Toe-spreading Reflex Research Articles

Morphological and functional evaluation of the effect of novel pyrimidine derivatives on regeneration of the sciatic nerve in rats

Two novel pyrimidine derivatives, RG2 and RG6, were studied using a rat’s model of peripheral nerve injury. Toe-spreading reflex and skin sensitivity to pinch in the foot were monitored to follow recovery of motor and sensory functions in the treated animals. The remyelation rate in the distal segment of the damaged nerve was also studied using morphological analysis of cross-sections of the nerve stained with methylene blue. The obtained data demonstrate a high stimulating effect of RG2 and RG6 on the restoration of motor and sensory functions of the sciatic nerve, as well as on the post-traumatic regeneration of myelin fibers. Possible mechanisms of the observed effects are discussed.

The Changes in Rats with Sciatic Nerve Crush Injury Supplemented with Evening Primrose Oil: Behavioural, Morphologic, and Morphometric Analysis.

Nerve crush injuries are commonly used models for axonotmesis to examine peripheral nerve regeneration. As evening primrose oil (EPO) is rich in omega-6 essential fatty acid component and gamma-linolenic acid, studies have shown the potential role of EPO in myelination. Seventy-two healthy adult Sprague-Dawley rats were classified into three groups: normal group, control group, and experimental group. The result indicates that there was significant difference in toe-spreading reflex between the normal and the control groups (1.9 ± 0.031, p < 0.05) and the normal and the EPO groups (0.4 ± 0.031, p < 0.05) and significant difference between EPO and the control groups (1.5 ± 0.031, p < 0.05). Regeneration of axons and myelin in nerve fibre in the EPO-treated group developed better and faster than in the control group. In the control group, the shape of the axon was irregular with a thinner myelin sheath. In the experimental group, the shape of the axons, the thickness of the myelin sheath, and the diameter of the axons were almost the same as in the normal group. In conclusion, EPO supplementation may be beneficial as a therapeutic option for disturbances of nerve interaction.

Enhanced axonal regeneration of the injured sciatic nerve by administration of Buyang Huanwu decoction

Ethnopharmacological relevanceBuyang Huanwu decoction (BYHWD) has been used in the traditional Chinese medicine for the treatment of cardiovascular and neurological symptoms, and recent experimental studies have begun to provide evidence showing its protective effects on neural cells. Yet, its function for the regenerative responses of axons in the peripheral nerve after injury is not known. Aim of the studyThe primary objective of the present study was to explore that BYHWD is involved in growth-promoting activity of the peripheral nerve axons after injury. We further examined whether the effect of BYHWD exerted directly on regrowing axons or Schwann cells. Materials and methodsSciatic nerves in rats were given crush injury, and BYHWD was injected by oral administration. Sciatic nerves or DRG tissues were prepared for immunofluorescence staining and western blot analysis. Levels of axonal regeneration were quantified by retrograde tracing technique. Cultured DRG sensory neurons and Schwann cells were prepared from rats and used to examine the effects of BYHWD on the neurite outgrowth. Behavioral analysis on functional recovery after nerve injury was assessed in mice by pin prick test, adhesive removal test, and toe-spreading reflex. ResultsImmunofluorescence and retrograde tracing analyses showed that the distal extension of the sciatic nerve axons was significantly improved by BYHWD treatment. Levels of axonal growth-associated protein GAP-43 were upregulated by BYHWD treatment in the sciatic nerve after injury and in the neurites of cultured DRG neurons. In vivo administration of BYHWD in rats upregulated the induction level of cell division cycle 2 (Cdc2) and its phosphorylation of vimentin in Schwann cells from injured sciatic nerve. Coculture of DRG neurons with Schwann cells prepared from preinjured sciatic nerves in animals administered with BYHWD led to the enhancement in neurite outgrowth. Behavioral tests in mice given sciatic nerve injury showed a significant improvement in sensorimotor activity by BYHWD administration. ConclusionsOur results suggest that BYHWD administration into animals given sciatic nerve injury facilitates axonal regeneration by acting on both the axons undergoing regeneration and neighboring Schwann cells and improves functional recovery.

