Reducing excessive GABA-mediated tonic inhibition promotes functional recovery after stroke

Stroke is a leading cause of disability, but no pharmacological therapy is currently available for promoting recovery. The brain region adjacent to stroke damage-the peri-infarct zone-is critical for rehabilitation, as it shows heightened neuroplasticity, allowing sensorimotor functions to re-map from damaged areas. Thus, understanding the neuronal properties constraining this plasticity is important for the development of new treatments. Here we show that after a stroke in mice, tonic neuronal inhibition is increased in the peri-infarct zone. This increased tonic inhibition is mediated by extrasynaptic GABA(A) receptors and is caused by an impairment in GABA (γ-aminobutyric acid) transporter (GAT-3/GAT-4) function. To counteract the heightened inhibition, we administered in vivo a benzodiazepine inverse agonist specific for α5-subunit-containing extrasynaptic GABA(A) receptors at a delay after stroke. This treatment produced an early and sustained recovery of motor function. Genetically lowering the number of α5- or δ-subunit-containing GABA(A) receptors responsible for tonic inhibition also proved beneficial for recovery after stroke, consistent with the therapeutic potential of diminishing extrasynaptic GABA(A) receptor function. Together, our results identify new pharmacological targets and provide the rationale for a novel strategy to promote recovery after stroke and possibly other brain injuries.     

Functional regeneration of respiratory pathways after spinal cord injury

Spinal cord injuries often occur at the cervical level above the phrenic motor pools, which innervate the diaphragm. The effects of impaired breathing are a leading cause of death from spinal cord injuries, underscoring the importance of developing strategies to restore respiratory activity. Here we show that, after cervical spinal cord injury, the expression of chondroitin sulphate proteoglycans (CSPGs) associated with the perineuronal net (PNN) is upregulated around the phrenic motor neurons. Digestion of these potently inhibitory extracellular matrix molecules with chondroitinase ABC (denoted ChABC) could, by itself, promote the plasticity of tracts that were spared and restore limited activity to the paralysed diaphragm. However, when combined with a peripheral nerve autograft, ChABC treatment resulted in lengthy regeneration of serotonin-containing axons and other bulbospinal fibres and remarkable recovery of diaphragmatic function. After recovery and initial transection of the graft bridge, there was an unusual, overall increase in tonic electromyographic activity of the diaphragm, suggesting that considerable remodelling of the spinal cord circuitry occurs after regeneration. This increase was followed by complete elimination of the restored activity, proving that regeneration is crucial for the return of function. Overall, these experiments present a way to markedly restore the function of a single muscle after debilitating trauma to the central nervous system, through both promoting the plasticity of spared tracts and regenerating essential pathways.     

硫酸软骨素裂解酶ABC产生菌的筛选及酶学性质的初步研究

从成都市井研县生猪屠宰厂等地采集的样品中筛选出一株高产、稳定的硫酸软骨素裂解酶ABC产生菌株GT38,初步鉴定为彭氏变形杆菌(P．penneri),其酶活力单位高达112U／L．酶学性质经初步研究表明：该酶的最适反应pH值为7．0,最适反应温度为30℃,在pH值6～8之间较稳定,温度在30℃～45℃较稳定,金属离子对酶活力影响不大．     

New strategies for the repair of spinal cord injury

Spinal cord injury （SCI） leads to several neu- rologic damages to patients with sensory and motor func- tion loss. Although some inspiring results were reported in animal, even in non-human primate models, the clinical translation of these research achievements barely move forward for the SCI patients. This review mainly focused on the pathogenesis of the primary and secondary injury, the clinical therapies and the new technologies in spinal cord injury. Moreover, the key scientific issues on this field were discussed in the review. Finally, some suggestions were proposed for the scientific and clinical researches.     

