Anatomical correlates of return of locomotor function after partial spinal cord lesions in cats

Summary We trained cats to walk on a moving treadmill belt, then subjected them to partial transverse sections of the thoracic spinal cord. Afterwards, we observed their ability to walk on the treadmill, over a period of several weeks, using gait analysis techniques to describe the resultant deficits. The extent of the lesions was verified histologically, and the identity of the spared descending axons from the brain stem was demonstrated by retrograde labeling with horseradish peroxidase.We found that significant sparing or recovery of hindlimb locomotor function is closely linked to sparing of axons in at least one ventrolateral quadrant of the cord. The essential elements probably belong to vestibulospinal and reticulospinal systems.Supported in part by grants by the NINCDS (NS-14546) and the Paraplegia Foundation of ArizonaMedical Investigator of the Veterans Administration     

Modulation of locomotor activity in complete spinal cord injury

The aim of this study was to evaluate the modulation of muscle activity during locomotor-like movements by different walking speeds in subjects with a motor complete spinal cord injury (SCI) compared to actively- and passively-walking control subjects without neurological deficit. Stepping movements on a treadmill were induced and assisted by a driven gait orthosis. Electromyographic (EMG) muscle activity of one leg (rectus and biceps femoris, tibialis anterior and gastrocnemius) was recorded and analyzed at three stepping velocities with similar body weight support in both subject groups. In SCI subjects, the EMG amplitude of biceps femoris, tibialis anterior and gastrocnemius was in general similar or weaker than in passively- and actively-stepping control subjects, but that of rectus femoris was larger. The degree of co-activation between tibialis anterior and gastrocnemius was higher in SCI than in control subjects. A significant velocity-dependent EMG modulation was present in all four-leg muscles in both subject groups. In SCI subjects, this EMG modulation was similar to that in actively stepping control subjects. It is concluded that in complete spastic SCI subjects, spinal neuronal circuits underlying locomotion can to a large extent adequately respond to a change in external drive to adapt the neuronal pattern to a new locomotion speed. The application of various speeds might enhance the effect of locomotor training in incomplete SCI subjects.     

Gonadotropin-releasing hormone treatment improves locomotor activity, urinary function and neurofilament protein expression after spinal cord injury in ovariectomized rats

It was reported that the hypothalamic decapeptide, gonadotropin-releasing hormone (GnRH) produces neurotrophic effects and that the spinal cord possesses GnRH receptors. The aim of the present study was to determine whether administration of GnRH improves locomotor activity, urinary function and neurofilament (NFs) protein expression after spinal cord injury (SCI) in ovariectomized rats. SCI was induced by balloon inflation model resulting in paraplegia. Locomotion was evaluated according to the Basso, Beattie, and Bresnahan Scale. Rats were subjected to bladder compression, twice daily until bladder reflex was established. NFs of 68, 160 and 200 kDa from spinal cords were analyzed by electrophoresis. GnRH (60 μg/kg) or physiologic NaCl solution was administered at 1 day after SCI and then daily for 15 days and the functional evaluation was realized for 5 weeks. Our results indicate that locomotor activity, restoration of urinary dysfunction and NFs expression of 160 and 200 kDa were improved in SCI animals given GnRH compared to those without treatment. These findings suggest that GnRH acts as a neurotrophic factor and may be used as a potential therapeutic agent for treatment of SCI.     

Atorvastatin prevents early apoptosis after thoracic spinal cord contusion injury and promotes locomotion recovery

The systemic administration of atorvastatin has been shown to be neuroprotective after spinal cord injury (SCI), by decreasing the inflammatory response at the lesion site and by reducing neuronal and oligodendrocyte apoptosis. The latter effect spares white matter at the injury site and improves locomotion. The aim of this study was to confirm the neuroprotective efficacy of atorvastatin as well as its early action in limiting apoptosis with its administration post-SCI. Female Sprague-Dawley rats received an intra peritoneal injection of: (1) statin/saline (5 mg/kg) at 2 h after the contusion injury; (2) physiological saline at 2 h post-SCI; or (3) physiological saline without injury. Statin-treated rats showed significant (p < 0.05) improvement in locomotion at week 4 post-SCI compared to vehicle-treated animals. Explaining this outcome, caspase-3 activity decreased by 50% (p < 0.05), and the histological TUNEL method revealed a decrease of approximately 20% in apoptotic cells at the injury site (p < 0.01) at 4 h post-SCI in atorvastatin-treated rats in comparison to vehicle-treated controls. These data demonstrate that atorvastatin is effective after experimental spinal cord contusion injury in preventing early apoptosis at the injury site within 2 h post-administration.     

