High-frequency spinal cord stimulation produces longlasting analgesic effects by restoring lysosomal function and autophagic flux in the spinal dorsal horn

High-frequency spinal cord stimulation(HF-SCS) has been established as an effective therapy for neuropathic pain. However, the analgesic mechanisms involved in HF-SCS remain to be clarified. In our study, adult rat neuropathic pain was induced by spinal nerve ligation. Two days after modeling, the rats were subjected to 4 hours of HF-SCS(motor threshold 50%, frequency 10,000 Hz, and pulse width 0.024 ms) in the dorsal horn of the spinal cord. The results revealed that the tactile allodynia of spinal nerve-injured rats was markedly alleviated by HFSCS, and the effects were sustained for 3 hours after the stimulation had ceased. HF-SCS restored lysosomal function, increased the levels of lysosome-associated membrane protein 2(LAMP2) and the mature form of cathepsin D(matu-CTSD), and alleviated the abnormally elevated levels of microtubule-associated protein 1 A/B-light chain 3(LC3)-II and sequestosome 1(P62) in spinal nerve-injured rats. HF-SCS also mostly restored the immunoreactivity of LAMP2, which was localized in neurons in the superficial layers of the spinal dorsal horn in spinal nerve-injured rats. In addition, intraperitoneal administration of 15 mg/kg chloroquine for 60 minutes reversed the expression of the aforementioned proteins and shortened the timing of the analgesic effects of HF-SCS. These findings suggest that HF-SCS may exhibit longlasting analgesic effects on neuropathic pain in rats through improving lysosomal dysfunction and alleviating autophagic flux. This study was approved by the Laboratory Animal Ethics Committee of China Medical University, Shenyang, China(approval No. 2017 PS196 K) on March 1, 2017.

Pulsed electrical stimulation protects neurons in the dorsal root and anterior horn of the spinal cord after peripheral nerve injury

Most studies on peripheral nerve injury have focused on repair at the site of injury, but very few have examined the effects of repair strategies on the more proximal neuronal cell bodies. In this study, an approximately 10-mm-long nerve segment from the ischial tuberosity in the rat was transected and its proximal and distal ends were inverted and sutured. The spinal cord was subjected to pulsed electrical stimulation at T10 and L3, at a current of 6.5 m A and a stimulation frequency of 15 Hz, 15 minutes per session, twice a day for 56 days. After pulsed electrical stimulation, the number of neurons in the dorsal root ganglion and anterior horn was increased in rats with sciatic nerve injury. The number of myelinated nerve fibers was increased in the sciatic nerve. The ultrastructure of neurons in the dorsal root ganglion and spinal cord was noticeably improved. Conduction velocity of the sciatic nerve was also increased. These results show that pulsed electrical stimulation protects sensory neurons in the dorsal root ganglia as well as motor neurons in the anterior horn of the spinal cord after peripheral nerve injury, and that it promotes the regeneration of peripheral nerve fibers.

“Three Methods and Three Points” regulates p38 mitogen-activated protein kinase in the dorsal horn of the spinal cord in a rat model of sciatic nerve injury

Tuina is a traditional Chinese treatment for sensory disturbances caused by peripheral nerve injury and related diseases. Our previous studies showed that tuina regulates relevant regions and indices of the spinal dorsal horn using the Dian, Bo, and Rou method in Yinmen（BL37）, Yanglingquan（GB34）, and Weizhong（BL40）. Treatment prevents muscle atrophy, protects spinal cord neurons, and promotes sciatic nerve repair. The mechanisms of action of tuina for treating peripheral nerve injury remain poorly understood. This study established rat models of sciatic nerve injury using the crushing method. Rats received Chinese tuina in accordance with the principle of ＂Three Methods and Three Points,＂ once daily for 20 days. Tuina intervention reduced paw withdrawal latency and improved wet weight of the gastrocnemius muscle, as well as promoting morphological recovery of sciatic nerve fibers, Schwann cells, and axons. The protein expression levels of phospho-p38 mitogen-activated protein kinase, tumor necrosis factor-α, and interleukin-1β also decreased. These findings indicate that ＂Three Methods and Three Points＂ promoted morphological recovery and improved behavior of rats with peripheral nerve injury.

