Interfacing peripheral nerve with macro-sieve electrodes following spinal cord injury

Macro-sieve electrodes were implanted in the sciatic nerve of five adult male Lewis rats following spinal cord injury to assess the ability of the macro-sieve electrode to interface regenerated peripheral nerve fibers post-spinal cord injury. Each spinal cord injury was performed via right lateral hemisection of the cord at the T_(9–10) site. Five months post-implantation, the ability of the macro-sieve electrode to interface the regenerated nerve was assessed by stimulating through the macro-sieve electrode and recording both electromyography signals and evoked muscle force from distal musculature. Electromyography measurements were recorded from the tibialis anterior and gastrocnemius muscles, while evoked muscle force measurements were recorded from the tibialis anterior, extensor digitorum longus, and gastrocnemius muscles. The macro-sieve electrode and regenerated sciatic nerve were then explanted for histological evaluation. Successful sciatic nerve regeneration across the macro-sieve electrode interface following spinal cord injury was seen in all five animals. Recorded electromyography signals and muscle force recordings obtained through macro-sieve electrode stimulation confirm the ability of the macro-sieve electrode to successfully recruit distal musculature in this injury model. Taken together, these results demonstrate the macro-sieve electrode as a viable interface for peripheral nerve stimulation in the context of spinal cord injury.     

Spatiotemporal expression of inducible nitric oxide synthase and cyclooxygenase 2 in the spinal cord during early stage sciatic nerve crush injury

BACKGROUND: Previous studies have shown that inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX-2) participate in inflammatory immune responses and neuropathic pain following peripheral nerve injury. However, few reports have addressed time-dependent expression of iNOS and COX-2 following peripheral nerve injury. OBJECTIVE: To investigate spatiotemporal expression of iNOS and COX-2 during early stage sciatic nerve crush injury.DESIGN, TIME AND SETTING: The randomized, controlled, animal experiment was performed at the Laboratory of Applied Anatomy, Department of Human Anatomy and Neurobiology, Central South University, China from September 2006 to September 2007.MATERIALS: Mouse anti-rat iNOS monoclonal antibody and goat anti-rat COX-2 monoclonal antibody (Transduction Laboratory, USA), as well as biotinylated rabbit anti-mouse lgG and biotinylated rabbit anti-goat IgG (Santa Cruz Biotechnology, USA) were used in the present study.METHODS: A total of 48 healthy, adult, Sprague Dawley rats were randomly assigned to three groups. In the model group (n = 32), crush injury to the right sciatic nerve was established using an artery clamp. The model group was further assigned to four subgroups according to survival time (6,12, 24, and 72 hours), respectively (n = 8). Sham surgery (n = 8) and normal control (n = 8) groups were also established.MAIN OUTCOME MEASURES: iNOS and COX-2 expression was detected in the L4-6 spinal cord with immunohistochemistry. Gray values of iNOS- and COX-2-postive cells in the anterior horn and posterior horn of spinal cord, as well as quantification of iNOS- and COX-2-positive cells in the anterior horn of spinal cord, were measured.RESULTS: iNOS and COX-2 expression gradually increased in the anterior horn and posterior horn of the spinal cord on the damaged side over time from 6 hours following sciatic nerve injury (P＜0.05) and peaked at 72 hours. Simultaneously, the number of iNOS- and COX-2-positive cells similarly increased in the anterior horn of spinal cord on the damaged side (P＜ 0.05).CONCLUSION: iNOS and COX-2 expression increased in the spinal cord during early stage sciatic nerve crush, which suggested that iNOS and COX-2 participate in occurrence and development of inflammatory immune responses following peripheral nerve injury.     

Nerve grafting from spinal cord to spinal nerve: a microsurgical technique in cats

A ventral surgical approach is described for the grafting of autologous saphenous nerves between the spinal cord and the avulsed C7 ventral root in the cat. To overcome serious blood loss from the epidural venous plexus, the cats were hyperventilated (end tidal

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to about 23 mmHg) and controlled hypotension was induced (mean arterial pressure to about 60 mmHg). After selective avulsion of the ventral rootlets C7 the saphenous grafts were implanted into the spinal cord and coaptated to the avulsed spinal nerve. The combination of advanced anesthetic methods and microsurgical techniques appeared to be mandatory to achieve a low surgical mortality. Regenerated axons were retrogradely traced using retrograde horseradish peroxidase (HRP), and their functional recovery was evaluated by means of electrophysiological methods.     