Electrically evoked locomotor activity in the turtle spinal cord hemi-enlargement preparation

We assessed the locomotor capacity of the left half of the spinal cord hindlimb enlargement in low-spinal turtles. Forward swimming was evoked in the left hindlimb by electrical stimulation of the right dorsolateral funiculus (DLF) at the anterior end of the third postcervical spinal segment (D3). Animals were held by a band-clamp in a water-filled tank so that hindlimb movements could be recorded from below with a digital video camera. Left hindlimb hip and knee movements were tracked while electromyograms (EMGs) were recorded from left hip and knee muscles. In turtles with intact spinal cords, electrical stimulation of the right D3 DLF evoked robust forward swimming movements of the left hindlimb, characterized by rhythmic alternation between hip flexor (HF) and hip extensor (HE) EMG discharge, with knee extensor (KE) bursts occurring during the latter part of each HE-off phase. After removing the right spinal hemi-enlargement (D8-S2), DLF stimulation still evoked rhythmic locomotor movements and EMG bursts in the left hindlimb that included HF-HE alternation and KE discharge. However, post-surgical movements and EMG bursts had longer cycle periods, and movements showed lower amplitudes compared to controls. These results show that (1) sufficient locomotor CPG circuitry resides within the turtle spinal hemi-enlargement to drive major components of the forward swim motor pattern, (2) contralateral circuitry contributes to the excitation of the locomotor CPG for a given limb, and (3) a sufficient portion of the descending DLF pathway crosses over to the contralateral cord anterior to the hindlimb enlargement to activate swimming.     

Level of spinal cord lesion determines locomotor activity in spinal man

Recent studies have demonstrated that coordinated stepping movements can be induced in patients with complete para-/tetraplegia, when they were standing on a moving treadmill with their body weight partially unloaded and external assistance. The aim of this study was to determine which part of the spinal cord generated the locomotor pattern. In patients with complete paraplegia due to lesions at different levels of the spinal cord, the locomotor pattern was compared with that of healthy subjects. Any similarities in electromyographic (EMG) activity of gastrocnemius and tibialis anterior muscles between the patients and healthy subjects were reflected by the analysis of the variation ratio and amplitudes of the EMG activity. It was found that the higher the level of spinal cord lesion the more ”normal” was the locomotor pattern. This suggests that neuronal circuits underlying locomotor ”pattern generation” in man are not restricted to any specific level(s) of the spinal cord, but that an intricate neuronal network contributing to bipedal locomotion extends from thoracolumbal to cervical levels. Received: 16 October 1998 / Accepted: 21 April 1999     

Interleukin-17 deficiency improves locomotor recovery and tissue sparing after spinal cord contusion injury in mice

Following the initial impact, spinal cord injury (SCI) triggers a number of inflammatory responses which can exacerbate tissue damage in the cord and impair functional recovery. The involvement of several pro-inflammatory cytokines in the secondary degenerative mechanisms of SCI has been well established, although the role of interleukin-17 (IL-17) remains unclear. In the present study, we used IL-17 knockout (KO) and C57BL/6J wildtype (WT) mice to investigate the effects of IL-17 deficiency on locomotor recovery, lesion size, glial activation and inflammatory cell response following spinal cord contusion injury. Our results show that compared to WT mice, IL-17 KO mice had a significantly smaller lesion size, corresponding with significantly improved locomotor functional recovery following SCI. At 6 weeks after injury, recruitment of B cells, dendritic cells and neutrophils was significantly lower in IL-17 KO than WT mice, however there was no difference in the presence of activated microglia and reactive astrocytes, in the injured spinal cord. These findings suggest that IL-17 is a mediator of secondary degeneration, which contributes to neuroinflammation and hinders functional recovery, though its actions do not affect glial activation following SCI.     