Effect of propofol on glutamate-induced activation and related inflammatory cytokines of astrocytes from spinal cord dorsal horn

BACKGROUND： Astrocytes participate in central nervous system-mediated physiological or pathological processes, such as pain. Activated dorsal horn astrocytes from the spinal cord produce nerve active substances and proinflammatory cytokines, such as interleukin-lbeta （IL-1 β ）, IL-6, and tumor necrosis factor- α （TNF-α ）, which play important roles in pain transduction and regulation. OBJECTIVE： To investigate the effects of different doses of propofol on activation of cultured spinal cord dorsal horn astrocytes induced by glutamate, as well as changes in IL-1β, IL-6, and TNF- α, and 1L-10 （anti-inflammatory cytokine） expression in rats, and to explore the dose relationship of propofol. DESIGN, TIME AND SETTING： The cellular and molecular biology experiment was performed at the Central Laboratory of Yunyang Medical College between March 2006 and December 2007. MATERIALS： Forty healthy, Wistar rats, aged 2-3 days, were selected. Propofol was provided by Zeneca, UK; glutamate by Sigma, USA; EPICS XL flow cytometry by Beckman culture, USA; rabbit-anti-mouse glial fibrillary acidic protein （GFAP） antibody kit and inflammatory cytokine detection kit were provided by Zhongshan Biotechnology Company Ltd., Beijing; multimedia color pathologic image analysis system was a product of Nikon, Japan. METHODS： Astrocytes were harvested from T11- L6 spinal cord dorsal horn of Wistar rats and incubated for 3 weeks. The cells were divided into seven groups, according to various treatment conditions： control group was cells cultured in Hank＇s buffered saline solution; intralipid group was cells cultured in intralipid （0.2 mL/L）; glutamate group was cells cultured with 100 u mol/L glutamate; propofol group was cells cultured with 250 u mol/L propofol; three glutamate plus propofol groups were cultured in 100 11 mol/L of glutamate, followed by 5, 25, and 250 u mol/L of propofol 10 minutes later. MAIN OUTCOME MEASURES： GFAP-labeled astrocytes were analyzed using a multimedia pathology imaging analysis system to detect area density （AD） and average optical density （AOD） of positive cells. The supernatant concentrations of IL-1β, TNF- α, IL-6, and IL-10 were determined using radioimmune assays. RESULTS： Compared with the control group, cells in the glutamate plus low-dose propofol group were activated and hypertrophic, and AD and AOD were significantly increased （P 〈 0.01 ）. Concentrations of IL-1β, TNF- α, and IL-6 were also significantly increased （P 〈 0.01）, while IL-10 levels remained unchanged （P 〉 0.05）, but still higher than the control and glutamate groups （P 〉 0.05）. Compared with the glutamate group, astrocyte activation was inhibited by moderate and high-dose propotol. In addition, with moderate and high-dose propofol, AD, AOD, IL-1β, TNF- α, and IL-6 concentrations were significantly decreased （P 〈 0.05-0.01）, and IL-10 levels were increased （P 〈 0.01 ）. CONCLUSION： Propofol can effectively inhibit glutamate-induced astrocyte activation in the spinal cord dorsal horn, significantly inhibit production of IL-1 β, TNF- α, and IL-6, and increase IL-10 synthesis and release in a dose-dependent manner.

Transcriptional control of GABAergic neuron development in the dorsal spinal cord

GABAergic neurons are the major inhibitory interneurons that powerfully control the function of spinal neuronal networks.In recent years,tremendous progresses have been made in understanding the transcriptional control of GABAergic neuron development in the dorsal spinal cord.New experimental approaches provide a relatively high throughput way to study the molecular regulation of subgroup fate determination.Our understanding of the molecular mechanisms on GABAergic neuron development in the dorsal spinal cord is rapidly expanding.Recent studies have defined several transcription factors that play essential roles in GABAergic neuron development in the spinal dorsal horn.Here,we review results of very recent analyses of the mechanisms that specify the GABAergic neuron development in the dorsal spinal cord,especially the progresses in the homeodomain(HD) and basic-helix-loop-helix(bHLH) containing transcription factors.