Spinal cord decompression reduces rat neural cell apoptosis secondary to spinal cord injury

Objective: To determine whether spinal cord decompression plays a role in neural cell apoptosis after spinal cord injury. Study design: We used an animal model of compressive spinal cord injury with incomplete paraparesis to evaluate neural cell apoptosis after decompression. Apoptosis and cellular damage were assessed by staining with terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate nick-end labelling (TUNEL) and immunostaining for caspase-3, Bcl-2 and Bax. Methods: Experiments were conducted in male Sprague-Dawley rats (n=78) weighing 300~400 g. The spinal cord was compressed posteriorly at T10 level using a custom-made screw for 6 h, 24 h or continuously, followed by decompression by removal of the screw. The rats were sacrificed on Day 1 or 3 or in Week 1 or 4 post-decompression. The spinal cord was removed en bloc and examined at lesion site, rostral site and caudal site (7.5 mm away from the lesion). Results: The numbers of TUNEL-positive cells were significantly lower at the site of decompression on Day 1, and also at the rostral and caudal sites between Day 3 and Week 4 post-decompression, compared with the persistently compressed group. The numbers of cells between Day 1 and Week 4 were immunoreactive to caspase-3 and B-cell lymphoma-2 (Bcl-2)-associated X-protein (Bax), but not to Bcl-2, correlated with those of TUNEL-positive cells. Conclusion: Our results suggest that decompression reduces neural cell apoptosis following spinal cord injury.     

基因重组碱性成纤维细胞生长因子对大鼠脊髓损伤神经保护作用

目的 :探索碱性成纤维细胞生长因子 (bFGF)对大鼠脊髓损伤的神经保护作用 方法 :将吸入bFGF的胶原蛋白海绵或空白海绵贴敷于大鼠脊髓损伤处 ,术后 1、2、3周 ,对大鼠机能进行评分 ,并对大脑运动皮质进行电镜观察分析 结果 :术后 1、2、3周 ,bFGF组大鼠运动评分均明显优于对照组 (Ρ <0 .0 5) ,运动皮质电镜结果显示bFGF组线粒体、内质网轻度肿胀 ,神经毡结构正常 ,无明显胶质细胞增生 程度较对照组明显减轻 结论 :bFGF对大鼠脊髓受损神经纤维起源脑区—运动皮质的神经细胞具有明显的保护作用 ,进而使大鼠运动功能受损明显减轻     

Same impact momentum causes different degrees of spinal cord injury

The weight-drop impact is widely used in making animal model of spinal cord injury （SCI）. But there has not yet been an appropriate unit for the quantification of the impact. In this study, we compared the degrees of the spinal cord injury caused by weight-drop impact with the same momentum but different combinations of drop weight and drop height, in order to test whether ＂momentum＂ is capable of being the unit for the quantification of weight- drop impact. Thirty adult rats were randomly allocated to three groups and were injured with 5 g-10 cm （group A）, 10g-2.5 cm （group B） and 15 g-l.ll cm （group C） impacts with the same momentum to the spinal cord, respectively. Open-field locomotor function was evaluated using the Basso-Beattie-Bresnahan （BBB） locomotor rat- ing scale. The percentage of spared tissue area （STA） at the epicenter, and 500, 1000 and 1500 gm from the epicenter was calculated using serial sections stained by hematoxylin and eosin. As a result, the behavioral recovery （BBB scores） and the STA percentage were similar in group B and group C. However, the BBB score in group A was significantly lower than that in groups B and C at the same time point post injury. The STA percentage was signifi- cantly less and the lesion/cavity length was significantlygreater in group A than in groups B and C. These suggested that the 5 g-10 cm weight-drop impact, compared with the other two impacts with different weights and heights, caused a greater damage of the spinal cord when the momentum was the same. So, these impacts with the same momentum but different weights and drop heights cause different degrees of spinal cord injury. Momentum alone is inadequate to be the unit for the qualification of weight- drop impact and to be used to predict the extent of injury.     

