Selectivity of distal reinnervation of regenerating mixed motor and sensory nerve fibres across muscle grafts in rats

This study investigated target specificity during axonal regeneration of a mixed motor and sensory nerve towards respective targets. The femoral nerves in rats were divided and allowed to grow across a 6 mm gap interposed with frozen and thawed muscle grafts towards their distal motor and sensory nerve stumps. Fourteen weeks later the number of motoneurons projecting axons into the motor and sensory branches were determined by retrograde axonal tracing using horse-radish peroxidase. There were significantly higher numbers of motoneurons (p = 0.0034) projecting into the motor nerve than the sensory nerve. Efferent axons of a mixed nerve selectivity grew into motor branches when allowed to regenerate across a 6 mm gap interposed with muscle grafts. It is possible that a deliberately created 'structured gap' during repair of mixed nerves could improve axonal matching by allowing expression of neurotropism.     

Differential response of sensory and motor axons in nerve allografts after withdrawal of immunosuppressive therapy

OBJECT: Rejection of nerve allografts and loss of regenerated host axons after withdrawal of immunosuppressive therapy poses an ongoing challenge in peripheral nerve repair. The present report is of a blinded prospective controlled study in which an established rat model of nerve allotransplantation is used to examine the effect of fiber type on survival and degeneration of nerve allografts after discontinuation of immunosuppression. The authors hypothesized that sensory axons will selectively resist a rejection response, whereas motor axons will degenerate. METHODS: Four-centimeter nerve segments from ACI rats were grafted into peroneal and sural (mixed) or saphenous (sensory) nerve gaps in Lewis rats. In some rats, L4-6 dorsal root ganglia were ablated before grafting, creating pure motor sural and peroneal nerves. All rats received 12 weeks of immunosuppressive therapy to support nerve regeneration into allografts. Immunosuppression with cyclosporin was then withdrawn. At planned death (12-18 weeks postsurgery), graft tissue was subjected to histomorphometric analysis for evaluation of axon survival and loss. Graft rejection led to loss of all axons in approximately 60% of the allograft segments. The mixed nerve group was most prone to complete rejection, with significantly lowered axon counts at Weeks 16 and 18 compared with the Week 12 baseline. Axons from the sensory nerve were least likely to degenerate. The pure motor nerve group axons demonstrated intermediate sensitivity, with a selective loss of larger axons at Week 16 and a significant decrease in axon counts from the Week 12 baseline at Week 18. CONCLUSIONS: Whereas the majority of axons are lost after withdrawal of immunosuppressive therapy from nerve allografts, there is a selective survival of axons from cutaneous sensory nerves and smaller-diameter motor fibers. The biological and molecular mechanisms that make some axons impervious to injury remain to be determined.     

Selective regeneration of motor and sensory axons in an experimental peripheral nerve model without endorgans.

We assessed the selectivity of motor and sensory axon regeneration towards the distal motor and sensory nerve segments that were disconnected from endorgans in a rat silicone Y chamber model. The L5 ventral root was used as a pure motor nerve, and the saphenous nerve was used as a sensory nerve. In experiment 1 (n=11), the proximal stump of the L5 ventral root, a 1-cm-long L5 ventral root segment and a saphenous nerve segment were inserted into a silicone Y chamber. In experiment 2 (n=11), the proximal stump of the saphenous nerve, a L5 ventral root segment and a saphenous nerve segment were inserted into a Y chamber. The distance between the nerve stumps was 5 mm. Six weeks later, the number of regenerated myelinated motor and sensory axons was measured and compared in the distal two channels. Motor axons showed no selective regeneration, but sensory axons did.     

Processed nerve allografts for peripheral nerve reconstruction: a multicenter study of utilization and outcomes in sensory, mixed, and motor nerve reconstructions