In vivo evaluation of sciatic nerve crush injury using diffusion tensor imaging: correlation with nerve function and histology.

The aim of this study was to prospectively estimate magnetic resonance diffusion tensor imaging parameters in distal portions of crushed sciatic nerves in rabbits and correlation with neurological function and histology. Thirty-two rabbits were randomly divided into 8 groups and followed up at 10 weeks. The right sciatic nerves were crushed, and the left sciatic nerves served as the control group with sham operation. Another 4 rabbits were chosen in the normal control group. Diffusion tensor imaging scan was obtained on bilateral sciatic nerves using spin-echo single-shot echo planar imaging. The values of fractional anisotropy (FA), apparent diffusion coefficient, and eigenvalue (λ║ and λ⊥) were measured on diffusion tensor imaging. Quantitative assessment of functional recovery in crushed nerves included toe-spreading reflex and modified Tarlov score, as well as pathology of the injured nerves. Fractional anisotropy of distal portions in injured nerves reduced slightly on the first day and dropped to the minimum at the fourth day after surgery. Then, FA increased gradually from the eighth day to the eighth week. After that, FA recovered nearly normal in the 10th week after injury. There was statistical significance in FA during 4 days to the eighth week and λ⊥ values between distal portions of injured nerves and normal control nerves (P < 0.05). Fractional anisotropy of distal portions to injured nerves correlated significantly with the nerve function score(r = 0.898, P < 0.01), whereas λ⊥ of distal portions of injured nerves demonstrated negative correlation with the nerve function score(r = -0.820, <0.01). Fractional anisotropy values of distal portions of injured nerves formed a similar time course as functional recovery, whereas λ⊥ indicated a opposite trend. The changes in FA and λ⊥ of distal portions of injured sciatic nerves correlate well with functional recovery and histology. Therefore, FA and λ⊥ of distal portions of injured sciatic nerves can be used as a marker to monitor Wallerian degeneration and regeneration of crushed sciatic nerves.

Safety of Local Anesthetics Administered Intrathecally in Diabetic Rats

Although retrospective studies show the risk of neurological complications after spinal anesthesia with local anesthetics is small in diabetic patients, there is still concern about the safety of different local anesthetics in diabetics undergoing neuroaxial anesthesia. We examined block duration and histology of spinal cord and roots with intrathecal local anesthetics in diabetic rats. Rats were made diabetic with streptozotocin injection. Blood glucose levels confirmed diabetes, and diabetic neuropathy was verified by tactile hypersensitivity. Diabetic and nondiabetic rats received four intrathecal injections at 3-4-day intervals of 0.75% bupivacaine, with/without 100 µg/mL epinephrine; and 2% lidocaine, with/without 100 µg/mL epinephrine, and duration of sensory (pinprick) and motor (toe-spreading reflex) response inhibition recorded. Four days after the last drug injection, histology of spinal cord and roots was performed. All streptozotocin rats became diabetic and had pronounced tactile allodynia. Intrathecal injection of local anesthetics showed longer duration of sensory and motor block in diabetic rats vs nondiabetics. Histology of caudal spinal cord showed no difference in neuropathology between diabetic and nondiabetic rats. Necrotic neurons were not seen in either group, and white-matter pathology involved less than 0.1% of fibers. Histology of the spinal roots also showed no difference in pathology between groups, and pathology involved less than 0.1% of fibers. Neuron somas in the dorsal root ganglia were normal. Duration of local anesthetic spinal block is longer in diabetic animals than in nondiabetics. However, there was no increased pathology of spinal cord, roots, or dorsal root ganglia.

