天麻素注射液对蛛网膜下腔出血大鼠NF-κB/Bax/Caspase-3通路及神经元凋亡的影响

目的 探讨天麻素注射液对蛛网膜下腔出血(Subarachnoid hemorrhage,SAH)大鼠核转录因子-κB/B细胞淋巴瘤-2基因相关X蛋白/半胱氨酸天冬氨酸蛋白水解酶-3(Nuclear factor-κB,NF-κB/B cell lymphoma-2 associated X protein,Bax/Cysteinyl aspartate specific proteinase-3,Caspase-3)通路及神经元凋亡的影响。方法 血管穿刺法建立SAH大鼠模型,模型成功50只,随机分为模型组、天麻素低、中、高剂量组(腹腔注射5、10、20 mg·kg^-1·d^-1天麻素注射液)、阳性对照组(腹腔注射0.5 mg·kg^-1·d^-1地塞米松磷酸钠注射液),每组各10只,另10只大鼠设为假手术组; 干预3周后测定大鼠神经功能评分; 流式细胞术检测脑皮层细胞凋亡情况; 原位末端标记法(TdT-mediated dUTP-biotin nick end labeling assay,TUNEL)检测神经元凋亡状态; 蛋白免疫印迹(Western blot,WB)检测大鼠脑皮层NF-κB,Bax,Caspase-3、生长相关蛋白-43(Growth-associated protein-43,GAP-43)、突触素(Synaptophysin,SYN)蛋白表达水平。结果 与假手术组比较,模型组神经功能评分、脑皮层神经元凋亡率、脑组织中NF-κB,Bax,Caspase-3蛋白表达水平升高,脑皮层组织中GAP-43,SYN蛋白表达水平降低(P<0.05)。与模型组比较,天麻素中、高剂量组、阳性对照组神经功能评分、脑皮层神经元凋亡率、脑组织中NF-κB,Bax,Caspase-3蛋白表达水平降低,脑皮层组织中GAP-43,SYN蛋白表达水平升高(P<0.05); 天麻素低剂量组脑皮层神经元凋亡率、脑组织中NF-κB,Bax,Caspase-3蛋白表达水平降低,脑皮层组织中GAP-43,SYN蛋白表达水平升高(P<0.05); 随着天麻素注射液剂量的升高,神经功能评分、脑皮层神经元凋亡率、脑组织中NF-κB,Bax,Caspase-3蛋白表达水平降低,脑皮层组织中GAP-43,SYN蛋白表达水平升高(P<0.05)。结论 天麻素注射液可能通过抑制NF-κB/Bax/Caspase-3通路来实现对神经元凋亡的缓解和对SAH大鼠的脑保护。     

Increased expression of the growth‐associated protein 43 gene in the sensorimotor cortex of the macaque monkey after lesioning the lateral corticospinal tract

To investigate the neural basis for functional recovery of the cerebral cortex following spinal cord injury, we measured the expression of growth‐associated protein 43 (GAP‐43), which is involved in the process of synaptic sprouting. We determined the GAP‐43 mRNA expression levels in the sensorimotor cortical areas of macaque monkeys with a unilateral lesion of the lateral corticospinal tract (l‐CST) at the C4/C5 level of the cervical cord and compared them with the levels in the corresponding regions of intact monkeys. Lesioned monkeys recovered finger dexterity during the first months after surgery, and the GAP‐43 mRNA levels increased in layers II–III in primary motor areas (M1), bilaterally. Double‐labeling analysis of the lesioned monkeys showed that GAP‐43 mRNA was expressed strongly in excitatory neurons but only rarely in inhibitory interneurons. Expression also increased in the medium‐sized (area, 500–1,000 μm²) and large pyramidal cells (area, >1,000 μm²) in layer V of the bilateral M1. The increased expression of GAP‐43 mRNA in the M1 contralateral to the lesion was more prominent during the early recovery stage than during the late recovery stage. In addition, GAP‐43 mRNA increased in layers II–III of both the contralesional ventral premotor area and the primary somatosensory area. These results suggest that GAP‐43 is involved in time‐dependent and brain region‐specific plastic changes after l‐CST lesioning. The expression patterns imply that plastic changes occur not only in M1 but also in the broad associative cortical network, including the ventral premotor and primary sensory areas. J. Comp. Neurol. 516:493–506, 2009. © 2009 Wiley‐Liss, Inc.     

