福尔马林致大鼠膀胱痛牵涉区的观测

目的 :研究大鼠膀胱致痛后牵涉区出现的所在部位 ,为探讨经牵涉区治疗内脏痛和阐明牵涉痛发生的机理提供实验和理论基础。方法 :SD大鼠 ,颈内静脉注入伊文思蓝染料 ,观察、比较动物在福尔马林致膀胱痛后该染料渗漏出现的部位。结果 :福尔马林致膀胱痛的动物伊文思蓝染料渗漏斑较恒定出现在耻骨联合腹侧局部皮区。结论 :根据牵涉痛发生机制的交感反射假说 ,初步认为大鼠膀胱痛所致牵涉区在耻骨联合腹侧皮肤。     

福尔马林致大鼠膀胱痛牵涉区的观测

目的：研究大鼠膀胱致痛后牵涉区出现的所在部位，为探讨经牵涉区治疗内脏痛和阐明牵涉痛发生的机理提供实验和理论基础。方法：SD大鼠，颈内静脉注入伊文思蓝染料，观察、比较动物在福尔马林致膀胱痛后该染料渗漏出现的部位。结果：福尔马林致膀胱痛的动物伊文思蓝染料渗漏斑较恒定出现在耻骨联合腹侧局部皮区。结论：根据牵涉痛发生机制的交感反射假说，初步认为大鼠膀胱痛所致牵涉区在耻骨联合腹侧皮肤。     

肩胛间区低频电刺激对家兔胆囊痛及其脊髓后角pERK1/2表达的影响

目的:探讨肩胛间区低频电刺激对家兔胆囊痛及脊髓后角pERK1/2表达的影响.方法:家兔随机分为5组:对照组、生理盐水组、甲醛溶液组、肩胛间区电刺激组和非肩胛间区电刺激组,每组6只.采用2Hz、3mA电流,应用生理功能学方法、免疫组织化学方法和Western blot技术,观察肩胛间区电刺激对胆囊痛模型呼吸频率和心率的改变及脊髓后角pERK1/2表达的影响.结果:肩胛间区电刺激组呼吸频率和心率分别为(50.23±0.81)和062.60±0..7)min-1(P<0.01),与甲醛溶液组比较分别降低约22%和28%,同时脊髓后角pERK1/2表达减少,与甲醛溶液组比较,减少约58%(P<0.01).结论:肩胛间区低频电刺激可在一定程度缓解家兔胆囊痛模型的生理指标改变,抑制其脊髓后角ERKl,2的激活.     

胃牵涉皮区不同频率电刺激对大鼠胃痛及相关神经元c-fos表达的影响

目的:为临床经牵涉区实施简便易行无创的电刺激缓解胃痛及阐释牵涉痛机制的提供实验依据。方法:(1)实验用SD大鼠,以定量10%Formalin灌胃法建立胃痛模型,经尾静脉注射伊文思蓝,以胃痛大鼠体表"渗漏斑"所在局部作为胃痛牵涉区;(2)用韩式仪以2 Hz、100 Hz、2/100 Hz交替等不同频率和方式,经胃痛牵涉区施加电刺激,观测各实验组大鼠不同时间点的疼痛行为学改变后,进一步用免疫组化法和免疫印迹法测试脑和脊髓等部位的c-fos表达,统计分析各因素的相关性。结果:正常大鼠体表未见明显伊文思蓝渗漏斑,胃痛大鼠在其肩胛间区见有明显渗漏斑;经该渗漏斑所在部位给胃痛大鼠不同频率和方式电刺激后,以2/100 Hz交替电刺激组疼痛行为学评分最低,且其相关部位c-fos表达最少,与其他各组相比,P<0.001。结论:大鼠胃痛牵涉区位于肩胛间区。经此区给予胃痛大鼠2/100 Hz交替电刺激可明显缓解胃痛,且与胃运动与感觉相关部位神经元Fos蛋白等信号物质表达量呈正相关。本研究为临床经牵涉区实施简便易行无创的电刺激缓解胃痛以及阐释牵涉痛发生机制的提供了科学的实验依据。     