Local Anesthetic Sciatic Nerve Block and Nerve Fiber Damage in Diabetic Rats

A concern for anesthesiologists is whether local anesthetics are more toxic to peripheral nerves in diabetic patients. A previous study in streptozotocin-induced diabetic rats showed that larger doses of lidocaine produce moderate nerve injury after nerve block in normal rats and worse injury in diabetic rats. However, it is not clear whether a smaller local anesthetic dose that produces negligible nerve fiber damage in normal rats will produce significant nerve damage in diabetic rats and if adding adjuvant drugs modulates this effect. Rats were intravenously injected with 50 mg/kg streptozotocin to induce diabetes (blood glucose levels 9250 mg/dL) and diabetic neuropathy. After waiting 35 days, an injection (0.1 mL) of 1% lidocaine alone, or with 5 kg/mL epinephrine or 7.5 kg/mL clonidine added, or 0.5% ropivacaine alone was performed at the left sciatic notch in both diabetic and nondiabetic rats. The duration of sensory (pin prick) and motor (toe spreading reflex) nerve block in the hind paws was determined.For histologic controls, all rats also received saline vehicle injection into the right sciatic notch. Another group of uninjected rats was used as naive controls. Left and right nerves were removed 2 days after injection and fixed in situ with a 4% glutaraldehyde solution. Myelinated axon profiles suggestive of neuropathy (myelin figures, pale and swollen,or dark-staining axoplasm) were counted and expressed as a percentage of the total number of fibers in each rat sciatic nerve. All streptozotocin-injected rats became diabetic and had pronounced tactile allodynia. All rats had sensory and motor nerve blocks lasting for at least 50 mins after injection of local anesthetic. The duration of sensory and motor nerve block was longer in diabetic rats than in nondiabetic rats for all drug groups tested. None of the sciatic nerves examined showed greater than 3% nerve fiber degeneration. Although lidocaine in diabetic rats did not produce nerve fiber damage,diabetic rats receiving lidocaine/clonidine or ropivacaine had more abnormal myelinated axon profiles than did nondiabetic rats receiving the same drug. The duration of sciatic nerve block with local anesthetics is longer in diabetic compared with nondiabetic rats. A small, but statistically significant, increase in nerve damage occurred in diabetic rats after nerve block with ropivacaine alone or when duration of lidocaine block was extended with clonidine. These findings may have implications for dosing of local anesthetics in diabetic patients undergoing regional analgesia with nerve blocks.

Capsaicin Protects Mouse Neuromuscular Junctions From The Paralytic Effects Of Botulinum Neurotoxin A

Botulinum neurotoxins (BoNTs) are the most toxic naturally occurring proteins. Botulinum serotype A (BoNT/A) selectively cleaves SNAP-25 and inhibits acetylcholine release from motor nerve endings resulting in life threatening paralysis of skeletal muscles. Here we report that capsaicin (8-methyl-N-vanillyl-6-nonenamide), an irritant active principle of chili peppers, protects against the paralytic effects of BoNT/A in mouse. In Triangularis sterini nerve muscle preparations, capsaicin pretreatment in vitro, significantly reduced fluorescent labeled BoNT/A uptake mediated by depolarization with 40 mM KCl or neural stimulations. In vivo injection of capsaicin (3 μl of a 1 mM stock solution) either coinjected or injected 4 or 8 hours before BoNT/A injection protected the mice from the inhibitory effects of BoNT/A measured by toe spread reflex. In controls the toe spreading reflex was inhibited within 24 hrs of poisoning with BoNT/A. Also wortmannin, a PIP5K inhibitor, injected prior to BoNT/A exposure, protected the mice from the paralytic effect of BoNT/A. In vitro muscle tension measurements demonstrate that capsaicin pretreatment partially protected the functions of the mouse Extensor digitorum longus nerve muscle preparations. Also pretreatment of cultured cholinergic neuroblastoma (N2a) cells with 10 μM capsaicin for 10 minutes prior to exposure of 10 pM BoNT/A significantly preserved labeling of synaptic vesicle pools by FM1-43. Our data collectively demonstrate that capsaicin offers protection from the paralyzing effect of BoNT/A by significantly reducing the uptake of the toxin in mouse neuromuscular junction probably by interfering with endocytosis of the toxin.Supported by HDTRA1-07-C-0031 CBT BAA.