Metallothionein-II improves motor function recovery and increases spared tissue after spinal cord injury in rats

After spinal cord injury (SCI), a complex cascade of pathophysiological processes rapidly damages the nervous tissue. The initial damage spreads to the surrounding tissue by different mechanisms, including oxidative stress. We have recently reported that the induction of metallothionein (MT) protein is an endogenous rapid-response mechanism after SCI. Since the participation of MT in neuroprotective processes after SCI is still unknown, the aim of the present study was to evaluate the possible neuroprotective effect of exogenously administered MT-II during the acute phase after SCI in rats. Female Wistar rats weighing 200-250g were submitted to spinal cord contusion by means of a computer-controlled device (NYU impactor). Rats received several doses of MT-II (3.2, 10 and 100μg) at 2 and 8h after SCI. Results of the BBB scale were statistically analysed using an ANOVA of repeated-measures, followed by Tukey's test. Among the three doses tested, only 10 and 100μg were able to significantly increase (p<0.05) BBB scale scores eight weeks after SCI from a mean of 7.88 in the control group, to means of 12.63 and 10.88 for the 10 and 100μg doses of MT-II, respectively. The amount of spared tissue was also higher in the groups treated with 10 and 100μg, as compared to the control group values. Results from the present study demonstrate a significant neuroprotective effect of exogenously administered MT-II. Further studies are needed in order to characterize the mechanisms involved in this neuroprotective action.     

5-Hydroxytryptamine-induced locomotor rhythm in the neonatal mouse spinal cord in vitro

We examined the 5-hydroxytryptamine (5-HT)-induced locomotor rhythm in isolated spinal cord preparations taken from neonatal mice on postnatal day (P) 0–3. Motor activity was recorded from L2 and L5 ventral roots. Bath application of 5-HT (15–100 μM) evoked rhythmic bursts that alternated between the two sides, and the bursts in the L2 ventral root alternated with those in the ipsilateral L5 ventral root. After transection of the mid-lumbar cord, the locomotor rhythm in L2 persisted, while that in the L5 ventral root was abolished. This suggests that the upper lumbar region has a greater ability to generate a locomotor rhythm than the lower lumbar spinal cord. Kynurenate, a broad-spectrum glutamate receptor antagonist, blocked the 5-HT-induced locomotor rhythm indicating that ionotropic glutamate receptors are required for the rhythm to be generated.     

观察脊柱脊髓损伤不同时间段手术的术后神经功能恢复情况

目的对脊髓损伤患者不同时间段尽早急诊手术的术后神经功能康复情况进行观察。方法选取35例脊柱脊髓损伤并进行创伤外科急诊手术治疗患者的临床资料,进行统计并分析比较其不同时间段进行手术,术后的神经功能康复情况。结果 35例患者术后神经功能恢复均较术前明显好转,8小时手术的患者神经功能康复较8小时的患者好,越早进行手术的患者神经功能康复越好,所有患者术后的脊髓功能ASIA评分(针刺觉评分、轻触觉评分、运动评分)均较术前提高,且时间越早手术的术后ASIA评分提高值就越高,与术前、不同时间段之间比较P0.05,差异具有统计学意义。结论对于脊髓损伤的患者需加快其院外及院内的抢救,以最大限度降低患者的损伤,提高其术后神经功能康复力。     