皮肤与内脏相关的神经基础研究

目的在皮内注药治疗遗尿症及内脏痛有效的基础上,研究皮肤与内脏相关的初级神经元和交感神经元分布规律。方法通过家兔静脉注射伊文氏蓝(EvansBlue),胃内注入甲醛致胃伤害性刺激,观察皮肤渗漏斑,找出皮肤牵涉区;在牵涉区皮肤和胃黏膜分别注射辣根过氧化物酶(HRP)和荧光素核黄(NY)用神经逆行追踪法分别观察初级神经元和交感神经元分布规律。结果家兔胃痛的皮肤牵涉区在肩及肩胛区,在牵涉区皮肤和胃黏膜分别注射HRP和NY在C8～T8脊神经节和交感神经节相互重叠。结论胃痛的皮肤牵涉区与胃黏膜初级神经元分布的特点是在脊神经节呈节段性分布且相互重叠,在交感神经节呈弥散性分布,无节段性分布,也相互重叠,这可能是皮内注药治疗内脏痛的神经基础。     

牵涉区皮下电刺激减轻大鼠子宫炎性痛

目的:探讨牵涉区皮下电刺激对大鼠子宫炎性痛的影响。方法:建立Formalin子宫炎性痛模型后,在子宫炎性痛牵涉区皮下电针施加不同频率(2 Hz、100 Hz、2/100 Hz)电刺激,通过疼痛行为学评分观察电刺激牵涉区后大鼠疼痛行为学的变化。结果:与Formalin组相比,Formalin+牵涉区电刺激组大鼠疼痛行为学评分明显降低(P<0.01),而以2/100 Hz电刺激最为明显。结论:牵涉区皮下电刺激可以减轻大鼠子宫炎性痛。     

感觉神经激肽1受体拮抗剂N-乙酰-L-色氨酸-苯甲基酯抗过敏反应效用评价

目的:评价神经激肽1(neurokinin 1,NK1)受体拮抗剂N-乙酰-L-色氨酸-苯甲基酯(N-Acetyl-L-tryptophan benzyl ester,ATBE)对P物质的拮抗效应,为过敏性疾病的治疗寻找敏感的药物。方法:实验于2004-09／2004-11在南方医科大学完成。①鼠皮肤炎症模型的建立:从BALB／c鼠尾静脉注射标定浓度的Evans蓝,麻醉后在脊背局部皮内分别注射组胺及生理盐水,随后在皮肤刺激区出现水泡反应(Evans蓝渗漏)。②NK1受体的拮抗:在P物质诱导鼠模型前20,30, 60,120．240 min给予NK1受体拮抗剂ATBE后处死动物,切下反应皮肤区,取少许反应组织放入二甲酰胺溶液的小瓶中保存。实验时取出组织孵育,孵化液用分光光度计进行测定,测量渗出的Evans蓝的吸光度,评估NK1受体拮抗剂ATBE拮抗作用。结果:NK1受体拮抗剂ATBE对局部炎症渗出都有明显的阻断作用,且抑制P物质造成的炎症渗出呈时间依赖模式(P<0．05)。Evans蓝在 0．312-100 mg/L浓度范围内与吸光度呈线性关系。结论:NK1受体拮抗剂ATBE能有效抑制P物质诱导的血浆渗出而控制皮肤的炎性损害;Evans蓝可用于由P物质诱导的鼠局部皮肤炎性损害的定量分析。     

感觉神经激肽1受体拮抗剂N-乙酰-L-色氨酸-苯甲基酯抗过敏反应效用评价

目的：评价神经激肽1（neurokinin 1,NK1）受体拮抗剂N-乙酰-L-色氨酸-苯甲基酯（N-Acetyl-L-tryptophan benzyl ester,ATBE）对P物质的拮抗效应,为过敏性疾病的治疗寻找敏感的药物.方法：实验于2004-09/2004-11在南方医科大学完成.①鼠皮肤炎症模型的建立：从BALB/c鼠尾静脉注射标定浓度的Evans蓝,麻醉后在脊背局部皮内分别注射组胺及生理盐水,随后在皮肤刺激区出现水泡反应（Evans蓝渗漏）.②NK1受体的拮抗：在P物质诱导鼠模型前20,30,60,120,240 min给予NK1受体拮抗剂ATBE后处死动物,切下反应皮肤区,取少许反应组织放入二甲酰胺溶液的小瓶中保存.实验时取出组织孵育,孵化液用分光光度计进行测定,测量渗出的Evans蓝的吸光度,评估NK1受体拮抗剂ATBE拮抗作用.结果：NK1受体拮抗剂ATBE对局部炎症渗出都有明显的阻断作用,且抑制P物质造成的炎症渗出呈时间依赖模式（P＜0.05）.Evans蓝在0.312～100 mg/L浓度范围内与吸光度呈线性关系.结论：NK1受体拮抗剂ATBE能有效抑制P物质诱导的血浆渗出而控制皮肤的炎性损害;Evans蓝可用于由P物质诱导的鼠局部皮肤炎性损害的定量分析.     