Purpose: As alternatives to autograft become more conventional, clinical outcomes data on their effectiveness in restoring meaningful function is essential. In this study we report on the outcomes from a multicenter study on processed nerve allografts (Avance® Nerve Graft, AxoGen, Inc). Patients and Methods: Twelve sites with 25 surgeons contributed data from 132 individual nerve injuries. Data was analyzed to determine the safety and efficacy of the nerve allograft. Sufficient data for efficacy analysis were reported in 76 injuries (49 sensory, 18 mixed, and 9 motor nerves). The mean age was 41 ± 17 (18–86) years. The mean graft length was 22 ± 11 (5–50) mm. Subgroup analysis was performed to determine the relationship to factors known to influence outcomes of nerve repair such as nerve type, gap length, patient age, time to repair, age of injury, and mechanism of injury. Results: Meaningful recovery was reported in 87% of the repairs reporting quantitative data. Subgroup analysis demonstrated consistency, showing no significant differences with regard to recovery outcomes between the groups (P > 0.05 Fisher's Exact Test). No graft related adverse experiences were reported and a 5% revision rate was observed. Conclusion: Processed nerve allografts performed well and were found to be safe and effective in sensory, mixed and motor nerve defects between 5 and 50 mm. The outcomes for safety and meaningful recovery observed in this study compare favorably to those reported in the literature for nerve autograft and are higher than those reported for nerve conduits. © 2011 Wiley Periodicals, Inc. Microsurgery, 2012.     

Preferential block of small myelinated sensory and motor fibers by lidocaine: in vivo electrophysiology in the rat sciatic nerve.

BACKGROUND: Controversy still surrounds the differential susceptibility of nerve fibers to local anesthetics and its relation to selective functional deficits. In the current study we report features of conduction blockade in different classes of rat sciatic nerve fibers after injection of lidocaine by a percutaneous procedure that closely resembles clinical applications. METHODS: In 30 adult male Sprague-Dawley rats (weight, 300-400 g) during general anesthesia, impulses were recorded in different classes of sensory axons (large, Aalpha and beta fibers; small, Adelta myelinated fibers and unmyelinated C fibers) and motor axons (large, Aalpha fibers; small, Agamma myelinated fibers) classified by conduction velocity. The sciatic nerve was stimulated distally, and impulses were recorded from small filaments teased from L4-L5 dorsal (sensory) and ventral (motor) roots sectioned acutely from the spinal cord. Lidocaine at concentration of 0.05-1% was injected percutaneously in 0.1-ml solutions at the sciatic notch. Both tonic (stimulated at 0.5 Hz) and use-dependent (stimulated at 40 Hz for Adelta and Agamma fibers and at 5 Hz for C fibers) impulse inhibitions by lidocaine were assayed. RESULTS: Minimal effective (threshold) lidocaine concentrations (i.e., to block conduction in 10% of fibers) were, for sensory, 0.03% for Adelta, 0.07% for Aalphabeta, and 0.09-0.1% for C fibers, and for motor, 0.03% for Agamma and 0.05% for Aalpha fibers. The order of fiber susceptibility, ranked by concentrations that gave peak tonic fiber blockade of 50% (IC50s), was Agamma > Adelta = Aalpha > Aalphabeta > C. Faster-conducting C fibers (conduction velocity > 1 m/s) were more susceptible (IC50 = 0.13%) than slower ones (conduction velocity < 1 m/s; IC50 = 0.30%). At 1% lidocaine, all fibers were tonically blocked. Use-dependent effects accounted for only a modest potentiation of block (at a lidocaine concentration of 0.25%) in Adelta and Agamma fibers, and in C fibers phasic stimulation had even smaller effects and sometimes relieved tonic block. CONCLUSIONS: Susceptibility to lidocaine does not strictly follow the "size principle" that smaller (slower) axons are always blocked first. This order of fiber blockade is qualitatively consistent with previous reports of the order of functional deficits in the rat after percutaneous lidocaine, that is, motor = proprioception > nociception, if we assume that motor deficits first arise from conduction failure in Agamma fibers and that nociception relies on C fiber conduction.     

大鼠腓总神经"π"式桥接于胫神经再生的原配连接

目的研究大鼠腓总神经被切断桥接于胫神经后，再生的腓总神经纤维的来源。方法将大鼠右侧腓总神经在胭窝处剪除约5mm后，将断裂的腓总神经近端和远端分别就近与同侧胫神经上的外膜窗施行端侧吻合，吻合口间距约10mm；动物存活24个月后，将手术侧近端吻合口以上胫神经剪断，再将辣根过氧化物酶注入远段腓总神经干内，72h后取材并经四甲基联苯胺显色显示脊髓L3-6节段神经元和L4、L5脊神经节细胞，同时取腓总神经远段行电镜观察神经纤维再生状态。结果远段腓总神经再生神经纤维清晰、明显，神经纤维比正常者略细；脊髓L3-6节段和L4、L5脊神经节均见到蓝染细胞。结论断裂腓总神经“π”式桥接于胫神经，至少有部分再生神经纤维经腓总神经近段来源于原配神经的胞体。     