Functional Recovery Enhancement Following Injury to Rodent Peroneal Nerve by Lion's Mane Mushroom, Hericium erinaceus (Bull.: Fr.) Pers. (Aphyllophoromycetideae)

Peripheral nerve injury represents a huge burden to society. Following peripheral nerve injury, improved behavioral outcome may be the most important evidence of functionality of axonal regeneration after any repair strategy. Nerve-crush injury is a well-established axonotmetic model in experimental regeneration studies to investigate the impact of various pharmacological treatments. Hericium erinaceus is a temperate mushroom but is now being cultivated in tropical Malaysia. In this study, we investigated the activity of aqueous extract of H. erinaceus fresh fruitbodies in promoting functional recovery following an axonotmetic peroneal nerve injury in adult female Sprague-Dawley rats with a long-term view toward the possible use of this mushroom in the treatment of nerve injury. Functional recovery was assessed in the behavioral experiment by walking-track analysis and toe-spreading reflex. The peroneal functional index (PFI) was determined before surgery and after surgery, as the rats showed signs of recovery. Analysis of the PFI indicated that the return of hind-limb function occurred by day 10 to 14 and by day 14 to 17 in the treated and control (nontreated) groups, respectively. Normal toe-spreading in the crushed limb was achieved by day 7 to 10 and day 12 to 17 in the treated and control group, respectively. These results suggest that daily administration of aqueous extract of H. erinaceus fresh firuitbodies has a beneficial effect on the recovery of injured rat peroneal nerve in the early stages of regeneration. The PFI and toe-spreading reflex improved faster in the treated group than in the nontreated group.

Relevance of Muscle Weight as a Functional Parameter for the Regeneration of the Peroneal Nerve in Rabbits

To evaluate the relevance of muscle weight as a functional parameter of motor regeneration, the nerve-dependent muscle weights were measured and compared with the toe-spreading reflex and the muscle action potential in different designs of nerve conditioning of the peroneal nerve in New Zealand White rabbits. From 100 peroneal nerves in 50 rabbits, 60 nerves received a conditioning crush lesion and 4 weeks later a test lesion--a transection with suturing at different sites. An additional 10 peroneal nerves were solely transected and sutured, 10 peroneal nerves were sham-operated, and 20 peroneal nerves remained unoperated for control. Eleven weeks after the final lesion, when the toe-spreading reflex had returned, the peroneal muscle weight showed a non-significant relation with the toe-spreading reflex (area 0.650, P = 0.149), whereas the relation of the amplitude (area 0.964, P < 0.001) and the conduction velocity (area 0.721, P = 0.033, left side) with the toe-spreading reflex were significant. Despite the lower sensitivity and specificity of the muscle weight, the possibility to measure the weight of all three peroneal nerve-dependent muscles separately represents a more detailed method of evaluation than both the toe-spreading reflex testing and the muscle action potential recording and justifies its use as a functional parameter in the early phase of peroneal motor regeneration in the rabbit.

Analysis of axonal regeneration in the central and peripheral nervous systems of the NG2-deficient mouse.