实时定量RT-PCR法检测健侧C7神经移位前后神经生长相关蛋白43的变化**★

背景：已有研究证实，周围神经损伤后大脑皮质会出现功能可塑性变化；神经生长相关蛋白43在发育中神经系统的表达特征提示，它与神经发育中突起延伸和突触形成有关，参与突触重建过程。 目的：实时定量RT-PCR方法检测实验动物健侧颈7神经根移位前后，不同阶段相应大脑皮质组织神经生长相关蛋白 mRNA 的相对表达量及动态变化,探讨周围神经修复与中枢神经可塑性的关系。 设计、时间及地点：随机对照动物实验，于2007-09/2008-12在上海市周围神经重点实验室和复旦大学实验动物科学部完成。 材料：成年SD大鼠108只，分为臂丛损伤未修复组(n＝48)、臂丛损伤修复组(n＝48)及对照组(n＝12)。 方法：大鼠左侧为实验侧，臂丛损伤未修复组在大鼠左侧取锁骨下横形切口，找到臂丛各神经根并造成全臂丛根性撕脱伤。臂丛损伤修复组同法将大鼠全臂丛神经根性撕脱后，取右侧锁骨下切口，显露健侧颈C7神经根备用；取左侧前臂尺神经通过皮下隧道桥接健侧颈7神经根与正中神经端端吻合。对照组为成年雌性正常大鼠，不进行任何处理。 主要观察指标：于术后1 d，3 d，1周，2周，1个月，3个月，6个月，10个月共8个时段及对照组取材，采用SYBR GreenⅠ实时定量RT-PCR法检测健侧颈7神经根移位术前后对侧(右侧)大脑前肢投射区域皮质组织生长相关蛋白43 mRNA相对表达量及动态变化。 结果：臂丛损伤未修复组对侧(右侧)大脑前肢投射区域皮质组织生长相关蛋白43 mRNA的相对表达量变化规律为术后第1天开始升高，第3天达到高峰，然后逐渐降低。术后1 d，3 d和1周与对照组比较差异具有显著性意义 (P < 0.05，P < 0.01)。臂丛损伤修复组对侧(右侧)大脑前肢投射区域皮质组织生长相关蛋白43 mRNA的相对表达量变化规律为术后第1天开始升高，第3天达到高峰，然后逐渐降低，至术后3个月再次升高，6个月达到高峰，然后逐渐降低。术后1 d，3 d，1周和6个月与对照组比较差异具有显著性意义(P < 0.05，P < 0.01)。 结论：臂丛损伤健侧颈7移位术前后相应大脑皮质生长相关蛋白表达变化与临床现象一致，提示生长相关蛋白在大脑皮质及突触可塑性方面发挥作用。     

Intracisternal antisense oligonucleotide to growth associated protein-43 blocks the recovery-promoting effects of basic fibroblast growth factor after focal stroke.

Focal infarction (stroke) of the lateral cerebral cortex of rats (including the sensorimotor cortex) produces deficits in sensorimotor function of the contralateral limbs that recover partially over time. In previous studies, we found that the intracisternal injection of basic fibroblast growth factor (bFGF), a potent neurotrophic growth factor, starting at 1 day after stroke, significantly enhanced recovery of sensorimotor function of the contralateral forelimb and hindlimb. Moreover, immunoreactivity (IR) for growth-associated protein-43 (GAP-43), a molecular marker of new axonal growth, was increased in the intact contralateral sensorimotor cortex following bFGF treatment. In the current study, we found that the intracisternal administration of antisense, but not missense, oligonucleotide to GAP-43 blocked the recovery-enhancing effects of bFGF and blocked the increase in GAP-43 IR in the contralateral cortex. These results suggest that upregulation of GAP-43 expression and consequent enhanced axonal sprouting in intact uninjured parts of the brain are likely mechanisms for the recovery-promoting effects of bFGF.     