Effects of intrathecal injection of glial cell inhibitor on spinal cord astrocytes following chronic compression of dorsal root ganglia in rats

BACKGROUND： Astrocytes are considered to provide nutritional support in the central nervous system. However, recent studies have confirmed that astrocytes also play an important role in chronic pain. OBJECTIVE： To investigate the effects of intrathecal injection of fluorocitrate, minocycline or both on astrocyte activation and proliferation in the spinal dorsal horn of compressed dorsal root ganglion in rats. DESIGN, TIME AND SETTING： The neurology randomized controlled animal study was performed at the Jiangsu Institute of Anesthesia Medicine, from September 2006 to April 2007. MATERIALS： A total of 96 male Sprague Dawley rats, aged 6-8 weeks, were selected for this study. Following intrathecal catheterization, 80 rats underwent steel bar insertion into the L4-5 intervertebral foramina to make a stable compression on the L4-5 posterior root ganglion. Thus rat models of ganglion compression were established. Minocycline and fluorocitrate were purchased from Sigma, USA. METHODS： A total of 96 rats were randomly and equally divided into six groups. Rat L4, L5 transverse process and intervertebral foramina were exposed in the sham operation group, but without steel bar insertion. The model group did not receive any manipulations. Rats in the phosphate buffered saline （PBS） group were intrathecally injected with 0.01 mmol/L PBS （20 μL）. Rats in the fluorocitrate group were subjected to 1 μmol/L fluorocitrate （20 μL）. Rats in the minocycline group were intrathecally injected with 5 g/L minocycline （20 μL）. Rats in the minocycline and fluorocitrate group received a mixture （20 μL） of 5 g/L minocycline and 1 μmol/L fluorocitrate. Following model establishment, drugs were administered once a day. MAIN OUTCOME MEASURES： At 7 and 14 days following model induction, glial fibrillary acidic protein expression in the spinal dorsal horn was measured by immunofluorescence microscopy. Six sections with significant glial fibrillary acidic protein -positive expression were obtained to count astrocytes under an inverted microscope. RESULTS： No significant differences in astrocyte count were detected between the fluorocitrate and model groups. Cell bodies were small with a few processes in the fluorocitrate group, compared with the model group. The astrocyte count decreased significantly in the minocycline group and the minocycline and fluorocitrate group compared with the sham operation, model, PBS and fluorocitrate groups （P 〈 0.01）. The decrease in astrocyte count was mainly found in layers Ⅲ–Ⅳ of the spinal dorsal horn. Cell body volume was smaller and process numbers were fewer in the minocycline group and the minocycline and fluorocitrate group, compared with the model and PBS groups. CONCLUSION： Fluorocitrate can inhibit astrocyte activation, but does not affect astrocyte proliferation. However, minocycline can inhibit the activation and proliferation of astrocytes.

Up-regulation and time course of protein kinase C immunoreactivity during persistent inflammation of the rat spinal cord