BackgroundThe chondroitin sulphate proteoglycan NG2 blocks neurite outgrowth in vitro and has been proposed as a major inhibitor of axonal regeneration in the CNS. Although a substantial body of evidence underpins this hypothesis, it is challenged by recent findings including strong expression of NG2 in regenerating peripheral nerve.ResultsWe studied axonal regeneration in the PNS and CNS of genetically engineered mice that do not express NG2, and in sex and age matched wild-type controls. In the CNS, we used anterograde tracing with BDA to study corticospinal tract (CST) axons after spinal cord injury and transganglionic labelling with CT-HRP to trace ascending sensory dorsal column (DC) axons after DC lesions and a conditioning lesion of the sciatic nerve. Injury to these fibre tracts resulted in no difference between knockout and wild-type mice in the ability of CST axons or DC axons to enter or cross the lesion site. Similarly, after dorsal root injury (with conditioning lesion), most regenerating dorsal root axons failed to grow across the dorsal root entry zone in both transgenic and wild-type mice.Following sciatic nerve injuries, functional recovery was assessed by analysis of the toe-spreading reflex and cutaneous sensitivity to Von Frey hairs. Anatomical correlates of regeneration were assessed by: retrograde labelling of regenerating dorsal root ganglion (DRG) cells with DiAsp; immunostaining with PGP 9.5 to visualise sensory reinnervation of plantar hindpaws; electron microscopic analysis of regenerating axons in tibial and digital nerves; and by silver-cholinesterase histochemical study of motor end plate reinnervation. We also examined functional and anatomical correlates of regeneration after injury of the facial nerve by assessing the time taken for whisker movements and corneal reflexes to recover and by retrograde labelling of regenerated axons with Fluorogold and DiAsp. None of the anatomical or functional analyses revealed significant differences between wild-type and knockout mice.ConclusionThese findings show that NG2 is unlikely to be a major inhibitor of axonal regeneration after injury to the CNS, and, further, that NG2 is unlikely to be necessary for regeneration or functional recovery following peripheral nerve injury.

Functional, electrophysiologic, and morphometric evaluation of nerve regeneration from coaptation on regenerated nerve fibers: experimental study in rabbits.

The importance of a sufficient number of nerve fibers at a proximal coaptation site is indisputable for the successful repair of nerves; however, the quality of nerve fibers required at this site has yet to be defined. The present study deals with the question of whether it is necessary to trim nerves back to unaffected neuronal tissue or whether the coaptation on recently regenerated nerve fibers, commonly believed to produce a poor quality of repair can, in fact, produce adequate nerve regeneration. Twenty New Zealand White rabbits received a standardized crush lesion on the peroneal nerves of both hind legs. Four weeks later, the nerves of the left hind legs (n = 20) were transected 10 mm distal to the previous crush lesion and coapted to the freshly regenerated nerve fibers. For comparison, on 10 right hind legs, the nerves were transected at the site of previous crushing (Group A, superimposition) or 10 mm proximal to the site of crushing on unscathed nerve fibers (Group B). Eleven weeks later, the quality of nerve regeneration was assessed by the toe-spreading reflex, electrophysiologic data, muscle weight, and histomorphologic evaluation. In the animals of Group A, the quality of nerve regeneration following coaptation on the regrown axons did not differ in any of the examined parameters from the quality of nerve fibers outgrown from the site of the superimposed lesion. Both lesions led to a completely functional reinnervation. Also in Group B, nerve action potential recording and histologic data on both sides did not reveal a significant difference between the number and maturation of nerve fibers equidistant from the suture site, shortly before muscle entrance. With this coaptation model, it could be demonstrated in the peroneal nerve of rabbits, that coaptation to recently regenerated nerve fibers leads to a significant functional regeneration.

Magnetic stimulation accelerating rehabilitation of peripheral nerve injury.

The effect of magnetic stimulation (MS) on sciatic nerve injury was observed. After sciatic nerve was crushed in 40 Sprague Dawley (SD) rats, one randomly selected group (group D) was subjected, from the 4th day post-operatively to 3 min of continuous 70% of maximum output of MS daily for 8 weeks. The other group (group E) served as a control group. The nerve regeneration and motor function recovery were evaluated by walking track analysis (sciatic function index, SFI; toe spreading reflex, TSR), electrophysiological, histological and acetylcholineesterase histochemistry. The SFI in the group D was greater than in the group E with the difference being statistically significant (P < 0.01). TSR reached its peak on the 4th day in the group D and on the 10th day in the group E respectively. The amplitude and velocity of MCAP and NCAP in the group D was greater than in the group E with the difference being statistically significant (P < 0.01), while the latency and duration of MCAP and NCAP in the group D were less than in the group E with the difference being also statistically significant (P < 0.01). Histological examination showed the mean axon count above the lesion for thick myelinated fibers (> 6.5 microns) in the group D was greater than in the control group with the difference being statistically significant (P < 0.01), while the mean axon count below the lesion for thick myelinated fibers was less than that in the group E with the difference being statistically significant (P < 0.01). The mean axon count above the lesion for thin myelinated fibers (2-6.5 microns) in the group D was greater than that in the group E with the difference being statistically significant (P < 0.05), while the mean axon count below the lesion for thin myelinated in the group D was greater than that in the group E with the difference being statistically significant (P < 0.01). Acetylcholine esterase examination showed that the MS could significantly increase the number of the motor neurons. There was no significant difference in the number of the motor neurons between the treatment side and the normal side (P > 0.05). It can be concluded that MS can enhance functional recovery and has a considerable effect in the treatment of the peripheral nerve injury.