皮内注药治疗内脏痛的神经基础与皮肤经络的实质

目的 研究家兔胃伤害性刺激牵涉区皮内神经末梢感受器的初级神经元和交感神经元的分布规律及其末梢分布. 方法 在家兔胃伤害性刺激牵涉区皮内注射辣根过氧化物酶( HRP)和荧光素核黄( NY)逆行神经追踪,观察初级神经元和交感神经元的分布规律及其末梢分布规律. 结果 在 C4～ T10脊神经节( SG)发现标记细胞 , 以 C6～ T8标记细胞较多.在颈、胸交感神经节及腹腔神经节发现标记细胞,以腹腔神经节标记细胞较多.在下丘脑、脊髓后角、骶脊肌的肌膜、胃、膀胱壁的外膜和黏膜、心肌的内、外膜,腹主动脉壁的内、外膜发现有荧光密集区. 结论胃伤害性刺激皮肤牵涉区初级神经元分布在 C4～ T10脊神经节,交感神经元分布在颈、胸交感神经节及腹腔神经节,以腹腔神经节标记细胞较多.其末梢分布于皮肤、血管壁、肌膜、空腔脏器的外膜和粘膜等感觉神经末梢分布区,初级神经元和交感神经元可能有递质上行至脊髓后角和下丘脑,这可能是皮内注药治疗内脏痛的神经基础也可能与皮肤经络的实质有关.     

五种化妆品的皮肤刺激性:平皿法鸡胚试验评价

背景:评价化妆品皮肤刺激性,传统多采用兔皮肤刺激实验,即将化学物涂抹在动物背部去毛的皮肤上,观察其变化,这些方法费时,费力,且给动物带来很大痛苦.各国实验室都在寻找动物替代实验的方法.目的:采用平皿法鸡胚试验方法作为替代方法评价化妆品引起人体皮肤不良反应潜在可能的可行性.设计、时间及地点:动物替代实验,于2008-03/05在中国检验检疫科学研究院毒理学实验室完成.材料:无特定病原体级受精鸡卵,购于北京梅里亚通实验动物技术有限公司.方法:将鸡胚置于平皿中,孵育第8天用于试验,记录出现出血,血管裂解及凝血的鸡胚数目,得出刺激因子X,以判断待测物质是否具有人体皮肤不良反应潜在可能.并与人体斑贴试验结果进行比较.主要观察指标:刺激因子X.结果:5种化妆品中2种有人体皮肤不良反应潜在可能,刺激因子X＞5.与人体斑贴试验结果经校正x2 检验比较,差异无显著性意义,其灵敏度、特异度、阳性预测价值、阴性预测价值均为1.结论:平皿法鸡胚试验和人体斑贴试验的结果一致,该方法可以用来评价化妆品的皮肤刺激性.     

Glucocorticoids inhibit neurogenic plasma extravasation and prevent virus-potentiated extravasation in the rat trachea.

Capsaicin increases the permeability of blood vessels in the rat tracheal mucosa through a mechanism involving the release of tachykinins from sensory nerves. This capsaicin-induced increase in vascular permeability is potentiated by viral infections of the respiratory tract. The present study was done to determine whether this "neurogenic plasma extravasation" can be inhibited by glucocorticoids, to learn the time course of this inhibition, and to determine whether glucocorticoids can prevent the potentiating effect of viral respiratory infections on neurogenic plasma extravasation. Groups of pathogen-free F344 rats were treated with dexamethasone for 2 or 8 h (4 mg/kg i.p.) or 48 or 120 h (0.5-4 mg/kg per d i.p.). Another group of rats was treated with dexamethasone for 120 h following the intranasal inoculation of Sendai virus. The magnitude of plasma extravasation produced by capsaicin or substance P was assessed after this treatment by using Monastral blue pigment and Evans blue dye as intravascular tracers. We found that dexamethasone reduced, in a dose-dependent fashion, the magnitude of plasma extravasation produced in the rat trachea by capsaicin and substance P. Significant inhibition was produced by a dose of dexamethasone as small as 0.5 mg/kg i.p. The effect of dexamethasone had a latency of several hours and reached a maximum after 2 d of treatment. Furthermore, dexamethasone prevented the potentiation of neurogenic plasma extravasation usually present after 5 d of Sendai virus respiratory infection.     