护理干预对大鼠脊髓损伤后运动功能修复的影响

目的探索护理干预对脊髓损伤大鼠运动功能恢复程度的影响。方法将60只大鼠随机分为A、B、C共3个组(每组20只),A组为正常对照组,B组为实验对照组,C组为实验组;B组给予常规护理,C组在给予常规护理的基础上给予肌肉按压、关节被动运动、皮肤护理等护理干预。应用BBB评分和斜坡试验评价大鼠脊髓损伤后不同时相点的行为学变化,采用多道生理信号采集系统和电子天平称量,观察大鼠腓肠肌在脊髓损伤后不同时间点肌电反应和肌湿重变化。结果 BBB评分和斜坡试验结果基本一致,C组与B组相比,C组大鼠后肢功能明显改善,但与A组(正常对照组)相比,其后肢功能评分相差很大;腓肠肌肌电反应纤颤电位波幅C组较B组高;腓肠肌湿重下降C组较B组减轻。结论护理干预可延缓肌肉萎缩速度、改善运动功能,促进损伤脊髓功能的部分运动功能恢复。     

护理干预对大鼠脊髓损伤后运动功能修复的脊髓形态学研究

目的研究护理干预对大鼠脊髓损伤后脊髓运动功能修复的脊髓形态学变化。方法将60只SD成年大鼠随机分为3组,分别为正常对照组、实验对照组、实验组,每组20只,每组分4个时相点,即脊髓损伤后1 d、7 d、30 d、60 d(n=5)。实验对照组和实验组均采用切割加挤压脊髓L4平面横断制备脊髓损伤大鼠模型。正常对照组为正常大鼠未经处理,实验对照组大鼠脊髓损伤后给予排尿、排便等常规护理,实验组除常规护理之外,还给予关节活动度训练和肌肉按摩训练,每日2次,每次10 min。采用常规HE染色、免疫组织化学染色法观察护理干预对脊髓损伤后大鼠脊髓的形态学变化。结果 HE染色结果以及GFAP和NF-200免疫组织化学染色结果显示,实验组和实验对照组未见明显区别,但与正常对照组比较差异非常明显。结论护理干预对损伤区脊髓组织形态学研究未见明显改变。     

c-Jun氨基末端激酶基因表达与急性脊髓损伤大鼠神经功能修复状态的关联研究

目的探索急性脊髓损伤后大鼠脊髓组织中c-Jun氨基末端激酶(c-Jun amino-terminal kinase,JNK)基因表达与神经功能修复状态的关联。方法取健康成年雄性大鼠50只,随机选取,假手术对照组18只,实验组32只。损伤后1、12、24、72 h行PCR检测,比较大鼠脊髓组织中JNK(JNK1、JNK2、JNK3)表达水平变化;损伤后12、72 h选取L_(4/5)节段脊髓行HE染色、,比较脊髓损伤大鼠脊髓组织细胞凋亡情况、神经功能变化状态(NSS)。结果 PCR结果显示,髓损伤大鼠脊髓组织中JNK1、JNK2、JNK3的表达,在损伤后1、12、24、72 h除JNK3在相应时点表达水平与假手术组比较水平下降外,其他两项表达与假手术对照组无明显差别。L4/5脊髓HE染色结果提示,与假手术组比较,损伤后12 h损伤的脊髓皮质周边区HE染色阳性细胞显著增多。随着时间推移,染色阳性细胞出现明显减少的趋势,损伤后12 h的染色阳性细胞明显少于72 h。损伤组NSS评分则明显更高,但脊髓损伤组损伤后不同时点比较,NSS评分渐减。结论脊髓损伤后出现JNK3表达下调的情况,可能与神经细胞凋亡以及促进损伤后神经功能的恢复有关联。     