The toe-spreading reflex of the rabbit revisited--functional evaluation of complete peroneal nerve lesions.

Although a variety of electrophysiological and morphological tests are available for studying nerve regeneration in animals, these endpoints do not necessarily correlate with the return of muscle function. Recent efforts have focused on the assessment of function as the endpoint of nerve regeneration. One of the best known of these tests is the sciatic function index in rats. For rabbits, the toe-spreading reflex has been suggested as a valuable index of peroneal function. We examined the reliability and sensitivity of the toe-spreading reflex in a study of nerve regeneration of the peroneal nerve in 10 New Zealand White rabbits. Eleven weeks after the transection and immediate suturing of the peroneal nerve in both hind legs (at two slightly different sites), a toe-spreading reflex could always be elicited on that side where the level of the severed nerve was closer to the dependent muscles. Also on this hind leg the muscle weight of the peroneal target muscles was significantly higher (P = 0.031) than on the contralateral side, which corresponds well to the results of the toe-spreading reflex. The toe-spreading reflex is an excellent and sensitive indicator of the onset of motor recovery in the peroneal nerve-dependent muscles of rabbits. Even small differences in the localization of lesions in both hind legs can be differentiated with this test.

Mice Lacking tPA, uPA, or Plasminogen Genes Showed Delayed Functional Recovery after Sciatic Nerve Crush

Axonal outgrowth during peripheral nerve regeneration relies on the ability of growth cones to traverse through an environment that has been altered structurally and along a basal lamina sheath to reinnervate synaptic targets. To promote migration, growth cones secrete proteases that are thought to dissolve cell-cell and cell-matrix adhesions. These proteases include the plasminogen activators (PAs), tissue PA (tPA) and urokinase PA (uPA), and their substrate, plasminogen. PA expression and secretion are upregulated in regenerating mammalian sensory neurons in culture. After sciatic nerve crush in mice, there was an induction of PA mRNAs in the sensory neurons contributing to the crushed nerve and an upregulation of PA-dependent activity in crushed nerve compared with sham counterparts during nerve regeneration. To further assess the role of the PA system during peripheral nerve regeneration, PA-dependent activity as well as recovery of sensory and motor function in the injured hindlimb were assessed in wild-type, tPA, uPA, and plasminogen knock-out mice. Protease activity visualized by gel zymography showed that after nerve crush, the upregulation of PA activity in the tPA and uPA knock-out mice was delayed compared with wild-type mice. Recovery of sensory function was assessed by toe pinch, footpad prick, and the toe-spreading reflex. All knock-out mice demonstrated a significant delay in hindlimb response to these sensory stimuli compared with wild-type mice. For each modality tested, the uPA knock-out mice were the most dramatically affected, showing the longest delay to initiate a response. These studies clearly showed that PAs were necessary for timely functional recovery by regenerating peripheral nerves.

Effects of methylprednisolone and MK-801 on functional recovery after experimental chronic spinal cord injury.