A comparison of the passage of regenerating axons through old degenerated nerve autografts and fresh nerve autografts in rats

Summary We have compared the passage of regenerating axons through old degenerated nerve autografts and fresh nerve autografts in two groups of Sprague Dawley rats. In one group the right sciatic nerve was divided and repaired with the graft taken from the distal segment of the left sciatic nerve, which had been severed 15 weeks before. In the other group the right sciatic nerve was repaired with the graft taken from the intact left sciatic nerve. Eight weeks later the myelinated axons were counted at three different locations in the grafted right sciatic nerve in both groups. The old degenerated nerve autografts were less effective in allowing the passage of regenerating axons than the fresh nerve autografts, but the difference was not great. This provides justification for the use of old degenerated nerve as an additional graft where a large number of autogenous nerve grafts are required, such as in the operative treatment of injuries to the brachial plexus.

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Résumé Les auteurs ont effectué un travail destiné à évaluer la possibilité du passage d'axones de régénération au travers d'autogreffes de nerfs dévitalisés depuis un certain temps, comparativement à celui d'autogreffes nerveuses fraîches. Dans ce but, une expérimentation a été menée sur deux groupes de rats blancs Sprague Dawley. Dans un des deux groupes, on a sectionné les nerfs sciatiques droits et on y a ensuite greffé les segments distaux de nerfs sciatiques gauches prélevés quinze semaines auparavant. Dans l'autre groupe, on a greffé sur les nerfs sciatiques droits sectionnés des segments prélevés sur les nerfs sciatiques gauches intacts. Après 8 semaines, on a comparé dans les deux groupes les axones myélinisés à trois endroits différents des nerfs sciatiques droits greffés: l'extrémité distale du segment proximal, le milieu de la greffe et l'extrémité proximale du segment distal. Les résultats ont montré que la capacité du nerf dégénéré à laisser passer les axones en régénérescence est 0.79 fois inférieure à celle du nerf frais (p[t]<0.05). Cependant la différence n'est pas très importante. Ceci signifie que des nerfs dégénérés peuvent être utilisés commes greffes complémentaires lorsqu'un grand nombre de greffes nerveuses autogènes est nécessaire, comme c'est le cas pour le traitement chirurgical des lésions du plexus brachial.

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Reprint requests to: H.J. Kim     

感觉性和运动性神经来源许旺细胞神经生长因子的表达

目的;研究感觉性和运动性许旺细胞神经生长因子的表达是否有区别,进而探讨对神经特异性再生的影响。方法：体外培养高纯度的运动性许旺细胞,每种细胞分别在常规和添加白细胞介素－1培养基中培养,用双抗体夹心ELISA法测量不同时间培养培养基中神经生长因子表达量。结果：两种培养条件一,感觉性许旺细胞的神经生长均呈双峰状,第二峰较第一峰明显增高,运动性许旺细胞的表达在第6、7d达峰值,但总体水平较感觉性许旺细胞为低。结论：两种许旺细胞神经生长因子的表达量和特点具有差别,这种差别可以为解释神经特异性再生提供依据。     

大鼠腓总神经“π”式桥接于胫神经后再生神经的电生理溯源

目的 研究大鼠胫神经原位桥接切断的腓总神经，观察腓总神经再生程度、神经纤维的来源等。方法 将断裂的腓总神经近端和远端分别就近与胫神经施行端侧吻合，存活18个月后，电生理检测再生神经纤维的动作电位传导，取腓总神经远段行光镜及电镜观察神经纤维再生数量及状态。结果 远段腓总神经有明显的神经纤维再生，远段腓总神经通过邻近神经的桥接与近段腓总神经之间有动作电位传导。结论 断裂腓总神经“π”式桥接于胫神经，部分再生神经纤维可能来源于原腓总神经近段，部分来自胫神经。     

Both stump area and volume of distal sensory nerve segments influence the regeneration of sensory axons in rats.