Craniocerebral injury promotes the repair of peripheral nerve injury

The increase in neurotrophic factors after craniocerebral injury has been shown to promote fracture healing. Moreover, neurotrophic factors play a key role in the regeneration and repair of peripheral nerve. However, whether craniocerebral injury alters the repair of peripheral nerve injuries remains poorly understood. Rat injury models were established by transecting the left sciatic nerve and using a free-fall device to induce craniocerebral injury. Compared with sciatic nerve injury alone after 6–12 weeks, rats with combined sciatic and craniocerebral injuries showed decreased sciatic functional index, increased recovery of gastrocnemius muscle wet weight, recovery of sciatic nerve ganglia and corresponding spinal cord segment neuron morphologies, and increased numbers of horseradish peroxidase-labeled cells. These results indicate that craniocerebral injury promotes the repair of peripheral nerve injury.     

Substance pin spinal cord dorsal horn decreases following peripheral nerve injury

Substance P was located in the spinal cord of rats by immunocytochemistry.Section and ligation of the sciatic nerve produced a depleted area low in substance P in the medial two-thirds of laminae 1 and 2 of segments L4 and 5.The time of depletion began about 5 days after the nerve had been cut and substance P reached a steady minimum by about 9 days and remained depleted for the entire period examined, 31 days.Crush lesions of the sciatic nerve failed to produce the marked and rapid changes of spinal cord substance P observed after section and ligation.     

Carbon filaments direct the growth of postlesional plastic axons after spinal cord injury

The effect of implantation of carbon filaments and fetal tissues on the axonal regeneration following contusion injury in a rat model was investigated by in situ immunofluorescence. Female Sprague-Dawley rats were subjected to severe contusion injury to the spinal cord at T9-T10. All animals were divided into 5 groups (N = 5/group): normal controls. surgical controls, with carbon filament implants, with fetal tissue implants and with implants consisting of fetal tissue cocultured with carbon filaments. After a 10-week survival period, the astroglial response was assessed by immunoreactive glial fibrillary acidic protein and the neuro-axonal profile by immunoreactive phosphorylated and nonphosphorylated neurofilament proteins. The contusion injury resulted in: (a) dramatically increased immunoreactivity of glial fibrillary acidic protein indicating injury-associated reactive astrogliosis, (b) increase in immunoreactive phosphorylated neurofilament protein indicating upregulated phosphorylation of neurofilament protein, (c) with no change in the highly differentiated nonphosphorylated neurofilament protein which normally occur in the nonregenerating mature neurons. Implantation of fetal tissues alone following contusion injury did not show any appreciable change with regard to the immunoreactivities for the glial and neuronal markers studied, compared to the injury controls. However, the implantation of carbon filaments alone or together with fetal tissues directed the growth of glial fibrillary acidic protein-positive astroglia and phosphoneurofilament-positive neurites along the carbon fibers, with no effect on nonphosphoneurofilament protein. In conclusion, implantation of carbon filaments appears to be critical for facilitating the attachment of astroglia forming a substrate and scaffolding that can further support and direct the growth of postlesional plastic axons across the lesion. In addition, carbon filament prostheses in combination with fetal tissue implants provides an improved combinational approach to promote regrowth of injured neurons following injury.     

Axonal regeneration from the spinal cord to peripheral nerve induced by end-to-side neurorrhaphy Evidence from acetylcholinesterase staining and Fluorogold retrograde tracing

BACKGROUND： In recent years, surgeons have advocated root or trunk repair of avulsed nerve roots for overall recovery. However, donor nerves pose a major problem, because they do not contain adequate numbers of axons. Moreover, the procedures lead to nerve deficits in the donor nerve following transplantation. OBJECTIVE： To observe whether axonal regeneration occurs by end-to-side neurorrhaphy in the peripheral nerve and spinal cord. DESIGN, TIME AND SETTING： A neuroanatomical, randomized, controlled, animal study was performed at Functional Anatomy Lab in Nagoya University School of Medicine from May 2002 to July 2003. MATERIALS： Fluorogold was purchased from Fluorochrome, LLC, USA. BX50 light microscope and fluorescent microscope were purchased from Olympus, Japan. METHODS： A total of 21 rats were randomly divided into three groups, and the posterior avulsion injury model （C6-8） of the brachial plexus was performed. In the ventral root graft group, the avulsed C7 ventral roots were reanastomosed to the small anterior lateral aspect window of the spinal cord via nerve grafts. In the dorsal root graft group, the C7 dorsal roots were reanastomosed at the small pia mater window of the posterior lateral aspect of the spinal cord via nerve grafts. In the control group, the avulsed nerve roots were not repaired. MAIN OUTCOME MEASURES： The nerve grafts were collected from the ventral and dorsal root graft groups, and the C7 proximal nerve end was collected from the control group. Acetylcholinesterase staining was performed on the tissue. Fluorogold retrograde tracing technique was applied to determine the origin of the regenerating axons. RESULTS： Results showed that acetylcholine-positive axons existed in nerve grafts of the ventral and dorsal root graft groups. However, axons were not found in the avulsed nerve roots of the control group. Fluorogold retrograde tracing confirmed the presence of fluorogold-containing neurons in the ventral and dorsal horn of the ventral and dorsal root graft groups. Fluorogold-positive neurons were not observed in the control group. CONCLUSION： End-to-side neurorrhaphy induced axonal regeneration from the spinal cord to the peripheral nervous system.     