An experimental study was conducted to evaluate the effects of methylprednisolone and MK-801 after the compressive injury of spinal cord in rats. To investigate the effect of methylprednisolone and non-competitive NMDA antagonist MK-801 in long-term functional outcome after spinal cord injury (SCI). A randomized group A of Sprague-Dawley rats were treated with MK-801 (1.0 mg/kg, n=10; Group A) after a compression injury. A group of methylprednisolone (MP)-treated (30 mg/kg, n=10; Group B) and non-treated animals (n=9; Group C) were included for comparison. The functional motor outcome such as inclined plane (IP), toe spreading reflex (TSR), and modified Tarlov scale (TS) were measured in each animal at regular time points up to 8 weeks post-treatment. Histologically the injury site was scored in four groups and immunohistochemically Wallerian Degeneration (WD), astrocytosis and expression of beta-amyloid protein was identified. In examining the IP data, no significant difference was recognized between the group means (P-value>0.5). For the TSR, there were no differences in the group responses. For the TS, the differences were not statistically significant. Only group B showed significance in cavitation scores compared to group A (P>0.0094), WD was significantly different than group C (P>0.03), astrocytosis was significantly higher than group A (P>0.001) and modest presence of beta-amyloid protein. Our data indicate that one time bolus administration of MK-801 lacks any significant effect on axonal function in chronically injured rats. Daily bolus administration of MP at 30 mg/kg also did not ensure a better functional outcome. Immunohistochemically we have been able to show significant differences in WD, astrocytosis and small insignificant changes in beta-amyloid protein.

Enhancement of Mouse Sciatic Nerve Regeneration by the Long Chain Fatty Alcohol,N-Hexacosanol

The purpose of the present study was to determine the effects ofn-hexacosanol (hexa) on nerve regeneration. Hexa, a long chain fatty alcohol has been shown to possess neurotrophic properties on cultured neurons and to attenuate the degeneration of cholinergic neurons after injury. The effects of daily intraperitoneal injections of hexa (1 mg/kg) on regeneration of nerve fibers were studied in mice following a sciatic nerve crush. Measurement of axonal regeneration using the pinch test 7 days postlesion showed a 40% increase of the regeneration rate of sensory fibers in hexa-treated mice compared to controls (1.67±0.15 mm/day and 1.09±0.03 mm/day, respectively). The recovery of neuromuscular function was significantly improved, as shown by quantitative electromyography and sensorimotor tests. Clinical signs of recovery evaluated with toe spreading reflex appeared earlier in hexa group than in control animals. Electrophysiological recordings were performed each 3 days during 34 days following nerve injury. Higher values of the compound muscle action potential (CMAP) were obtained in hexa-treated animals that correspond to an improved regeneration. Moreover, hexa induced a significantly faster regeneration rate (hexa: 2.87±0.15 mV/day; control: 2.00±0.06 mV/day), as measured by the slope of CMAP increase (44% enhancement). A morphometric analysis performed 7 days following crush showed an increased number of regenerating fibers, as well as increased diameter and thickness of the myelin in hexa-treated mice. Thus, hexa increased the regeneration of both sensory and motor axons in lesioned nerve, leading to an improved functional recovery.

Long‐term effects of deprivation of cell support in the distal stump on peripheral nerve regeneration

The distal stump of an injured peripheral nerve supports regenerating axons by offering a favourable growth substratum and several cell-produced growth factors. Deprivation of cellular factors alone has been shown not to prevent fairly rapid axonal elongation after nerve injury if the growth substratum was preserved. The present study examined possible long-term untoward effects of cell support deprivation during an early phase of nerve regeneration. Rat sciatic nerve was crushed and a 25 mm long distal nerve segment was made acellular by freezing-thawing, while the integrity of the growth substratum for the regenerating axons was preserved. Toe-spreading reflex and skin sensitivity to pinch in the foot were monitored to follow recovery of motor and sensory function, respectively. The number of myelinated axons was determined in the sciatic nerve proximally to the lesion site, and distally in the predominantly sensory sural nerve as well as in the mixed motor nerve to the soleus muscle. Except for a short delay in the onset of recovery, explainable by the reduced elongation rate of axons growing through the acellular nerve segment, we found no deleterious effect of cell support deprivation on sensory or motor function recovery after nerve crush. Most of regenerating sensory neurons did not critically depend on the distal stump cell support. However, a 15% and 25% loss of myelinated axons both proximally to the lesion and distally in the sensory sural nerve, respectively, indicated that a corresponding minor loss of injured sensory neurons occurred when they were deprived of such cell support even if provided with a favourable growth substratum for successful regeneration.(ABSTRACT TRUNCATED AT 250 WORDS)