美金刚对急性脊髓损伤大鼠运动功能的影响

目的:探讨美金刚对急性脊髓损伤大鼠运动功能恢复的影响及相关作用机制。方法:80只健康成年雄性SD大鼠,采用改良的Allen打击法(10 g×2.5 cm)建立大鼠T12脊髓损伤动物模型,并随机分为假手术组(A组)、模型组(B组)、美金刚组(C组)及MK-801组(D组),每组20只。术前及术后24 h、72 h、5 d、7 d进行运动功能BBB评分,并采用黄嘌呤氧化法和硫代巴比妥酸法检测损伤脊髓节段组织匀浆超氧化物歧化酶(SOD)和丙二醛(MDA)的水平;HE、尼氏染色观察损伤脊髓组织的病理学改变及Tunel法检测神经细胞的凋亡。结果:与模型组比较,美金刚组及MK-801组大鼠伤后各时间点的脊髓组织SOD活性显著升高、MDA含量显著下调、BBB运动功能评分增加、神经细胞凋亡率降低,差异均具有统计学意义(P0.05)。HE及尼氏染色观察可见美金刚组及MK-801组大鼠较模型组灰质出血量、白质结构紊乱、炎性细胞浸润程度等都有明显改善。结论:美金刚可以通过阻断过度激活的NMDA受体功能,减轻大鼠脊髓损伤后氧化应激反应,减少脊髓损伤处神经细胞的凋亡,促进脊髓损伤大鼠运动功能的恢复。     

活性氧在脊髓损伤中的作用及损伤机制

目的研究脊髓损伤(SCI)后脊髓组织中丙二醛(MDA)的动态水平变化和神经细胞凋亡及其凋亡相关因子表达的变化,探讨活性氧(ROS)在SCI后的作用及其分子机制。方法成年雄性大鼠132只,用改良的Allen法复制大鼠急性SCI模型,致大鼠脊髓(T10)中度挫伤,采用化学比色法测定不同时间段受损脊髓组织的MDA含量;免疫组织化学法分析受损脊髓组织区域凋亡相关因子Caspase-3、Bcl-2和Bax的表达变化;荧光原位缺口末端标记法(TUNEL)检测细胞凋亡。结果SCI后6h,脊髓组织中MDA含量明显增高并维持到损伤后5d,期间在6h和3d出现两次高峰,7d基本恢复正常;SCI后6h脊髓组织中凋亡细胞开始增多,3d达高峰,以后逐渐减少,各时间点与假手术组比较有显著性的差异;Bcl-2和Bax蛋白损伤后6h开始有较多的表达,以后快速增多,5d达高峰,然后逐渐回落。Caspase-3蛋白在损伤后6h开始增多,3d达高峰,以后逐渐减少。3种凋亡相关因子在各时间点与假手术组比较有显著性差异;甲基强的松龙(MPSS)治疗组与SCI组比较:MDA含量、凋亡细胞数、凋亡因子Caspase-3和Bax表达减少,Bcl-2表达增加,并且在部分时间点有显著性差异。结论在SCI后ROS可能通过促进Caspase-3表达和降低Bcl-2/Bax之间的比值诱导神经细胞凋亡,从而加重了SCI继发性损伤。     

Recovery of hindlimb motor functions after spinal cord transection is enhanced by grafts of the embryonic raphe nuclei

In this study, a piece of embryonic tissue from the raphe nucleus was transplanted into the spinal cord below the lesion 1 month after transection. Two months later the recovery of hindlimb motor function in rats which had received a transplant of neural tissue (ST rats) was much better than in spinal control animals without the graft (SC rats). Analysis of the electromyographic (EMG) activity showed that the timing of muscle activity during locomotor-like movement of hindlimbs in ST rats was more regular than in SC rats. In SC rats the relationships between EMG burst duration (soleus, tibialis anterior) and step cycle duration were significantly altered. The restoration of hindlimb motor function of ST rats was also reflected in the better interlimb coordination during locomotor-like hindlimb movements. The results of several behavioural tests demonstrated that the responses to stimulation of various receptors, such as tactile or proprioceptive, in ST rats were more complex than in SC rats. Additionally, unlike in SC animals, in ST rats long-lasting spontaneous episodes of air stepping movement of hindlimbs accompanied by a relatively high amplitude of EMG activity were obtained. These results confirm that grafted embryonic raphe nuclei which contain serotoninergic cells are likely to increase the excitability of neuronal circuitry in the injured spinal cord. Moreover, transplantation of embryonic raphe nuclei encourages the recovery of hindlimb motor function in adult rats even when the grafting is carried out several weeks after spinal cord injury.     