Extensive spinal decussation and bilateral termination of cervical corticospinal projections in rhesus monkeys

To examine neuroanatomical mechanisms underlying fine motor control of the primate hand, adult rhesus monkeys underwent injections of biotinylated dextran amine (BDA) into the right motor cortex. Spinal axonal anatomy was examined using detailed serial‐section reconstruction and modified stereological quantification. Eighty‐seven percent of corticospinal tract (CST) axons decussated in the medullary pyramids and descended through the contralateral dorsolateral tract of the spinal cord. Eleven percent of CST axons projected through the dorsolateral CST ipsilateral to the hemisphere of origin, and 2% of axons projected through the ipsilateral ventromedial CST. Notably, corticospinal axons decussated extensively across the spinal cord midline. Remarkably, nearly 2‐fold more CST axons decussated across the cervical spinal cord midline (≈12,000 axons) than were labeled in all descending components of the CST (≈6,700 axons). These findings suggest that CST axons extend multiple segmental collaterals. Furthermore, serial‐section reconstructions revealed that individual axons descending in either the ipsilateral or contralateral dorsolateral CST can: 1) terminate in the gray matter ipsilateral to the hemisphere of origin; 2) terminate in the gray matter contralateral to the hemisphere of origin; or 3) branch in the spinal cord and terminate on both sides of the spinal cord. These results reveal a previously unappreciated degree of bilaterality and complexity of corticospinal projections in the primate spinal cord. This bilaterality is more extensive than that of the rat CST, and may resemble human CST organization. Thus, augmentation of sprouting of these extensive bilateral CST projections may provide a novel target for enhancing recovery after spinal cord injury. J. Comp. Neurol. 513:151–163, 2009. © 2009 Wiley‐Liss, Inc.     

RNA干扰抑制NgR表达对脑梗死大鼠皮质脊髓束可塑性的影响

目的研究应用RNA干扰技术下调Nogo受体(NgR)基因的表达对脑梗死大鼠神经功能恢复及皮质脊髓束可塑性的影响。方法线栓法制作左侧大脑中动脉闭塞脑梗死大鼠模型,用携带U6启动子和NgR特异短发夹RNA(shRNA)编码序列的质粒pNgR-shRNA1、pNgR-shRNA2行缺血侧侧脑室注射给药,以含非特异性shRNA编码序列的无关质粒pGenesil-Con(pCon)注射组及模型组作为对照,应用改良的神经功能缺损评分(mNSS)进行神经功能评定,Western blot检测梗死灶周围脑组织NgR的表达,免疫组织化学技术检测梗死灶周围脑组织生长相关蛋白43(GAP-43)的表达,生物素化葡聚糖胺(BDA)顺行示踪法观察皮质脊髓束可塑性变化。结果 pNgR-shRNA1组、pNgR-shRNA2组mNSS评分明显低于pCon组及模型组(P<0.05),pCon组及模型组mNSS评分差异不明显(P>0.05)。与pCon组及模型组相比,pNgR-shRNA1组、pNgR-shRNA2组梗死灶周围脑组织NgR的表达显著降低(P<0.05),GAP-43的表达明显增强(P<0.05),BDA顺行示踪见病变侧大脑脚水平及病变对侧C1-3脊髓水平BDA阳性纤维数量明显增多(P<0.05)。结论 RNA干扰抑制NgR表达能促进脑梗死大鼠皮质脊髓束可塑性改变及神经功能恢复。     

Subacute intranasal administration of tissue plasminogen activator increases functional recovery and axonal remodeling after stroke in rats

As a thrombolytic agent, application of recombinant tissue plasminogen activator (tPA) to ischemic stroke is limited by the narrow time window and side effects on brain edema and hemorrhage. This study examined whether tPA, administered by intranasal delivery directly targeting the brain and spinal cord, provides therapeutic benefit during the subacute phase after stroke. Adult male Wistar rats were subjected to permanent right middle cerebral artery occlusion (MCAo). Animals were treated intranasally with saline, 60 μg or 600 μg recombinant human tPA at 7 and 14days after MCAo (n=8/group), respectively. An adhesive-removal test and a foot-fault test were used to monitor functional recovery. Biotinylated dextran amine (BDA) was injected into the left motor cortex to anterogradely label the corticorubral tract (CRT) and the corticospinal tract (CST). Naive rats (n=6) were employed as normal control. Animals were euthanized 8 weeks after stroke. Compared with saline treated animals, significant functional improvements were evident in rats treated with 600 μg tPA (p<0.05), but not in 60 μg tPA treated rats. Furthermore, 600 μg tPA treatment significantly enhanced both CRT and CST sprouting originating from the contralesional cortex into the denervated side of the red nucleus and cervical gray matter compared with control group (p<0.01), respectively. The behavioral outcomes were highly correlated with CRT and CST axonal remodeling. Our data suggest that delayed tPA intranasal treatment provides therapeutic benefits for neurological recovery after stroke by, at least in part, promoting neuronal remodeling in the brain and spinal cord.     

