245–255, 2009. © 2009 Wiley‐Liss, Inc.     

BDNF/TrkB signaling regulates HNK-1 carbohydrate expression in regenerating motor nerves and promotes functional recovery after peripheral nerve repair

Functional recovery after peripheral nerve injury is often poor despite high regenerative capacity of peripheral neurons. In search for novel treatments, brief electrical stimulation of the acutely lesioned nerve has recently been identified as a clinically feasible approach increasing precision of axonal regrowth. The effects of this stimulation appear to be mediated by BDNF and its receptor, TrkB, but the down-stream effectors are unknown. A potential candidate is the HNK-1 carbohydrate known to be selectively reexpressed in motor but not sensory nerve branches of the mouse femoral nerve and to enhance growth of motor but not sensory axons in vitro. Here, we show that short-term low-frequency electrical stimulation (1 h, 20 Hz) of the lesioned and surgically repaired femoral nerve in wild-type mice causes a motor nerve-specific enhancement of HNK-1 expression correlating with previously reported acceleration of muscle reinnervation. Such enhanced HNK-1 expression was not observed after electrical stimulation in heterozygous BDNF or TrkB-deficient mice. Accordingly, the degree of proper reinnervation of the quadriceps muscle, as indicated by retrograde labeling of motoneurons, was reduced in TrkB+/- mice compared to wild-type littermates. Also, recovery of quadriceps muscle function, evaluated by a novel single-frame motion analysis approach, and axonal regrowth into the distal nerve stump, assessed morphologically, were considerably delayed in TrkB+/- mice. These findings indicate that BDNF/TrkB signaling is important for functional recovery after nerve repair and suggest that up-regulation of the HNK-1 glycan is linked to this phenomenon.     

Electrical stimulation impairs early functional recovery and accentuates skeletal muscle atrophy after sciatic nerve crush injury in rats

Neuromuscular recovery after peripheral nerve lesion depends on the regeneration of severed axons that re‐establish their functional connection with the denervated muscle. The aim of this study was to determine the effects of electrical stimulation (ES) on the neuromuscular recovery after nerve crush injury in rats. Electrical stimulation was carried out on the tibialis anterior (TA) muscle after sciatic nerve crush injury in a rat model. Six ES sessions were administered every other day starting from day 3 postinjury until the end of the experiment (day 14). The sciatic functional index was calculated. Muscle excitability, neural cell adhesion molecule (N‐CAM) expression, and muscle fiber cross‐sectional area (CSA) were accessed from TA muscle. Regenerated sciatic nerves were analyzed by light and confocal microscopy. Both treated (crush+ES) and untreated (crush) groups had their muscle weight and CSA decreased compared with the normal group (P < 0.05). Electrical stimulation accentuated muscle fiber atrophy more in the crush+ES than in the crush group (P < 0.05). N‐CAM expression increased in both crush and crush+ES groups compared with the normal group (P < 0.05). Regenerated nerves revealed no difference between the crush and crush+ES groups. Nevertheless, functional recovery at day 14 post‐injury was significantly lower in crush+ES group compared with the crush group. In addition, the crush+ES group had chronaxie values significantly higher on days 7 and 13 compared with the crush group, which indicates a decrease in muscle excitability in the crush+ES animals. The results of this study do not support a benefit of the tested protocol of ES during the period of motor nerve recovery following injury. Muscle Nerve, 2010     

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.