The lack of an effect of applied d.c. electric fields on peripheral nerve regeneration in the guinea pig

This study was undertaken to provide evidence of enhanced regeneration of mammalian peripheral nerves in response to applied d.c. electric fields. Peroneal nerves of adult guinea-pigs were crushed or transected and anastomosed. Constant current d.c. stimulators (20 μA) were implanted in the flank with platinum/iridium electrodes routed to the ankles. Animals with crush lesions were tested for toe spreading ability from the 14th to the 23rd day following the lesion. Animals with transection lesions were allowed to recover for 40 days and isometric force measurements of toe abduction and foot flexion were made. Both myelinated and unmyelinated fiber densities were determined. There proved to be no difference between legs treated with an anode, a cathode, or a sham electrode as evaluated by: the time to return of the toe spreading reflex, the isometric force of either twitches or tonic contractions, the latency between stimulation and contraction, or the number or density of either myelinated or unmyelinated fibers. These negative results are at variance with other studies that have reported beneficial effects of d.c. electric fields on peripheral nerve regeneration. The stimulation and analysis techniques used in this study were well within the variety of protocols that have yielded reports of highly significant positive effects with smaller numbers of animals than used in this study. The conclusion is that either there is a subtle but highly specific and critical difference between the present protocol and others, or the other studies need to be reevaluated. In either case, it seems that the ability of applied d.c. fields to enhance peripheral nerve regeneration in vivo remains open to question.

A comparison of cryoprobe and crush lesions in the rat sciatic nerve

This study examines the behavioral, sensory, motor and structural recovery during the first 2 months following a freeze (cryoprobe) lesion compared to a nerve crush (forceps). There is a complete loss of sensory and motor function following either type of lesion during the first 2 weeks of recovery. The toe spreading reflex and the sciatic functional index of locomotion behavior returned to normal without significant group differences. Latency times for the pain withdrawal reflex were slightly shorter in the cryoprobe group, but both groups returned to baseline during the second month. An improved regenerative pattern was suggested for the motor recovery in the cryoprobe group as expressed by the amplitude of the digital twitch tension curves. However, the respective curve areas were not different. Morphometric analysis indicated a significant reduction in the distal nerve cross-sectional area, an increase in the mean myelinated fiber density and an increase in the estimated total number of myelinated nerve fibers in both experimental groups. Mean fiber diameter and myelin sheath thickness had not fully returned to normal in either experimental group. Both the fiber size and myelin sheath thickness were significantly reduced in the cryoprobe group. In conclusion, the two lesion types have remarkably similar patterns of recovery. Functional data suggest that motor recovery precedes sensory recovery following a cryoprobe lesion.

Chronic access to endoneurial space using an arterial autograft

A technique for direct chronic infusion of compounds onto peripheral axons has been investigated in the rat sciatic nerve. A 2 cm segment of the femoral artery was removed and one end inserted into the endoneurial space of the contralateral peroneal nerve fascicle of the same animal. The other end of the artery was connected to a catheter system to allow infusion into the endoneurium, thus bypassing the barrier that the perineurium presents to hydrophilic compounds. The patency of this arterial access system was evaluated by the ability of 20 μl of 2% lidocaine to inhibit the toe-spreading reflex. The results of the study were that the infusion system remained operational for 3–7 days after transplantation. There were histologic changes in the nerve but there were no functional deficits due to the surgery. Although a long-term endoneurial infusion was not achieved, the limited access time to the axons might be long enough for applications such as the delivery of nerve growth factors to injured nerve.