Changes in the somatotopic organization of the cat lumbar spinal cord following peripheral nerve transection and regeneration

Glass microelectrodes were used to record the activity of neurones in the left dorsal horn of the L6 segment of the spinal cord of normal cats and cats in which the left sciatic and saphenous nerves had been cut 1 or 9 months previously. In the normal animals the receptive fields of L6 dorsal horn neurones excited by tactile stimulation of the leg were somatotopically organized, with neurones in the medial and central dorsal horn having receptive fields on the distal parts of the leg, particularly the toes, and neurones in the lateral dorsal horn having receptive fields on the proximal parts of the leg, buttock and lower back. This somatotopy has been shown before. One month after nerve section no cells responded to tactile stimulation of the distal leg and cells in the medial and central parts of the dorsal horn now had receptive fields on the proximal leg, buttock and back. There did not appear to be any somatotopic organization of these new receptive fields. Lateral dorsal horn neurones had normal receptive fields. Nine months after nerve section neurones in the medial and central parts of the lumbar dorsal horn had receptive fields on the distal leg but they showed several abnormal features and there was no evidence of a return of the somatotopic organization seen in normal animals. Lateral dorsal horn cells still had normal receptive fields.     

Leukotriene synthases and the receptors induced by peripheral nerve injury in the spinal cord contribute to the generation of neuropathic pain

Leukotrienes (LTs) belong to a large family of lipid mediators, termed eicosanoids, which are derived from arachidonic acids and released from the cell membrane by phospholipases. LTs are involved in the pathogenesis of inflammatory diseases, such as asthma, rheumatoid arthritis, and peripheral inflammatory pain. In the present study, we examined whether LTs were implicated in pathomechanism of neuropathic pain following peripheral nerve injury. Using the spared nerve injury (SNI) model in rats, we investigated the expression of LT synthases (5‐lipoxygenase; 5‐LO, Five lipoxygenase activating protein; FLAP, LTA4 hydrolase; LTA4h and LTC4 synthase; LTC4s) and receptors (BLT1, 2 and CysLT1, 2) mRNAs in the rat spinal cord. Semi‐quantitative RT‐PCR revealed that 5‐LO, FLAP, LTC4s, BLT1, and CysLT1 mRNAs increased following SNI, but not CysLT2 mRNAs. Using double labeling analysis of in situ hybridization with immunohistochemistry, we observed that 5‐LO, FLAP, and CysLT1 mRNAs were expressed in spinal microglia. LTA4h and LTC4s mRNAs were expressed in both spinal neurons and microglia. BLT1 mRNA was expressed in spinal neurons. The p38 mitogen‐activated protein kinase inhibitor, but not MEK inhibitor, reduced the increase in 5‐LO in spinal microglia. Continuous intrathecal administration of the 5‐LO inhibitor or BLT1 and CysLT1 receptor antagonists suppressed mechanical allodynia induced by SNI. Our findings suggest that the increase of LT synthesis in spinal microglia produced via p38 MAPK plays a role in the generation of neuropathic pain. © 2009 Wiley‐Liss, Inc.     

Apelin inhibits motor neuron apoptosis in the anterior horn following acute spinal cord and sciatic nerve injuries

Rat models of acute spinal cord injury and sciatic nerve injury were established.Apelin expression in spinal cord tissue was determined.In normal rat spinal cords,apelin expression was visible;however,2 hours post spinal cord injury,apelin expression peaked.Apelin expression increased 1 day post ligation of the sciatic nerve compared with normal rat spinal cords,and peaked at 3 days.Apelin expression was greater in the posterior horn compared with the anterior horn at each time point when compared with the normal group.The onset of neuronal apoptosis was significantly delayed following injection of apelin protein at the stump of the sciatic nerve,and the number of apoptotic cells after injury was reduced when compared with normal spinal cords.Our results indicate that apelin is expressed in the normal spinal cord and central nervous system after peripheral nerve injury.Apelin protein can reduce motor neuron apoptosis in the spinal cord anterior horn and delay the onset of apoptosis.     