补阳还五汤对脊髓损伤大鼠体重及后肢运动功能的影响

目的探讨补阳还五汤对脊髓损伤大鼠体重及后肢运动功能影响与其神经保护作用的关系。方法采用Allen’s打击法制作中度SCI模型,应用SPF级Wistar大鼠,随机分为4组：正常组和3个模型组,即模型空白组、BYHWD组、金纳多组,每组16只大鼠。模型组大鼠分别给予蒸馏水、BYHWD、金纳多水溶液,正常组给予蒸馏水,分别于造模前即0 d、造模后3 d、1 w、2 w、3 w,称量大鼠体重,并进行后肢运动功能评价,灌胃治疗2 w后,结束实验。结果SCI模型大鼠于造模后3 d体重及后肢运动功能均明显降低,灌胃治疗后,模型空白组大鼠体重缓慢增加,但后肢运动功能无明显改善;BYHWD组及金纳多组大鼠体重及后肢运动功能均有显著改善,且BYHWD组改善效果更明显,与金纳多组比较差异有显著性。结论BYHWD通过保护SCI大鼠脊髓组织,显著改善大鼠后肢运动功能,增加大鼠体重,为其治疗中医＂痿证＂提供了新的实验依据。     

Changes in spinal cord reflexes after cross-anastomosis of cutaneous and muscle nerves in the adult rat

Evidence exists that the specification of afferent nerves and their central connections in the embryo may depend in part on influences from the peripheral target innervated. We have now investigated whether such peripheral determination persists in the adult rat using the unmyelinated afferent system of C fibres, which differ chemically in the adult depending on their target. We have previously shown that if the cutaneous sural nerve and the muscle gastrocnemius nerve are cross-anastomosed so that they grow to each other's target, the A fibres establish functional endings and the C fibres change their chemistry to that which is appropriate for the new target. Here we report that in normal adult rats, a short train of stimuli to the cutaneous sural nerve produced a brief facilitation of the flexion reflex, lasting on average only 5 min, whereas similar stimulation of the gastrocnemius-muscle nerve enhanced this reflex for an average of 54 min. In cross-anastomosed animals, stimulation of the gastrocnemius nerve (innervating skin) induced a brief potentiation of the flexion reflex, lasting on average only 3 min. By contrast, stimulation of sural nerve (innervating muscle) produced a potentiation of this reflex lasting 57 min. Thus the ability of adult afferent nerves to potentiate the flexion reflex depends on the target with which they make contact. We propose that tissue-specific factors influence some of the central actions of primary afferent neurons in the adult.     

A topographic map of recruitment in spinal cord

Animals move over a range of speeds by using rhythmic networks of neurons located in the spinal cord. Here we use electrophysiology and in vivo imaging in larval zebrafish (Danio rerio) to reveal a systematic relationship between the location of a spinal neuron and the minimal swimming frequency at which the neuron is active. Ventral motor neurons and excitatory interneurons are rhythmically active at the lowest swimming frequencies, with increasingly more dorsal excitatory neurons engaged as swimming frequency rises. Inhibitory interneurons follow the opposite pattern. These inverted patterns of recruitment are independent of cell soma size among interneurons, but may be partly explained by concomitant dorso-ventral gradients in input resistance. Laser ablations of ventral, but not dorsal, excitatory interneurons perturb slow movements, supporting a behavioural role for the topography. Our results reveal an unexpected pattern of organization within zebrafish spinal cord that underlies the production of movements of varying speeds.     