Bone marrow stromal cells enhance inter- and intracortical axonal connections after ischemic stroke in adult rats

We investigated axonal plasticity in the bilateral motor cortices in rats after unilateral stroke and bone marrow stromal cell (BMSC) treatment. Rats were subjected to permanent right middle cerebral artery occlusion followed by intravenous administration of phosphate-buffered saline or BMSCs 1 day later. Adhesive-removal test and modified neurologic severity score were performed weekly to monitor limb functional deficit and recovery. Anterograde tracing with biotinylated dextran amine injected into the right motor cortex was used to assess axonal sprouting in the contralateral motor cortex and ipsilateral rostral forelimb area. Animals were killed 28 days after stroke. Progressive functional recovery was significantly enhanced by BMSCs. Compared with normal animals, axonal density in both contralateral motor cortex and ipsilateral rostral forelimb area significantly increased after stroke. Bone marrow stromal cells markedly enhanced such interhemispheric and intracortical connections. However, labeled transcallosal axons in the corpus callosum were not altered with either stroke or treatment. Both interhemispheric and intracortical axonal sprouting were significantly and highly correlated with behavioral outcome after stroke. This study suggests that, after stroke, cortical neurons surviving in the peri-infarct motor cortex undergo axonal sprouting to restore connections between different cerebral areas. Bone marrow stromal cells enhance axonal plasticity, which may underlie neurologic functional improvement.     

Decreased GAP-43/B-50 phosphorylation in striatal synaptic plasma membranes after circling motor behavior during development

We evaluated the in vitro phosphorylation of the presynaptic substrate of protein kinase C (PKC), GAP-43/B-50 and the PKC activity in the striatum of rats submitted to a circling training (CT) test during postnatal development. Motor activity at 30 days of age, but not at other ages, produced a unilateral reduction (-29.5%; p<0.001) in the level of GAP-43/B-50 endogenous phosphorylation in the contralateral striatum with respect to the ipsilateral side, while non-trained control animals did not show asymmetric differences. Compared to controls, the contralateral striatum of trained animals also showed a significant reduction (-29.3%; p<0. 001) in the incorporation of 32P-phosphate into GAP-43. This decreased in vitro GAP-43 phosphorylation was seen at 30 min, but not immediately after circling motor behavior. This contralateral change in GAP-43 phosphorylation correlated with the running speed developed by the animals [(r=0.9443, p=0.0046, n=6, relative to control group) and (r=0.8813, p=0.0203, n=6, with respect to the ipsilateral side of the exercised animals)]. On the contrary, GAP-43/B-50 immunoblots did not show changes in the amount of this phosphoprotein among the different experimental groups. Back phosphorylation assays, performed in the presence of bovine purified PKC, increased the level of GAP-43/B-50 phosphorylation in the striatum contralateral to the sense of turning [(+22%; p<0.05, with respect to ipsilateral side of the same trained group) and (+21%; p<0.05, relative to control group)]. Taken together, these results demonstrate that the activity developed in the CT test induces a reduction in the phosphorylation state of GAP-43/B-50 in the specific site for PKC. We conclude that general markers of activity-dependent neuronal plasticity are also altered in the same period that long-lasting changes in striatal neuroreceptors are triggered by circling motor behavior.     

Cavernous nerve injury elicits GAP-43 mRNA expression but not regeneration of injured pelvic ganglion neurons