BACKGROUND： It has been reported that activation and/or translocation of protein kinase C （PKC） is related to hyperalgesia, and changes in PKC expression in the dorsal horn of spinal cord take place during inflammatory pain. OBJECTIVE： To observe PKC changes in the dorsal horn of spinal cord using immunohistochemistry and to measure the time-course during persistent pain produced by chemical stimulation with a right hind-paw injection of formalin. DESIGN： Randomized controlled animal experiment. SETTING： Institute of Basic Medical Science, Hebei Medical University MATERIALS： The present experiment was performed at the Department of Pathophysiology, Institute of Basic Medical Science, Hebei Medical University between September 2000 and June 2002. Forty-two Sprague-Dawley rats, weighing 260-280 g, irrespective of gender, were provided by the Center of Animal Experimentation at Hebei Medical University. PKC antibody was provided by Sigma, USA. Immunohistochemistry kits were purchased from Zhongshan Biotechnology Company, Beijing. HPIAS-1000 definition multicolor system was provided by Qianping Wuxiang Project Company of Tongji Medical University. Animal use during experimentation was consistent with the standards of Animal Ethics Committee. METHODS： Sprague-Dawley rats were divided randomly into control （n = 6） and experimental groups （n = 36）. Experimental rats were given an intracutaneous injection of 5% formalin into the planta surface of the right hind-paw. Animals with inflammatory pain were anesthetized and sacrificed to obtain the L5 spinal region at 1, 3, 12 hours, 1, 3, and 7 days after formalin treatment, with 6 rats in each time group. The spinal cords at the L5 region were collected from the control group following sodium chloride injections into the planta surface of the right hind-paw, identical to the experimental group. MAIN OUTCOME MEASURES： Pain reaction of experimental rats after formalin treatment. PKC-positive neurons, and distribution of PKC-immunoreactive particles, in the ipsi- and contralateral dorsal horn were investigated during different stages of inflammatory pain using immunohistochemistry. RESULTS： All 42 rats were included in the final analysis, without any loss. Pain reaction： consistent with previous findings, it was determined that a unilateral injection of formalin into the hind-paw resulted in significant edema and induced a series of nociceptive responses, such as licking, biting, or shaking the injected paw. The maximal inflammation change was observed 1 day after formalin injection and changes did not disappear until the day 7. Number of the PKC positive neurons： results demonstrated that the number of PKC immunoreactive neurons in the dorsal horn increased slightly after formalin injection at 1 hour, compared with the control group. PKC immunoreactivity was up-regulated at day 1, reduced at day 3, and appeared to recover at day 7. The number of PKC-positive neurons in the contralateral side was less than the ipsilateral side at each time sampled. Distribution of PKC immunoparticles over the neurons： PKC immunoreactivity was observed in the nucleus and cytoplasm, as well as on or near the membrane of neurons and synaptosomes in the spinal cord of the control group. PKC activated and translocated from nucleus to the membrane-associated site following formalin treatment. Significant changes were observed at 1 hour and 1 day. The intensity of staining was stronger in the ipsilateral side than the contralateral side at all time points following formalin injection （P 〈 0.01）, whereas the expression patterns of PKC immunoreactivity in the nuclei were very similar in the right and left hemispheres. CONCLUSION： PKC expression in the dorsal horn of the spinal cord peaked at 1 hour and 24 hours, and was very obvious at 24 hours. Protein kinase C expression in the spinal cord increased bilaterally, although it was greater in the ipsilateral hemisphere. In addition, PKC expression at the neuronal membrane and synaptosome was significantly increased. These results indicate that PKC expression is activated in the dorsal horn of the spinal cord during hyperalgesia.

Spinal processing of bee venom-induced pain and hyperalgesia

Subcutaneous injection of bee venom causes long-term neural activation and hypersensitization in the dorsal horn of the spinal cord,which contributes to the development and maintenance of various pain-related behaviors.The unique behavioral 'phenotypes' of nociception and hypersensitivity identified in the rodent bee venom test are believed to reflect a complex pathological state of inflammatory pain and might be appropriate to the study of phenotype-based mechanisms of pain and hyperalgesia.In this review,the spinal processing of the bee venom-induced different 'phenotypes' of pain and hyperalgesia will be described.The accumulative electrophysiological,pharmacological,and behavioral data strongly suggest that different 'phenotypes' of pain and hyperalgesia are mediated by different spinal signaling pathways.Unraveling the phenotype-based mechanisms of pain might be useful in development of novel therapeutic drugs against complex clinic pathological pain.

Analgesic effect of intrathecal bumetanide is accompanied by changes in spinal sodium-potassium-chloride co-transporter 1 and potassium-chloride co-transporter 2 expression in a rat model of incisional pain

Accumulating evidence has demonstrated that the sodium-potassium-chloride co-transporter 1 and potassium-chloride co-transporter 2 have a role in the modulation of pain transmission at the spinal level through chloride regulation in the pain pathway and by effecting neuronal excitability and pain sensitization. The present study aimed to investigate the analgesic effect of the speciifc sodium-potassium-chloride co-transporter 1 inhibitor bumetanide, and the change in spinal sodium-potassium-chloride co-transporter 1 and potassium-chloride co-transporter 2 expression in a rat model of incisional pain. Results showed that intrathecal bumetanide could decrease cumulative pain scores, and could increase thermal and mechanical pain thresholds in a rat model of incisional pain. Sodium-potassium-chloride co-transporter 1 expression in-creased in neurons from dorsal root ganglion and the deep laminae of the ipsilateral dorsal horn following incision. By contrast, potassium-chloride co-transporter 2 expression decreased in neurons of the deep laminae from the ipsilateral dorsal horn. These ifndings suggest that spinal sodium-potassium-chloride co-transporter 1 expression was up-regulated and spinal potassi-um-chloride co-transporter 2 expression was down-regulated following incision. Intrathecal bumetanide has analgesic effects on incisional pain through inhibition of sodium-potassi-um-chloride co-transporter 1.