The effect of peripheral nerve injury on dorsal root potentials and on transmission of afferent signals into the spinal cord

The sciatic nerve adults rats was either cut and ligated or was crushed on one side. The response of the spinal cord to stimulation of the proximal part of the injured nerve was examined at various times after the lesion and compared to the effects of stimulating the intact nerve on the other side. During the first 10 days after nerve section the following measures were not affected: (i) the size of the input volley (compound action potential, CAP, measured on a dorsal root that carried sciatic nerve afferents (L5); (ii) the volley running in the dorsal columns; (iii) the dorsal root potential (DRP) evoked on neighbouring dorsal roots which do not contain sciatic afferents (L2 and L3); (iv) the post-synaptic volleys ascending in the spinal cord. However, by the fourth day after nerve section, there was a decrease of the DRP evoked on the ipsilateral L5 dorsal root by stimulation of the cut nerve. By 10 days this DRP had decreased by 50%. There was also a decrease in the DRP on the L5 root evoked by stimulation of the contralateral intact nerve. Crush lesions of the sciatic nerve did not produce DRP charge. Beginning 10–20 days after nerve cut, there was a decrease in the amplitude of the afferent CAP and of all the measures of central response to the afferent volley. We discuss the possibility that the loss of the DRP may be associated with a disinhibition which results in novel receptive fields which we observe in cord cells deafferented by the peripheral nerve section. The decrease of DRP and the appearance of novel receptive fields do not occur if the peripheral nerve is crushed rather than cut.     

Regenerating motor bridge axons refine connections and synapse on lumbar motoneurons to bypass chronic spinal cord injury

To restore motor control after spinal cord injury requires reconnecting the brain with spinal motor circuits below the lesion. A bridge around the injury is an important alternative to promoting axon regeneration through the injury. Previously, we reported a novel motor bridge in rats. The thirteenth thoracic nerve was detached from the muscle it innervates and the cut end implanted caudally into the lumbar gray matter where motor bridge axons regenerate. In this study, we first determined that regenerating bridge axons project to spinal motor circuits. Stable projections were present in ventral motor laminae of the cord, including putative synapses directly on motoneurons, 2 months after insertion in the intact cord. At this time, earlier-forming dorsal horn projections were mostly eliminated. Regenerating axons were effective in evoking leg motor activity as early as 2 weeks. We next determined that bridge axons could regenerate caudal to a chronic injury. We hemisected the spinal cord at L2 and inserted the bridge nerve 1 month later at L5 and found ventral laminae projections similar to those in intact animals, including onto motoneurons directly. Finally, we determined that the bridge circuit could be activated by neural pathways rostral to its origin. For spinally hemisected animals, we electrically stimulated the rostral spinal cord and recorded evoked potentials from the bridge and, in turn, motor responses in the sciatic nerve. Our findings suggests that bridge motoneurons could be used by descending motor pathways as premotor interneurons to transmit neural signals to bypass a chronic spinal injury.     

Edaravone promotes functional recovery after mechanical peripheral nerve injury

Edaravone has been shown to reduce ischemia/reperfusion-induced peripheral nerve injury. However, the therapeutic effect of edaravone on peripheral nerve injury caused by mechanical factors is unknown. In the present study, we established a peripheral nerve injury model by crushing the sciatic nerve using hemostatic forceps, and then administered edaravone 3 mg/kg intraperitoneally. The sciatic functional index and superoxide dismutase activity of the sciatic nerve were increased, and the malondialdehyde level was decreased in animals in the edaravone group compared with those in the model group. Bcl-2 expression was increased, but Bax expression was decreased in anterior horn cells of the L4–6 spinal cord segments. These results indicated that edaravone has a neuroprotective effect following peripheral nerve injury caused by mechanical factors through alleviating free radical damage to cells and inhibiting lipid peroxidation, as well as regulating apoptosis-related protein expression.     

修复周围神经损伤的神经移植材料

摘要：周围神经大段损伤一直是康复领域的重大难题,所以修复损伤的周围神经逐渐成为热点话题。在众多的修复方法中,移植修复周围神经损伤已经成为研究热点。自体移植物是临床上的“黄金标准”,人工组织工程材料移植物来源广泛,同种异体移植物可避免异体移植造成的免疫排斥反应。文章采用文献收集及总结的方法,探讨了自体移植物,人工组织工程材料移植物和同种异体移植物的研究进展。 关键词：周围神经;损伤;再生;移植;综述文献