Recovery of erectile dysfunction after cavernous nerve injury takes a long period. To elucidate this mechanism, unilateral cavernous nerve of male rat was cut, and the expression level of a nerve regeneration marker, the growth associated protein-43 (GAP-43) mRNA was evaluated by in situ hybridization and RT-PCR. While GAP-43 mRNA expression was transiently increased in the injured neurons of the major pelvic ganglion (MPG) at 7 days after nerve injury, continuous increase of GAP-43 mRNA was observed in the contralateral MPG from 7 days to 6 months after the nerve injury. Histochemical double-labeling studies for either neuronal NOS (nNOS) or tyrosine hydroxylase (TH) and the GAP-43 mRNA expression demonstrated that in injured MPG the transient up-regulation of GAP-43 mRNA was mainly seen in nNOS negative and/or TH positive neurons, suggesting non-parasympathetic post-ganglionic neurons, and also demonstrated that in contralateral MPG GAP-43 mRNA positive neurons were gradually increased in nNOS positive but TH negative neurons, suggesting parasympathetic post-ganglionic neurons. When a retrograde tracer Fluorogold (FG) was injected into the penile crus 7 days before histological experiments, FG-positive neurons were, if any, hardly seen in nNOS-positive neurons of the injured MPG for at least 6 months, whereas numerous FG-positive cells were seen in nNOS-positive neurons of the contralateral MPG. These results suggest that post-ganglionic projecting neurons of the intact side, which express increased GAP-43 mRNA, would be most likely to contribute to the recovery of the erectile function after unilateral cavernous nerve injury possibly by a plastic change such as nerve sprouting.     

Growth-associated protein 43 and neural cell adhesion molecule expression following bone marrow-derived mesenchymal stem cell transplantation in a rat model of ischemic brain injury

BACKGROUND:Transplantation of bone marrow-derived mesenchymal stem cells(BMSCs)improves motor functional recovery,but the mechanisms remain unclear.OBJECTIVE: To investigate expression of growth-associated protein 43(GAP-43)and neural cell adhesion molecule following BMSC transplantation to the lateral ventricle in rats with acute focal cerebral ischemic brain damage.DESIGN,TIME AND SETTING: A randomized,controlled,animal experiment using immunohistochemistry was performed at the laboratories of Department of Neurology,Renmin Hospital of Wuhan University and Doctoral Scientific Research Work Station of C-BONS PHARMA,Hubei Province,China,from January 2007 to December 2008.MATERIALS: Monoclonal mouse anti-rat 5-bromo-2-deoxyuridine and neural cell adhesion molecule antibodies were purchased from Sigma,USA;monoclonal mouse anti-rat GAP-43 antibody was purchased from Wuhan Boster,China.METHODS: Rat models of right middle cerebral artery occlusion were established using the thread method.At 1 day after middle cerebral artery occlusion,20 μL culture solution,containing 5 × 105 BMSCs,was transplanted to the left lateral ventricle using micro-injection.MAIN OUTCOME MEASURES: Scores of neurological impairment were measured to assess neural function.Expression of GAP-43 and neural cell adhesion molecule at the lesion areas was examined by immunohistechemistry.RESULTS: GAP-43 and neural cell adhesion molecule expression was low in brain tissues of the sham-operated group,but expression increased at the ischemic boundary(P < 0.05).Transplantation of BMSCs further enhanced expression of GAP-43 and neural cell adhesion molecule(P < 0.05)and remarkably improved neurological impairment of ischemic rats(P< 0.05).CONCLUSION: BMSC transplantation promoted neurological recovery in rats by upregulating expression of GAP-43 and neural cell adhesion molecule.     

Somatosensory corticospinal tract axons sprout within the cervical cord following a dorsal root/dorsal column spinal injury in the rat

The corticospinal tract (CST) is the major descending pathway controlling voluntary hand function in primates, and though less dominant, it mediates voluntary paw movements in rats. As with primates, the CST in rats originates from multiple (albeit fewer) cortical sites, and functionally different motor and somatosensory subcomponents terminate in different regions of the spinal gray matter. We recently reported in monkeys that following a combined cervical dorsal root/dorsal column lesion (DRL/DCL), both motor and S1 CSTs sprout well beyond their normal terminal range. The S1 CST sprouting response is particularly dramatic, indicating an important, if poorly understood, somatosensory role in the recovery process. As rats are used extensively to model spinal cord injury, we asked if the S1 CST response is conserved in rodents. Rats were divided into sham controls, and two groups surviving post-lesion for ~6 and 10 weeks. A DRL/DCL was made to partially deafferent one paw. Behavioral testing showed a post-lesion deficit and recovery over several weeks. Three weeks prior to ending the experiment, S1 cortex was mapped electrophysiologically, for tracer injection placement to determine S1 CST termination patterns within the cord. Synaptogenesis was also assessed for labeled S1 CST terminals within the dorsal horn. Our findings show that the affected S1 CST sprouts well beyond its normal range in response to a DRL/DCL, much as it does in macaque monkeys. This, along with evidence for increased synaptogenesis post-lesion, indicates that CST terminal sprouting following a central sensory lesion, is a robust and conserved response.     