Estrogen affects neuropathic pain through upregulating N-methyl-D-aspartate acid receptor 1 expression in the dorsal root ganglion of rats

Estrogen affects the generation and transmission of neuropathic pain,but the specific regulatory mechanism is still unclear.Activation of the N-methyl-D-aspartate acid receptor 1（NMDAR1） plays an important role in the production and maintenance of hyperalgesia and allodynia.The present study was conducted to determine whether a relationship exists between estrogen and NMDAR1 in peripheral nerve pain.A chronic sciatic nerve constriction injury model of chronic neuropathic pain was established in rats.These rats were then subcutaneously injected with 17β-estradiol,the NMDAR1 antagonist D（-）-2-amino-5-phosphonopentanoic acid（AP-5）,or both once daily for 15 days.Compared with injured drug na？ve rats,rats with chronic sciatic nerve injury that were administered estradiol showed a lower paw withdrawal mechanical threshold and a shorter paw withdrawal thermal latency,indicating increased sensitivity to mechanical and thermal pain.Estrogen administration was also associated with increased expression of NMDAR1 immunoreactivity（as assessed by immunohistochemistry） and protein（as determined by western blot assay） in spinal dorsal root ganglia.This 17β-estradiol-induced increase in NMDAR1 expression was blocked by co-administration with AP-5,whereas AP-5 alone did not affect NMDAR1 expression.These results suggest that 17β-estradiol administration significantly reduced mechanical and thermal pain thresholds in rats with chronic constriction of the sciatic nerve,and that the mechanism for this increased sensitivity may be related to the upregulation of NMDAR1 expression in dorsal root ganglia.

The hypothalamic-spinal dopaminergic system:a target for pain modulation

Nociceptive signals conveyed to the dorsal horn of the spinal cord by primary nociceptors are subject to extensive modulation by local neurons and by supraspinal descending pathways to the spinal cord before being relayed to higher brain centers. Descending modulatory pathways to the spinal cord comprise,among others, noradrenergic, serotonergic, γ-aminobutyric acid(GABA)ergic, and dopaminergic fibers.The contributions of noradrenaline, serotonin, and GABA to pain modulation have been extensively investigated. In contrast, the contributions of dopamine to pain modulation remain poorly understood.The focus of this review is to summarize the current knowledge of the contributions of dopamine to pain modulation. Hypothalamic A11 dopaminergic neurons project to all levels of the spinal cord and provide the main source of spinal dopamine. Dopamine receptors are expressed in primary nociceptors as well as in spinal neurons located in different laminae in the dorsal horn of the spinal cord, suggesting that dopamine can modulate pain signals by acting at both presynaptic and postsynaptic targets. Here, I will review the literature on the effects of dopamine and dopamine receptor agonists/antagonists on the excitability of primary nociceptors, the effects of dopamine on the synaptic transmission between primary nociceptors and dorsal horn neurons, and the effects of dopamine on pain in rodents. Published data support both anti-nociceptive effects of dopamine mediated by D2-like receptors and pro-nociceptive effects mediated by D1-like receptors.

电针对紫杉醇诱导大鼠神经病理性疼痛的影响

目的观察电针(EA)对紫杉醇诱导神经病理性疼痛大鼠脊髓背角NKCC1、KCC2表达和小胶质细胞活化的影响及其可能机制。方法将48只雄性SD大鼠随机分为溶媒组(Vehicle组)、紫杉醇组(PTX组)、紫杉醇+电针组(PTX+EA组)、紫杉醇+假电针组(PTX+Sham EA组),每组12只。采用腹腔注射PTX的方法建立PTX诱导的神经病理性疼痛大鼠动物模型,建模完成后,PTX+EA组给予“足三里”、“阳陵泉”电针刺激,连续7 d。于紫杉醇注射前2 d和注射后第1、3、5、7、14、21天进行机械撤足阈值和热缩足潜伏期痛行为学测试。利用免疫荧光染色技术和Western blot技术检测脊髓背角组织中钠钾氯联合转运蛋白1(NKCC1)、钾氯联合转运蛋白2(KCC2)和小胶质细胞标志物离子钙结合衔接分子1(Iba1)表达的变化。结果与Vehicle组比较,PTX组大鼠出现双后足机械和热痛觉过敏,脊髓背角组织中NKCC1表达升高和活化的小胶质细胞数增加。与PTX组比较,PTX+EA组大鼠在第14、21天的机械和热痛觉过敏得到显著改善,脊髓背角组织中NKCC1和Iba1表达量降低,4组间KCC2表达差异无统计学意义。结论电针可有效缓解紫杉醇诱导的神经病理性疼痛,其机制可能与抑制大鼠脊髓背角组织中NKCC1表达和小胶质细胞活化有关。