Changes in the Spatiotemporal Expression of Local and Referred Pain Following Repeated Intramuscular Injections of Hypertonic Saline: A Longitudinal Study

Intramuscular injection of hypertonic saline produces a dull ache that is felt in the muscle belly but also often refers into distal structures. We have previously observed in 2 subjects that the pattern of pain referral alters during painful stimuli separated by a week. In this investigation, we tested the hypothesis that the intensity and area of pain in the local and referred regions exhibits plasticity when an identical noxious stimulus is delivered to the same site over sequential trials. Bolus 1 mL intramuscular injections of 5% hypertonic saline were made into the same site of the tibialis anterior (TA) muscle on the same day each week for 4 consecutive weeks. Twenty-one subjects mapped the areas of local and referred pain and rated the intensities on a visual analog scale every 30 seconds until the cessation of pain. Over 4 weeks there was a progressive reduction in the area and intensity of local pain and a reciprocal increase in the expression of referred pain. We conclude that the decrease in perceived local pain and increase in perceived referred pain reflects plastic processes occurring centrally.PerspectiveWhat happens to the intensity of pain induced by repeated noxious stimuli over time? Does it stay the same, increase or decrease? Here we show that weekly injections of hypertonic saline into the tibialis anterior cause decreases in local but increases in referred pain, suggesting central changes in processing noxious inputs.     

The NMDA receptor antagonist MK-801 attenuates c-Fos expression in the lumbosacral spinal cord following repetitive noxious and non-noxious colorectal distention.

The effects of pretreatment with an NMDA receptor antagonist, MK-801, on c-Fos (Fos) expression in the lumbosacral spinal cord following repetitive, noxious (80 mmHg) or non-noxious (20 mmHg) colorectal distention (CRD) was examined immunocytochemically in awake and urethane anesthetized rats. In awake rats, noxious CRD induced Fos expression in the lumbosacral spinal cord. Pretreatment with MK-801 (0.1-1.0 mg/kg, i.p.) produced no change or an increase in noxious CRD induced-Fos expression and caused aversive side effects. In order to examine greater doses of MK-801, further experiments were performed in rats anesthetized with urethane. Both noxious and non-noxious CRD induced Fos in the lumbosacral spinal cord. Pretreatment with MK-801 (0.5, 1.0, 5.0 mg/kg, i.p.) dose-dependently attenuated noxious CRD-induced Fos by 20-40%. Five mg/kg MK-801 attenuated non-noxious CRD-induced Fos by 20%. Lesser doses did not significantly attenuate Fos expression. The laminar distribution of Fos following MK-801 pretreatment revealed a tendency towards the deeper laminae showing the greatest attenuation at the highest dose of MK-801. Protein plasma extravasation in the colon measured with Evan's blue dye showed no difference between rats without balloons, rats with balloons that were not distended and non-noxious CRD. There was significantly more extravasation following noxious CRD. Pretreatment with systemic MK-801 had no effect on plasma extravasation produced by noxious CRD. These data suggest that the induction of Fos in the lumbosacral spinal cord by noxious and non-noxious CRD is partially NMDA receptor mediated. However, NMDA receptor activation contributes significantly more to noxious than non-noxious CRD-induced Fos. Inflammation of the colon following noxious CRD likely contributes to sensitization of colonic afferents which may contribute to the increased NMDA receptor-mediated Fos following the noxious stimulus.     

Offset analgesia: a temporal contrast mechanism for nociceptive information

Temporal filtering of afferent information is an intrinsic component of the processing of numerous types of sensory information. To date, no temporal filtering mechanism has been identified for nociceptive information. The phenomenon of offset analgesia, the disproportionately large decrease in perceived pain following slight decreases in noxious thermal intensity, however, suggests the existence of such a mechanism. To test the hypothesis that a temporal filtering mechanism is engaged during noxious stimulus offset, subjects rated heat pain intensity while stimulus fall rates were varied from -0.5 to -5.0 degrees C/s. In the absence of a temporal filtering mechanism, pain intensity would be expected to decrease in direct proportion to the stimulus fall rate. However, psychophysical fall rates were considerably faster than stimulus fall rates, such that subjects reported no pain while stimulus temperatures were clearly within the noxious range (47.2 degrees C). In addition, paired noxious stimuli were presented simultaneously to determine if offset analgesia evoked by one stimulus could inhibit pain arising from a separate population of primary afferent neurons. Pain ratings were significantly lower than those reported from two constant 49 degrees C stimuli when offset analgesia was induced proximal to, but not distal to, a second noxious stimulus. These asymmetric spatial interactions are not readily explained by peripheral mechanisms. Taken together, these findings indicate that offset analgesia is mediated in part by central mechanisms and reflect a temporal filtering of the sensory information that enhances the contrast of dynamic decreases in noxious stimulus intensity.     