Neonatal sciatic nerve lesion triggers the sprouting of fibers in the contralateral ventral root of the rat

We explored the possibility that unilateral neurectomy of the sciatic nerve of the rat at the neonatal stage triggers sprouting of afferent fibers in the contralateral ventral root. 3 months after neonatal sciatic neurectomy, the numbers of both myelinated and unmyelinated fibers in the L5 and L3 ventral roots were counted on electron micrographic montages. Age-matched littermates were used as unoperated controls. To identify regenerating axons, electron microscopic immunohistochemistry was done on the ventral roots using antibody against growth-associated phosphoprotein (GAP-43). Neonatal sciatic neurectomy resulted in: (1) about a three-fold increase in the number of unmyelinated fibers in the contralateral L5 ventral root as compared with the unoperated control; (2) about a 25-fold increase in the number of unmyelinated fibers in the ipsilateral L5 ventral root as compared with the control; (3) 25% of the unmyelinated fibers in the contralateral L5 ventral root expressing GAP-43; and (4) no significant change in the number of unmyelinated fibers in the L3 ventral root of either side as compared with the control. The data suggest that a neonatal sciatic neurectomy of the rat triggers sprouting of unmyelinated afferent fibers in the ventral root of the contralateral as well as the ipsilateral side. The sprouting is restricted, however, to spinal segments with receive inputs from the sciatic nerve.     

Altered protein markers related to neural plasticity and motor function following electro-acupuncture treatment in rat model of ischemic-hypoxic brain injury

BACKGROUND: Previous studies have demonstrated that acupuncture treatment could ameliorate impaired motor function, and these positive effects might be due to neural plasticity. OBJECTIVE: Myelin basic protein (MBP), microtubule-associated protein 2 (MAP2), growth-associated protein-43 (GAP-43), and synaptophysin (SYN) were selected as markers of neural remodeling, and expression of these markers was evaluated with regard to altered motor function following brain injury and acupuncture treatment. DESIGN, TIME AND SETTING: A completely randomized experiment was performed at the Central Laboratory of Peking University First Hospital from November 2006 to May 2007. MATERIALS: Twenty-four Sprague Dawley rat pups, aged 7 days, were selected for the present experiment. The left common carotid artery was ligated to establish a rat model of ischemic-hypoxic brain injury.METHODS: All animals were randomly divided into three groups: sham operation, model, and electro-acupuncture treatment, with 8 rats in each group. Rats in the model and electro-acupuncture treatment group underwent establishment of ischemic-hypoxic brain injury. Upon model established, rats underwent hypobaric oxygen intervention for 24 hours. Only the left common carotid artery was exposed in rats of the sham operation group, without model establishment or oxygen intervention. The rats in the electro-acupuncture treatment group were treated with electro-acupuncture. One acupuncture needle electrode was inserted into the subcutaneous layer at the Baihui and Dazhui acupoint. The stimulation condition of the electro-acupuncture simulator was set to an amplitude-modulated wave of 0-100% and alternative frequency of 100 cycles/second, as well as frequency-modulated wave of 2-100 Hz and an alternative frequency of 3 cycles/second. Maximal current through the two dectrodes was limited to 3-5 mA. The stimulation lasted for 30 minutes per day for 2 weeks. Rats in the sham operation and model groups were not treated with electro-acupuncture, but only fixed to the table for the same time period. MAIN OUTCOME MEASURES: After 2 weeks stimulation, expression of MBP, MAP2, GAP-43, and SYN were detected in the brain by immunohistochemistry. Motor function was evaluated in the three groups. RESULTS: In the sham operation group, MBP was abundant in the myelinated nerve fibers. In the electro-acupuncture treatment group, however, the corpus callosum exhibited more MBP staining than the model group. MAP2 expression was increased in the model group, and increased further in the electro-acupuncture treatment group compared with the sham operation group. GAP-43 expression in the cerebral cortex was less in model group than in sham operation, but present in abundance in the electro-acupuncture treatment group. SYN expression in the cerebral cortex was less in the model and electro-acupuncture treatment group compared with the sham operation group. There was no significant difference in SYN expression and distribution between the model and electro-acupuncture treatment groups. Motor function of rats in the electro-aeapuncture treatment group was significantly better than the model group (P < 0.05), although function remained lower than the sham operation group (P < 0.05). CONCLUSION: Two weeks of electro-acupuncture treatment improved motor function in rats, and protein markers related to neural plasticity also changed, which may be a mechanism for improved motor function in rats with ischemic-hypoxic brain injury.     