P2X4 receptors induced in spinal microglia gate tactile allodynia after nerve injury

Pain after nerve damage is an expression of pathological operation of the nervous system, one hallmark of which is tactile allodynia-pain hypersensitivity evoked by innocuous stimuli. Effective therapy for this pain is lacking, and the underlying mechanisms are poorly understood. Here we report that pharmacological blockade of spinal P2X4 receptors (P2X4Rs), a subtype of ionotropic ATP receptor, reversed tactile allodynia caused by peripheral nerve injury without affecting acute pain behaviours in naive animals. After nerve injury, P2X4R expression increased strikingly in the ipsilateral spinal cord, and P2X4Rs were induced in hyperactive microglia but not in neurons or astrocytes. Intraspinal administration of P2X4R antisense oligodeoxynucleotide decreased the induction of P2X4Rs and suppressed tactile allodynia after nerve injury. Conversely, intraspinal administration of microglia in which P2X4Rs had been induced and stimulated, produced tactile allodynia in naive rats. Taken together, our results demonstrate that activation of P2X4Rs in hyperactive microglia is necessary for tactile allodynia after nerve injury and is sufficient to produce tactile allodynia in normal animals. Thus, blocking P2X4Rs in microglia might be a new therapeutic strategy for pain induced by nerve injury.     

bFGF对大鼠坐骨神经损伤后腓肠肌功能恢复的影响

目的：观察碱性成纤维细胞生长因子（ｂ Ｆ Ｇ Ｆ） 对大鼠坐骨神经损伤后腓肠肌功能恢复的作用。方法：以钳夹损伤左侧坐骨神经大鼠为模型,治疗组肌肉注射ｂ Ｆ Ｇ Ｆ,对照组注射等渗盐水,隔日１ 次。术后２ 、３ 、４ 周分别对治疗组和对照组损伤侧及健侧腓肠肌进行肌诱发电位及肌收缩力检测,以健侧为１００ ％ ,求出损伤侧各指标的恢复率。结果：与对照组相比,治疗组腓肠肌诱发电位幅值恢复率明显加快（ Ｐ＜ ００５） ;腓肠肌单收缩力和强直收缩力的恢复率也显著加强（ Ｐ＜ ００５ 和 Ｐ＜ ００１） 。结论：ｂ Ｆ Ｇ Ｆ 能促进大鼠坐骨神经损伤后骨骼肌运动功能的恢复     

Effect of Jiaji electroacupuncture in transected rat spinal cord

We evaluated the effect of Jiaji electroacupuncture on cell proliferation and the expression of markers of endogenous neural stem cell activation after complete spinal cord transection. Female Wistar rats were assigned to 4 groups (n = 24 each): a sham-operated group, a control group, a Jiaji electroacupuncture group, and a Jiaji electroacupuncture preconditioning group. Motor function was significantly improved in the acupuncture groups compared to the control group at 7 and 14 d. Numbers of bromodeoxyuridine (BrdU)-, nestin-, and glial fibrillary acidic protein (GFAP)-positive cells were significantly greater in the acupuncture groups than in the controls at each time point. Expression of nestin and GFAP mRNA was significantly higher in the acupuncture groups than in the controls at each time point. Thus, Jiaji electroacupuncture and preconditioning may promote the proliferation of endogenous neural stem cells after spinal cord transection.     

Functional regeneration of sensory axons into the adult spinal cord

The arrest of dorsal root axonal regeneration at the transitional zone between the peripheral and central nervous system has been repeatedly described since the early twentieth century. Here we show that, with trophic support to damaged sensory axons, this regenerative barrier is surmountable. In adult rats with injured dorsal roots, treatment with nerve growth factor (NGF), neurotrophin-3 (NT3) and glial-cell-line-derived neurotrophic factor (GDNF), but not brain-derived neurotrophic factor (BDNF), resulted in selective regrowth of damaged axons across the dorsal root entry zone and into the spinal cord. Dorsal horn neurons were found to be synaptically driven by peripheral nerve stimulation in rats treated with NGF, NT3 and GDNF, demonstrating functional reconnection. In behavioural studies, rats treated with NGF and GDNF recovered sensitivity to noxious heat and pressure. The observed effects of neurotrophic factors corresponded to their known actions on distinct subpopulations of sensory neurons. Neurotrophic factor treatment may thus serve as a viable treatment in promoting recovery from root avulsion injuries. I