Increased phosphorylation of cyclic AMP response element binding protein(CREB)in the dorsal root ganglia and superficial dorsal horn neurons following chronic constriction injury

Objective To investigate whether chronic constriction injury(CCI)of the sciatic nerve of rats could produce alterations in the phosphorylation of cyclic AMP response element binding(CREB)protein in dorsal root ganglia(DRG)and superficial dorsal horn neurons of the spinal cord.Methods Chronic constriction injury(CCI)of the sciatic nerve was employed as a model of neuropathic pain.Thirty-two Sprague-Dawley rats were randomly divided into Na⒍ve,Sham,CCI2w(received CCI for2weeks)and CCI4w(received CCI for4weeks)groups.Hind pawwithdrawal threshold to mechanical stimuli and withdrawal latency to thermal stimuli were used to determine the mechanical and thermal hyperalgesia.Then all the rats were deeply anesthetized and perfused intracardially with paraformaldehyde.The fixed L 4-5 spinal cord and the L 5 DRG ipsilateral to CCI were harvested for fixation.The pCREB-immunoreactive(pCREB-IR)cells in both DRG and superficial dorsal horn neurons were quantified for analysis using immunohistochemistry methods.Results On the14th day after sciatic nerve injury,all the rats exhibited significant mechanical and thermal hyperalgesia.The mechanical withdrawal thresholds to von Frey filament from CCI2w group decreased significantly compared to both baseline values and those of Sham group(P<0.01);Thermal withdwal latencies from CCI2w group decreased significantly compared to both baseline values and those of Sham group(P<0.01).Some rats from Sham group also showed mechanical hyperalgesia compared to both baseline values and those of Na⒍ve group(P<0.01).28days after CCI,both mechanical and thermal hypersensitivity were significantly alleviated,with no statistical significance compared to those of Sham group.On the14th day after CCI,the number of pCREB-IR cells significantly increased in ipsilateral L 5 DRGs and superficial dorsal horns(P<0.01)compared to Sham group.The number of phosphorylated CREB-IR cells in the ipsilateral DRGs from Sham group also increased compared to that of Naive rats(P<0.05).There were no significant statistical differences of numbers of CREB-IR neuron between Sham group and CCI4wgroup.Conclusion CCI increases CREB phosphorylation both in DRG and superficial dorsal horn neurons of the lumbar spinal cord,and may be one of the key molecular mechanisms of central and peripheral sensitization following peripheral nerve injury.

补体系统参与病理性疼痛的机制研究进展

慢性病理性疼痛是由于躯体感觉系统或者组织损伤而导致的疼痛,在临床上很常见,给患者的身心健康带来很大危害。其治疗方法涵盖了药物治疗、经皮神经电刺激、运动及心理治疗以及介入治疗等方法。然而,这些治疗方法也存在一些缺陷,如疗效欠佳、药物耐受、患者接受度不高等。

Chinese Tuina remodels the synaptic structure in neuropathic pain rats by downregulating the expression of N-methyl D-aspartate receptor subtype 2B and postsynaptic density protein-95 in the spinal cord dorsal horn