The representation of prolonged and intense, noxious somatic and visceral stimuli in the ventrolateral orbital cortex of the cat.

The responses of single neurones in the ventrolateral orbital (VLO) cortex to noxious pinch, heating of the skin, twisting of the joints and distension of the gall bladder were studied in cats anaesthetized with halothane. Of 60 neurones studied, 44 responded to prolonged (greater than 10 sec) stimuli that were well within the noxious range. Neurones were relatively unresponsive to innocuous stimuli or to the transient application of noxious stimuli. Many single neurones responded to a variety of modalities of noxious stimuli (e.g., skin heating and gall bladder distension). Many neurones studied showed a fluctuating level (5-15 Hz) of ongoing spontaneous activity. Neurones responded with either an increased frequency of spikes (excitation) or an inhibition of spontaneous discharge, irrespective of the source of noxious stimulation. Noxious stimuli delivered simultaneously to two different tissues (e.g., skin and visceral) sometimes produced excitation of the neurone under study, to levels above that produced by the application a noxious stimulus to only one of the tissues. Receptive fields were often large involving both contralateral and ipsilateral areas of the body, as well as both fore and hind limbs. No evidence of somatotopic organization was obtained. The responses of some neurones outlasted the application of the stimuli by many minutes. It is concluded that single neurones in the ventrolateral orbital cortex respond to the prolonged application of intensely noxious stimuli to a variety of body tissues, in a manner that is in keeping with the involvement of this cortical area in both the physiological, autonomic and experiential components of the affective-motivational aspect of pain. Furthermore, from the consequences of lesion studies in man and animals, it is proposed that the activation of cells in the orbital cortex by a variety of noxious stimuli reflects its more general role in the development and maintenance of behaviour in response to negative reinforcement of both social and physical origins.     

Autonomic responses and pain perception in Alzheimer's disease.

We analysed the effects of electrical noxious stimulation on the autonomic nervous system of Alzheimer's disease (AD) patients who were assessed by means of the Mini Mental State Examination test (MMSE). To do this, we used electrical stimuli at two different intensities: just above pain threshold and twice pain threshold. We recorded heart rate and systolic blood pressure by using conventional electrocardiography and finger photo-plethysmography. When a pain stimulus just above threshold was delivered, AD patients were found to have blunted autonomic responses compared to controls of the same age. Similarly, prestimulus expectation produced a less pronounced increase of the responses in AD patients compared to the controls. However, when the painful stimulus was increased to twice the pain threshold, the systolic blood pressure increase of AD patients did not differ from the controls, whereas heart rate increase was still slightly diminished. By contrast, pain perception was similar in the two groups when the stimulus was at pain threshold, whereas it was blunted in AD patients when the stimulus was twice the pain threshold. These findings show that in AD mild noxious stimulation produces blunted autonomic responses and normal pain perception, whereas strong noxious stimulation produces quasi-normal autonomic responses and blunted pain perception. These results indicate that AD patients have an increased threshold for both autonomic activation and pain tolerance.     

The visceromotor responses to colorectal distension and skin pinch are inhibited by simultaneous jejunal distension

Noxious stimuli that are applied to different somatic sites interact; often one stimulus diminishes the sensation elicited from another site. By contrast, inhibitory interactions between visceral stimuli are not well documented. We investigated the interaction between the effects of noxious distension of the colorectum and noxious stimuli applied to the jejunum, in the rat. Colorectal distension elicited a visceromotor reflex, which was quantified using electromyographic (EMG) recordings from the external oblique muscle of the upper abdomen. The same motor units were activated when a strong pinch was applied to the flank skin. Distension of the jejunum did not provoke an EMG response at this site, but when it was applied during colorectal distension it blocked the EMG response. Jejunal distension also inhibited the response to noxious skin pinch. The inhibition of the visceromotor response to colorectal distension was prevented by local application of tetrodotoxin to the jejunum, and was markedly reduced when nicardipine was infused into the local jejunal circulation. Chronic sub-diaphragmatic vagotomy had no effect on the colorectal distension-induced EMG activity or its inhibition by jejunal distension. The nicotinic antagonist hexamethonium suppressed phasic contractile activity in the jejunum, had only a small effect on the inhibition of visceromotor response by jejunal distension. It is concluded that signals that arise from skin pinch and colorectal distension converge in the central nervous system with pathways that are activated by jejunal spinal afferents; the jejunal signals strongly inhibit the abdominal motor activity evoked by noxious stimuli.