Bilateral corticospinal projections arise from each motor cortex in the macaque monkey: a quantitative study

The corticospinal projection is considered to influence fine motor function through nearly exclusively contralateral projections from the cortex in primates. However, unilateral lesions to this system in various species are frequently followed by significant functional improvement, raising the possibility that bilateral projections of this pathway may exist or emerge after injury. To examine the detailed anatomy and projections of the corticospinal motor neurons in rhesus monkeys (n = 4), we injected the high-resolution anterograde tracer biotinylated dextran amine (BDA) into 126 sites centered about the right lower extremity (LE) primary motor cortex. Projection and termination patterns were quantified at lumbar levels L1, L4, and L7 and mapped by using serial-section reconstructions. Notably, a mean of 10.1 +/- 0.6% (+/- SEM) of corticospinal tract (CST) axons descended in the lateral CST ipsilateral to the cortical BDA injection, and 87.9 +/- 1.0% of total CST axons projected in the contralateral lateral CST. The ipsilateral ventral CST contained only 1.0 +/- 0% of all projecting CST axons, whereas the contralateral ventral CST contained 0.3 +/- 0.2% of all axons. In addition, a minor dorsal column CST projection was identified. Measurement of BDA-labeled terminals in the spinal cord gray matter revealed that 11.2 +/- 2.2% of CST axons terminated ipsilateral to the side of cortical injection, and the remainder terminated contralaterally. As previously reported, most CST axons terminated in spinal cord laminae V-VIII, as well as the laterodorsal motoneuronal group of lamina IX (which innervates distal extremity muscles). Notably, many CST axons crossed the spinal cord midline (mean 19.9 +/- 4.9 axons per 40-microm-thick section). Detailed single-axon reconstructions revealed that most ipsilaterally projecting lateral CST axons terminated in ipsilateral gray matter. Notably, we found that the bouton-like swellings of many ipsilateral CST axons descending in the dorsolateral tract were located within Rexed's lamina IX, in close proximity to motoneuronal somata. Thus, bilateral projections of corticospinal axons originating from a single motor cortex could contribute to bilateral control of spinal motor neurons and to the highly evolved degree of fine motor control in primates. Furthermore, bilateral CST projections from a single motor cortex could represent a potential source of plasticity after injury, as well as a target of therapeutic effort in neural regeneration strategies.     

表皮生长因子对大鼠脑梗死后神经功能恢复的影响

目的 观察表皮生长因子（EGF）对大鼠脑梗死后神经功能恢复的影响。方法 采用肾血管性高血压大鼠制作一侧大脑中动脉皮层支闭塞（MCAO）模型。MCAO术后24 h，32只大鼠侧脑室注入10μl EGF（100μg/L），连续2d，共2μg（EGF组）；32只大鼠只注入不含EGF的等量溶液（对照组）。MCAO术后1、2、3和4w，行Bederson神经功能评分后，免疫印迹检测双侧大脑半球生长相关蛋白（GAP43）、突触囊泡蛋白（SYN）和胶质原纤维酸性蛋白（GFAP）的表达。结果 与同期对照组相比，EGF组大鼠在MCAO术后1、2w神经运动功能恢复更好，脑梗死灶同侧半球GAP43、SYN和GFAP有更早期和更高表达（P<0.05）。结论 GAP43、SYN和GFAP等蛋白的早期高峰表达可能与EGF引起的早期神经可塑性改善有关。     

Functional reorganization of the brain in recovery from striatocapsular infarction in man.