OBJECTIVE:To investigate whether the Chinese massage system,Tuina,exerts analgesic effects in a rat model of chronic constriction injury(CCI)by remodeling the synaptic structure in the spinal cord dorsal horn(SCDH).METHODS:Sixty-nine male Sprague–Dawley rats were randomly and evenly divided into the normal group,sham group,CCI group,CCI+Tuina group,CCI+MK-801[an N-methyl D-aspartate receptor subtype 2B(NR2B)antagonist]group,and CCI+MK-801+Tuina group.The neuropathic pain model was established using CCI with right sciatic nerve ligation.Tuina was administered 4 d after CCI surgery,using pressing manipulation for 10 min,once daily.Motor function was observed with the inclined plate test,and pain behaviors were observed by the Von Frey test and acetone spray test.At 19 d after surgery,the L3-L5 spinal cord segments were removed.Glutamate,interleukin 1β(IL-1β),and tumor necrosis factor-α(TNF-α)levels were detected by enzyme-linked immunosorbent assay.The protein expression levels of NR2B and postsynaptic density protein-95(PSD-95)were detected by Western blot,and the synaptic structure was observed by transmission electron microscopy(TEM).RESULTS:CCI reduced motor function and caused mechanical and cold allodynia in rats,increased glutamate concentration and TNF-αand IL-1βlevels,and increased expression of synapse-related proteins NR2B and PSD-95 in the SCDH.TEM revealed that the synaptic structure of SCDH neurons was altered.Most of these disease-induced changes were reversed by Tuina and intrathecal injection of MK-801(P<0.05 or<0.01).For the majority of experiments,no significant differences were found between the CCI+MK-801 and CCI+MK-801+Tuina groups.CONCLUSIONS:Chinese Tuina can alleviate pain by remodeling the synaptic structure,and NR2B and PSD-95 receptors in the SCDH may be among its targets.

Microsurgical dorsal root entry zone coagulation for chronic neuropathic pain due to spinal cord and/or cauda equina injuries

Chronic pain is a major problem for patients suffering from spinal cord injury （SCI）.The prevalence of SCI-related pain ranges from 48% to 94% depending on the method of data acquisition and type of pain considered.1 SCI-related neuropathic pain is classified into three subtypes：above-,at-,and below-lesion-level pain.After SCI,it is believed that the dorsal horn of the injured spinal cord expressed increased electrical activity.2 Thus,the neurosurgical procedures for coagulation in the dorsal root entry zone （DREZ） have proved to be most effective in chronic neuropathic pain due to SCI.The aim of this study was to describe the effect and safety ofmicrosurgical DREZ coagulation in treating chronic SCI-related neuropathic pain.

THE PROPERTY OF PERIPHERAL INPUT OF PROJECTION NEURONS IN THE SPINAL CORD DORSAL HORN

Antidromic and orthodromic responses of the projection neurons in the dorsal horn ofthe spinal cord have been recorded by a glass microelectrode in anesthetized and paralyz-ed cats. Furthermore, the effect of cervical segment antidromic stimulation to orthodromicresponse of the projection neurons has been observed by way of conditioning-test stimulation. Among all the spinocervical tract neurons (SCT), the dorsal column postsynaptic neu-rons (DCPS) and the spinocervical tract-dorsal column postsynaptic neurons (SCT-DCPS),which were identified by cervical segment antidromic stimulation, 46% are low-thresholdmechanoreceptive (LTM) and 54% are wide-dynamic-range (WDR) neurons. Most LTMneurons can evoke the same response to both 10 times (10 T) and 50 times (50 T) the thresh-old stimulation on the peroneal nerve. Most WDR neurons to 50 T intensity stimulation arestronger than the 10 T stimulation. Under the antidromic-cervical segment conditioningstimulation, the amount of orthodromic-discharging in most WDR and few LTM neuronsreduced significantly. The result shows that both LTM and WDR projection neurons in the spinal cord canrespond to all peripheral Aβ fibers and part of the Aδ fibers; there are some inhibitionaldescending fibers which affect the projection neurons in the cervical segment dorsal col-umn and dorsolateral funiculi.

