NGF对鸡胚背根神经节神经突起生长作用的机制研究

目的探讨神经生长因子(nerve growth factor,NGF)促进鸡胚背根神经节(dorsal root ganglion,DRG)神经突起生长的作用机制。方法实验采用9 d的鸡胚分离背根神经节,原代培养法,观察鸡胚DRG的体外生长情况。通过半定量PCR检测诱导型一氧化氮合酶(iNOS)mRNA表达,采用NO检测试剂盒检测NO释放水平。结果 NGF能明显促进鸡胚背根神经节神经突起生长,同时可见NGF抑制iNOS mRNA表达,NO检测结果显示,添加NGF培养的背根神经节上清NO分泌水平明显降低,与阴性对照组比较差异显著(P0.05)。结论 NGF可促进鸡胚背根神经节神经突起生长,其作用与其下调iNOS mRNA表达及抑制神经损伤因子NO释放有关。     

激活素A及激活素结合蛋白用于脑血管病治疗前后的变化

目的探讨激活素A以及激活素结合蛋白在脑血管病治疗前后的变化。方法收集2013-01—2014-01我院收治的脑出血患者22例为A组,脑梗死患者22例为B组,健康志愿者22例为C组,分别采集A、B组治疗前后及C组的外周静脉血,采用放射免疫法测定血清激活素A以及激活素结合蛋白水平。结果 A、B组治疗前的激活素A水平显著高于C组(P0.05);治疗后A、B组的激活素A均显著降低(P0.05),但与C组无明显差异(P0.05);A组治疗前激活素结合蛋白水平显著高于C组(P0.05),治疗后显著降低(P0.05),但与C组无明显差异(P0.05);B组激活素结合蛋白治疗前后均与C组无明显差异(P0.05)。结论脑出血患者存在外周血激活素A以及激活素结合蛋白水平明显升高现象,脑梗死仅表现为激活素A明显升高,治疗后均可恢复至正常水平。     

激活素A维持小鼠大脑皮质神经元存活和增加神经元电压依赖性钠电流

背景：研究表明,激活素A作为神经细胞生存因子,参与神经损伤的保护作用以及维持海马神经元的存活。目的：观察激活素A对大脑皮层神经元长期存活及其对神经元电压依赖性钠电流的影响。设计、时间及地点：随机的细胞和分子生物学及电生理学实验,于2007-2009年在吉林大学白求恩医学院免疫学系完成。材料：孕17天小鼠购于吉林大学动物中心;小鼠源性Neuro-2a 细胞来自美国ATCC;人重组激活素A购于R&D公司。方法：①取孕17天小鼠胚胎的大脑皮层神经元进行原代培养,于加入5ng/ml 激活素A和4ng/ml NGF后的第1、3、5、7和9天进邢盍觳猓虎诩?ng/ml 激活素A对Neuro-2a 细胞Na+电流（INa）的影响;③RT-PCR检测激活素A对Neuro-2a细胞ActRII、VIP及iNOS mRNA表达的影响。主要观察指标：①台酚蓝染色观察活细胞,通过甲苯胺蓝染色尼氏小体计数神经元;②膜片钳实验记录全细胞INa。结果：①对照组存活的神经元呈现时间依赖性减少,在培养第7天,几乎没有存活的神经元,而NGF组和激活素A组在培养第9天仍有部分存活的神经元;②与对照组 INa (0.817±0.21 nA)相比较,激活素A增加Neuro-2a细胞INa (1.044±0.22 nA) (p< 0.05) (n=10);③激活素A诱导Neuro-2a细胞ActRIIA、ActRIIB 和VIP mRNA的表达,降低iNOS mRNA表达。结论：激活素A维持小鼠大脑皮层神经元长期存活,增加神经元电压依赖性Na+电流。     

电针刺激增强备用背根猫背核组织提取液的促神经突起生长效应

切除１０只雄猫的一侧Ｌ１～Ｌ５、Ｌ７～Ｓ２背根节（ＤＲＧ）。保留Ｌ８背根为备用背根。电针刺激５只猫术侧Ｌ６背根分布区的足三里和悬钟、三阴交和伏兔两组穴位１０天，另５只不针刺。术后第１１天取脊髓背核组织，制备提取液，加入培养液。参照组以Ｈａｎｋｓ平衡盐溶液代替提取液。用不同的条件培养液培养鸡胚ＤＲＧ。于培养２４与４８小时测定各ＤＲＧ神经突起的长度，以同一批培养各组平均长度与参照组平均长度的比值衡量各组提取液对ＤＲＧ神经突起生长的影响。组间比较发现，针刺手术侧组的平均比值明显大于非针刺手术侧组和非手术侧组，非针刺的求侧组又明显大于非手术侧组。提示，部分去背根传入猫的背核组织提取液，其促进神经突起生长的作用增强，电针刺激穴位能进一步提高其促神经突起生长的效应。     