We examined the influence of both stump area and volume of a distal sensory nerve segment on neurotropic induction of regenerating sensory axons in a rat saphenous nerve model. In group 1 (n = 10) the proximal stump of the severed saphenous nerve was inserted into the proximal channel, and a 2 cm free nerve segment and a double-barrelled 1 cm free nerve segment were inserted into the distal two channels of a silicone Y-chamber. In group 2 (n = 10), 2 cm and 1 cm free nerve segments were inserted into the distal two channels of a Y-chamber. The gap between the stumps was set at 4 mm. After six weeks, we counted and compared the number of regenerated myelinated sensory axons in the distal two channels. Significantly more axons regenerated in the wider stump area channel of group 1 and in the larger volume channel of group 2 than in the opposite channel in either group (p < 0.05 in each case).     

Use of an intact sensory nerve to bridge a motor nerve defect: an experimental study

OBJECT: End-to-side neurorrhaphy has recently became popular for peripheral nerve repair. Although this method is mainly indicated in nerve defects in which there is an absent proximal nerve stump, bridging a motor nerve defect by coapting the proximal and distal ends of the defect to a neighboring mixed nerve in an end-to-side fashion has been another experimental use of this method. In this situation, however, the source of the regenerating axons is unclear because the axons in both the proximal end of the defect and the bridging intact nerve have the capacity for regeneration. The goal of this study was to identify the source of the regenerating axons. METHODS: In this experimental study, the authors used a sensory nerve to bridge a motor nerve defect so that they could elucidate the source of the regenerating motor axons in the distal part of the motor nerve. One advantage of using a sensory nerve was that it eradicated the risk of damaging another motor nerve. Tests used in the analysis included gait evaluation, electrophysiological tests, and histological assessment. CONCLUSIONS: Results of this study showed that, in the rat model, a sensory nerve can be used to bridge a motor nerve defect, thereby eliminating the need for nerve grafting.     

Reverse end-to-side neurotization in a regenerating nerve

Bypass grafting around a neuroma-in-continuity entails coapting a nerve graft above and below the injured segment using two sequential end-to-side repairs. The proximal repair is analogous to what has been classically described as an end-to-side repair; the axons from the intact nerve sprout into the end of a recipient nerve and travel distally. At the distal connection, however, axons in the graft must enter the side of the intact nerve and find their way to appropriate end organs. This process has not been well investigated. To examine this, a reverse end-to-side repair, suturing the distal end of the peroneal nerve to the side of a transected and repaired tibial nerve, was performed in 20 rats. A primary end-to-end repair of the tibial nerve was performed in 10 additional rats. Twelve weeks later, contraction forces of the gastrocnemius muscle were measured following proximal stimulation. Measurements were repeated following elimination of potential axonal pathways to identify which axons (peroneal or tibial) had achieved greater reinnervation. The results indicated that both groups of axons had achieved significant reinnervation. This study supports the idea that a reverse end-to-side repair can result in axonal invasion of an intact but regenerating nerve and achieve functional recovery.     

Correlation between target reinnervation and distribution of motor axons in the injured rat sciatic nerve

Peripheral nerve injuries are rarely followed by complete return of function. Deficits are particularly important for motor function, resulting in paralysis and muscle atrophy. In different groups, the sciatic nerve was either crushed or transected and repaired by direct suture or by tube repair using silicone or collagen tubes. After 60 days, nerve regeneration was assessed by electrophysiological and functional tests, nerve morphology and immunohistochemistry against choline acetyltransferase (ChAT) for labeling motor axons. Suture and tube repair resulted in similar levels of muscle reinnervation, but significantly lower than after nerve crush. Recovery of walking track pattern was poor in all groups after nerve section. The numbers of regenerated myelinated fibers and of ChAT+ fibers were similar to control values after nerve crush, but increased after section and repair. The normal fascicular architecture and grouping of ChAT+ fibers were maintained after nerve crush, but lost after section and repair, where motor fibers were scattered within small regenerated fascicles throughout the nerve. The loss of fascicular organization was related to the deficient recovery of locomotor function. Thus, labeling of motor axons by ChAT immunohistochemistry provides useful information for the study of the degree and specificity of nerve regeneration.     

Subblocking concentrations of local anesthetics: effects on impulse generation and conduction in single myelinated sciatic nerve axons in frog.