瑞舒伐他汀部分恢复吗啡耐受大鼠的吗啡镇痛效能

目的探讨瑞舒伐他汀对吗啡耐受大鼠的吗啡镇痛效能的影响及其相关的分子机制。方法 48只♂SD大鼠随机分成6组(n=8):Ⅰ组为空白对照组;Ⅱ组为吗啡耐受组;Ⅲ组为10 mg.kg-1瑞舒伐他汀对照组;Ⅳ组、Ⅴ组、Ⅵ组分别为0.4、2、10 mg.kg-1瑞舒伐他汀处理组。Ⅱ、Ⅳ、Ⅴ、Ⅵ组皮下注射吗啡10 mg.kg-1,Ⅰ组和Ⅲ组皮下注射生理盐水,每天8∶00和16∶00各1次,连续10 d。d 6起,上午皮下注射前30 min,Ⅰ、Ⅱ组给予生理盐水灌胃,Ⅲ、Ⅳ、Ⅴ、Ⅵ组给予相应剂量的瑞舒伐他汀灌胃,连续5 d。d 6、11,测定大鼠基础热缩足潜伏期(PWTL)后,计算尾静脉注射吗啡30 min时各组大鼠的MPE值。d 11行为学检测后处死大鼠,留取腰5脊髓,比较各组脊髓内细胞外调节蛋白激酶(ERK)、磷酸化细胞外调节蛋白激酶(p-ERK)及星形胶质细胞表面标志物GFAP的表达水平。结果①d 6,6组的基础PWTL无明显差异。与Ⅰ组相比,Ⅱ、Ⅳ、Ⅴ和Ⅵ组MPE值明显降低(P<0.05)。d 11,6组的基础PWTL无差异。与Ⅰ组相比,Ⅱ、Ⅳ组尾静脉注射吗啡后30 min的MPE值明显降低(P<0.05);与Ⅱ组相比,Ⅴ、Ⅵ组的MPE值明显升高(P<0.05)。②d 11,6组大鼠腰5脊髓内总ERK表达水平差异无显著性。与Ⅰ组相比,Ⅱ、Ⅳ组腰5脊髓内p-ERK表达明显升高(P<0.05);与Ⅱ组相比,Ⅲ、Ⅴ、Ⅵ组腰5脊髓内p-ERK表达明显降低(P<0.05)。与Ⅰ组相比,Ⅱ组腰5脊髓内GFAP的荧光强度明显升高(P<0.05);与Ⅱ组相比,Ⅵ组腰5脊髓内GFAP的强度明显降低(P<0.05)。结论瑞舒伐他汀能够部分恢复吗啡耐受大鼠的吗啡镇痛效果,这可能与其抑制脊髓内ERK的磷酸化,减少星形胶质细胞活化有关。

磷酸化钙/钙调蛋白依赖性蛋白激酶Ⅱ在大鼠脊髓背角C-纤维诱发电位长时程增强的诱导和维持中的作用

实验旨在探讨钙/钙调蛋白依赖性蛋白激酶Ⅱ(calcium/calmodulin—dependent protein kinase Ⅱ,CaMKⅡ)在脊髓背角C-纤维诱发电位长时程增强(long-term potentiation,LTP)的诱导和维持中的作用。用Western blot技术分别检测LTP形成30 min和3 h脊髓背角(L4-L6)CaMK Ⅱ的含量及其磷酸化水平。同时观察脊髓局部给予CaMK Ⅱ选择性抑制剂KN-93后对脊髓背角LTP和CaMK Ⅱ磷酸化的影响。观察结果如下:(1)诱导LTP后30 min.CaMK Ⅱ的磷酸化水平明显高于对照组,而CaMKⅡ的总量无变化;诱导LTP后3 h CaMK Ⅱ的磷酸化水平进一步升高,而且CaMK Ⅱ的总量也明显增加(n=4);(2)强直刺激前30 min 于脊髓局部给予CaMK Ⅱ的特异性抑制剂KN-93(100μmol/L),可阻断LTP的诱导,同时明显抑制CaMK Ⅱ的磷酸化水平;(3)诱导LTP后30 min给予KN-93,可显著抑制LTP的维持,同时CaMKⅡ的磷酸化水平与未用药组相比也明显降低(n=3);(4)LTP 3 h后给予KN-93,LTP的幅值不受影响,磷酸化的CaMKⅡ的含量与用药前相比也无差别(n=3)。根据上述实验结果可以认为,CaMK Ⅱ的激活参与脊髓背角C-纤维诱发电位LTP的诱导和早期维持过程。

大鼠坐骨神经压榨损伤后早期降钙素基因相关肽的变化

目的:研究大鼠坐骨神经压榨损伤后早期降钙素基因相关肽(CGRP)的动态变化及与神经再生的关系。方法:SD大鼠坐骨神经压榨损伤后分别存活1d到21d,免疫组化技术观察CGRP分布和含量的变化。结果:(1)1d组神经CGRP大量堆积,压榨近端明显多于远端,随即下降,21d组基本消失。(2)1d组背根节、脊髓后角和前角CGRP开始增高,并分别在3～5d、5～7d和7d组达峰值,随后渐降,21d组脊髓前角CGRP阳性运动神经元仍明显高于假手术组和对照侧。结论:神经压榨损伤后CGRP表达变化呈明显的时空模式,可能参与了神经元保护并介导了损伤信号的传导。