We used positron emission tomography (PET) to study organizational changes in the functional anatomy of the brain in 10 patients following recovery from striatocapsular motor strokes. Comparisons of regional cerebral blood flow maps at rest between the patients and 10 normal subjects revealed significantly lower regional cerebral blood flow in the basal ganglia, thalamus, sensorimotor, insular, and dorsolateral prefrontal cortices, in the brainstem, and in the ipsilateral cerebellum in patients, contralateral to the side of the recovered hand. These deficits reflect the distribution of dysfunction caused by the ischemic lesion. Regional cerebral blood flow was significantly increased in the contralateral posterior cingulate and premotor cortices, and in the caudate nucleus ipsilateral to the recovered hand. During the performance of a motor task by the recovered hand, patients activated the contralateral cortical motor areas and ipsilateral cerebellum to the same extent as did normal subjects. However, activation was greater than in normal subjects in both insulae; in the inferior parietal (area 40), prefrontal and anterior cingulate cortices; in the ipsilateral premotor cortex and basal ganglia; and in the contralateral cerebellum. The pattern of cortical activation was also abnormal when the unaffected hand, contralateral to the hemiplegia, performed the task. We showed that bilateral activation of motor pathways and the recruitment of additional sensorimotor areas and of other specific cortical areas are associated with recovery from motor stroke due to striatocapsular infarction. Activation of anterior and posterior cingulate and prefrontal cortices suggests that selective attentional and intentional mechanisms may be important in the recovery process. Our findings suggest that there is considerable scope for functional plasticity in the adult human cerebral cortex.     

Beneficial effects of gfap/vimentin reactive astrocytes for axonal remodeling and motor behavioral recovery in mice after stroke

The functional role of reactive astrocytes after stroke is controversial. To elucidate whether reactive astrocytes contribute to neurological recovery, we compared behavioral outcome, axonal remodeling of the corticospinal tract (CST), and the spatio‐temporal change of chondroitin sulfate proteoglycan (CSPG) expression between wild‐type (WT) and glial fibrillary acidic protein/vimentin double knockout (GFAP^–/–Vim^–/–) mice subjected to Rose Bengal induced cerebral cortical photothrombotic stroke in the right forelimb motor area. A foot‐fault test and a single pellet reaching test were performed prior to and on day 3 after stroke, and weekly thereafter to monitor functional deficit and recovery. Biotinylated dextran amine (BDA) was injected into the left motor cortex to anterogradely label the CST axons. Compared with WT mice, the motor functional recovery and BDA‐positive CST axonal length in the denervated side of the cervical gray matter were significantly reduced in GFAP^–/–Vim^–/– mice (n = 10/group, P < 0.01). Immunohistological data showed that in GFAP^–/–Vim^–/– mice, in which astrocytic reactivity is attenuated, CSPG expression was significantly increased in the lesion remote areas in both hemispheres, but decreased in the ischemic lesion boundary zone, compared with WT mice (n = 12/group, P < 0.001). Our data suggest that attenuated astrocytic reactivity impairs or delays neurological recovery by reducing CST axonal remodeling in the denervated spinal cord. Thus, manipulation of astrocytic reactivity post stroke may represent a therapeutic target for neurorestorative strategies. GLIA 2014;62:2022–2033     

Expression of GAP-43 in the Granule Cells of Rat Hippocampus After Seizure-induced Sprouting of Mossy Fibres: In Situ Hybridization and Immunocytochemical Studies

The axonal growth-associated protein GAP-43 is believed to play some role in the synaptic remodelling that takes place in the hippocampus of adult rats after certain experimental lesions. GAP-43 mRNA is highly expressed in adult CA3 pyramidal cells but almost absent in the dentate granule cells. We analysed whether the sprouting of granule cell axons, the mossy fibres of the hippocampus, caused by kainic acid-induced seizures in adult rats was associated with any induction of GAP-43 mRNA in granule cells and with any changes in the immunostaining pattern of GAP-43 in the hippocampus. Increased GAP-43 mRNA expression was found to be induced in granule cells 18, 24 and 30 h after a systemic injection of kainic acid which induced generalized seizures in adult rats, and returned to control levels by 48 h post-treatment. No effect was observed in other regions of the hippocampus. However, when kainic acid was injected into 15-day-old rats, which responded with generalized seizures but no sprouting of mossy fibres, there was no induction of GAP-43 mRNA in the granule cells, suggesting a close relation between GAP-43 expression and sprouting of these cells. Seven days after kainic acid injections, GAP-43 immunostaining was decreased in the inner molecular layer of the dentate gyrus except for a thin supragranular band, whereas 30 days after treatment all animals showed increased GAP-43 immunoreactivity in the whole inner molecular layer. Since collaterals of mossy fibres grow in the inner molecular layer after kainic acid-induced seizures, these results support the theory that GAP-43 plays a role in synaptic remodelling in the adult central nervous system.