Quadriplegia recovery after hemi-section and transplant model of spinal cord at the level of C5 and C6

A spinal cord hemi-section with a homologous transplant of medullar tissue at the level of C5–C6 and preservation of the anterior spinal artery was used to evaluate the histological characteristics such as quantity and quality of axons, myelin index and blood vessels after quadriplegia recovery. Vascular changes after spinal injury results in severe endothelial damage, axonal edema, neuronal necrosis and demyelinization as well as cysts and infarction. Preservation of the anterior spinal artery has demonstrated clinical recuperation; therefore, in addition to the lesion we included a homologous transplant to visualize changes at a cellular level. Two groups of dogs (hemi-section and transplant) went through a traumatic spinal cord hemi-section of 50% at the level of C5–C6. The transplant group formed by animals which simultaneously had 4 mm of spinal cord removed and the equal amount substituted from a donor animal at the level of C5–C6 corresponding to the half right side; both preserving the anterior spinal artery. Histological evaluation of all groups took place at days 3 (acute) and 28 (chronic) post-operation. Changes of degeneration and axonal regeneration were found in the hemi-section and transplant groups at acute and chronic time, as well as same quadriplegia recovery at chronic time in the hemi-section and transplant groups which closely related to mechanisms which participate in regeneration and functional recuperation due to the preservation of the anterior spinal artery and presence of new blood vessels.     

c-Jun N-terminal kinase induces axonal degeneration and limits motor recovery after spinal cord injury in mice

C-Jun N-terminal kinase (JNK) mediates neuronal death in response to stress and injury in the CNS and peripheral nervous system. Here, we show that JNK also regulates retrograde axonal degeneration (axonal dieback) after spinal cord injury (SCI) in mice. Activated phospho-JNK was highly expressed in damaged corticospinal tract (CST) axons after thoracic SCI by hemisection. Local administration of SP600125, a JNK inhibitor, prevented accumulation of amyloid-β precursor protein and retraction of the severed CST axons as well as preserved the axonal arbors rostral to the injury site. The treatment with SP600125 also improved functional recovery of the hindlimbs, assessed by Basso mouse scale open-field scores and the grid-walking test. In Jnk1^−/− and Jnk3^−/− mice, we observed prevention of axonal degeneration and enhancement of motor recovery after SCI. These results indicate that both JNK1 and JNK3 induce axonal degeneration and limit motor recovery after SCI. Thus, a JNK inhibitor may be a suitable therapeutic agent for SCI.     

Simvastatin treatment improves functional recovery after experimental spinal cord injury by upregulating the expression of BDNF and GDNF

The aim of this study was to determine the therapeutic efficacy of simvastatin treatment starting 1 day after spinal cord injury (SCI) in rat and to investigate the underlying mechanism. Spinal cord injury was induced in adult female Sprague–Dawley rats after laminectomy at T9-T10. Then additionally with sham group (laminectomy only) the SCI animals were randomly divided into 3 groups: vehicle-treated group; 5-mg/kg simvastatin-treated group; and 10-mg/kg simvastatin-treated group. Simvastatin or vehicle was administered orally at 1 day after SCI and then daily for 5 weeks. Locomotor functional recovery was assessed during 8 weeks postoperation by performing open-field locomotor test and inclined-plane test. At the end of study, motor evoked potentials (MEPs) and somatosensory evoked potentials (SEPs) were assessed to evaluate the integrity of spinal cord pathways. Then, the animals were killed, and 1-cm segments of spinal cord encompassing the injury site were removed for histopathological analysis. Immunohistochemistry was performed to observe the expression of brain-derived neurotrophic factor (BDNF), glial cell line-derived neurotrophic factor (GDNF) in the spinal cord. Results show that the simvastatin-treated animals showed significantly better locomotor function recovery, better electrophysiological outcome, less myelin loss, and higher expression of BDNF and GDNF. These findings suggest that simvastatin treatment starting 1 day after SCI can significantly improve locomotor recovery, and this neuroprotective effect may be related to the upregulation of BDNF and GDNF. Therefore, simvastatin may be useful as a promising therapeutic agent for SCI.