Phenomena seen in axons exposed to subblocking doses serve as the basis for interpreting clinical and behavioral observations during onset and recovery of peripheral nerve block. To delineate the changes in excitability and in impulse conduction caused by subblocking concentrations of local anesthetics (LAs) in myelinated peripheral nerve fibers, LAs were applied to excised frog sciatic nerves while impulse conduction was monitored in single axons. For concentrations ranging from 0.01 to 1.2 times the LA concentration needed to block impulse conduction, three measures of susceptibility to LA were made to quantify the action of the drugs on "resting" fibers (firing rates < or = 0.5 Hz): the increase in the threshold for electrical activation of impulses, the increase in conduction latency reflecting the slowing of impulse conduction in the region exposed to LA, and the "critical blocking concentration" of LA just sufficient to prevent impulse conduction in the recorded fiber. Wide interfiber variation in these variables was observed (e.g., for lidocaine, latency increases at block ranged from 66% to 257% of control, blocking concentrations ranged from 0.29 to 1.40 mM), which was not correlated with fiber diameter (as indicated by resting conduction velocity). Mathematical modeling of impulse conduction in fibers exposed to LA demonstrated that the interfiber variation in susceptibility to LA block could result from interfiber differences in the density of sodium and potassium channels. The effects of LA were also studied in active fibers (firing rates > 0.5 Hz). Local anesthetics reversibly inhibited two normally occurring afteroscillations in membrane threshold related to afterpotentials following an impulse. These were "superexcitability," a transient lowering of threshold lasting as long as 1 s, and "depression," a phase of raised threshold peaking within 2-4 s after an impulse and recovering slowly over several minutes. Impulse activity also transiently increased the apparent potency of LAs. Such "use-dependent" increases in threshold and decreases in conduction velocity showed kinetics that were agent specific, lasting 1 s after a burst of impulses for lidocaine and lasting > 10 s for bupivacaine. At low concentrations, within the range of nontoxic plasma concentrations after systemic administration, the predominant actions of LAs on conducting fibers were transient decreases in excitability and conduction velocity in combination with a reduction of intrinsic oscillatory aftereffects of impulse discharge. These effects may degrade decoding of information in discharge patterns without actually blocking conduction of infrequent impulses, suggesting how functional blockade of coordinated movement and perception may occur even without complete blockade of impulse conduction.     

感觉/运动神经切断影响大鼠骨折愈合的实验研究

[目的]通过建立切断组成大鼠坐骨神经的不同成分、联合胫骨骨折的动物模型,来研究感觉/运动神经损伤对骨折愈合的影响,并进一步探讨周围神经系统对骨组织的生理作用。[方法]Wistar大鼠90只平均分为3组：对照组,运动神经（前根）切断组,感觉神经（后根）切断组,选择性切断一侧L4～6神经根。所有大鼠于神经根暴露侧/切断侧胫骨造成横行骨折,髓钉固定。30d后取大鼠双侧胫骨,测患侧胫骨近端骨密度、在影像学资料上测量患侧骨痂宽度及健侧胫骨相应位置的皮质宽度、取所有大鼠双侧胫骨行湿重称量、骨痂组织学切片观察、双侧胫骨行生物力学拉伸测试。所得结果均以患侧数据除以相应健侧数据得出一相对数,以相对数数据进行统计分析。[结果]患侧胫骨近端骨密度无显著差异（P〉0.05）;检测项目示神经切断组骨折处骨痂形成多,力学强度差,骨痂成熟度差（P〈0.05）,以感觉神经（后根）切断组尤甚（P〈0.05）[结论]感觉神经纤维对骨折愈合的影响较运动神经纤维显著;完整的神经支配是正常骨折愈合的必要条件。     

Effect of infra-red low-power laser irradiation on regeneration of myelinated axons

The purpose of the present study was to examine the regeneration of myelinated axons under the effect of laser irradiation at various wavelengths and energy densities. Laser irradiation at 890 nm with an energy dose of 0.33 J cm^–2 as well as He-Ne laser irradiation failed to change the number of regenerating myelinated axons in distal nerve stumps on the 30th day after cutting the nerve. An increase of dose delivered to the skin to 9.33 J cm^–2 resulted in a 49% decrease in the number of myelinated axons. A 24% suppression of nerve regeneration was also registered using 1220 nm wavelength laser (dose 0.98 J cm^–2). This phenomenon is likely to be attributed to the stimulating effect of laser irradiation of the near-infra-red range on proliferation of fibroblasts and scar formation. 1220 nm of 7.2 J cm^–2 dose effected neither the growth nor the myelinization of axons in distal nerve stumps on the 20th day following nerve damage.