表皮生长因子对卵巢激活素基因表达的调节及其信号通路**

背景：激活素作为卵巢内调控分子,对卵巢卵泡发育起着重要作用。 目的：探索表皮生长因子在激活素基因表达过程中所起的重要作用以及可能参与调节的信号通路。 方法：分离斑马鱼卵巢卵泡,体外培养6 d,消化后传代培养24 h。表皮生长因子单独或与其他分子抑制剂(AG1478、H89、GF109203X)或激动剂(FK、PMA)联合处理细胞,提取细胞RNA,反转录PCR检测细胞激活素表达量。 结果与结论：表皮生长因子可快速提高激活素表达量,其作用可能是通过磷酸化信号分子丝裂原活化蛋白激酶实现,而蛋白激酶C特异性抑制剂或激动剂可减弱或加强表皮生长因子对丝裂原活化蛋白激酶信号分子的激活,显示卵巢内激活素表达受表皮生长因子调节,蛋白激酶C/丝裂原活化蛋白激酶信号通路参与了这种调节作用。蛋白激酶A抑制剂也能抑制表皮生长因子对激活素表达的促进作用。     

Aβ25-35激活小胶质细胞机制的研究

目的以Aβ25-35为工具药，观察小胶质细胞激活后形态和功能的变化，以探讨阿尔茨海默病发病过程中活化的小胶质细胞对神经元损伤作用的可能机制。方法用Aβ25-35处理体外培养的大鼠小胶质细胞，采用倒置相差显微镜观察细胞形态，RT-PCR方法检测炎性因子肿瘤坏死因子（TNF）-α、白介素（IL）-1β、诱导型一氧化氮合酶（iNOS）mRNA表达，Griess反应检测一氧化氮（NO）生成量，超氧化物岐化酶可抑制的WST-1还原法检测细胞外超氧化物产生量。结果Aβ25-35作用后，小胶质细胞形态发生明显变化，细胞胞体延长，由胞体伸出一个或多个树枝状突起，变为类巨噬样细胞；细胞外超氧化物产生量明显增加；但TNF—α、IL-1β、iNOS mRNA表达及NO释放量无明显改变。结论Aβ25-35激活小胶质细胞以活性氧类物质产生为主，活性氧类物质在介导Aβ诱导的神经毒性中起了关键作用。     

血管紧张素Ⅱ和缬沙坦对血管平滑肌细胞增殖和基质金属蛋白酶9表达的影响

目的研究血管紧张素Ⅱ（AngⅡ）对大鼠血管平滑肌细胞（VSMC）的增殖及基质金属蛋白酶-9（MMP-9）表达的影响及缬沙坦（Valsartan）的干预作用。方法分离大鼠胸主动脉平滑肌细胞,贴块法原代培养大鼠血管平滑肌细胞,取第5～10代细胞进行实验,随机分成AngⅡ组（A组）、AngⅡ＋Valsartan组（A＋V组）、Valsartan组（V组）及对照组4组,各组细胞分别采用四唑盐（MTT）比色法检测血管平滑肌细胞增殖,逆转录聚合酶链反应和免疫印迹法测定MMP-9mRNA和蛋白的表达水平。结果与对照组比较,AngⅡ组VSMC的增殖率升高;与AngⅡ组比较,AngⅡ＋Valsartan组VSMC的增殖率降低,差异均明显（P〈0．05）。AngⅡ组与对照组比较,MMP-9mRNA和蛋白的表达均升高;AnⅡValsartan组与AngⅡ组比较,MMP_9mRNA和蛋白的表达均降低,差异均显著（P〈O．05）。结论血管紧张素Ⅱ可以诱导血管平滑肌细胞的增殖和基质金属蛋白酶-9mRNA和蛋白的表达增加;血管紧张素Ⅱ1型受体（AT1R）拮抗剂一缬沙坦可抑制此作用,ATR参与介导此过程。     

小鼠癫痫持续状态后海马活化素A、抑制素A蛋白及活化素受体ⅡAmRNA表达的变化

目的观察小鼠癫痫持续状态(status epilepticus，SE)后海马活化素A、抑制素A蛋白及活化素受体ⅡA mRNA表达的变化。方法采用匹鲁卡品诱导的SE小鼠模型．并以正常及未成SE的小鼠（NSE)作对照，分别应用Western blot与RT-PCR观察SE后活化素A与抑制素A蛋白及活化素受体ⅡA mRNA表达的动态变化。结果（1)正常小鼠海马活化素A含量很低；SE后3h开始增高，6h增高达显著性差异水平，24h至高峰，48h仍维持在较高水平，而NSE鼠活化素A表达无明显增加；(2)抑制素A蛋白SE后无明显增加，同正常及NSE小鼠比较无显著差异；(3)活化素受体ⅡA mRNA在SE后表达无明显变化。结论海马活化素A蛋白在匹鲁卡品诱导SE后明显上调．但活化素受体ⅡA mRNA及抑制素A蛋白无明显上调。     

脑神经肽对人胚神经细胞生长发育及对小鼠急性缺血脑组织的影响

目的和方法应用体外培养的人胚背根神经节和大脑皮层神经元，研究了神经肽对神经组织和神经细胞生长发育的影响，还通过小鼠的急性脑缺血模型研究神经肽对急性脑缺血的作用。结果发现神经肽可使背根神经节突起的长度和密度增加，皮层神经元存活数增加，神经元分化率增高，胞体增大，突起延长，细胞生长加快。超微结构观察显示神经肽可增强细胞内的合成代谢和细胞间连接。脑神经肽可使小鼠脑组织水肿较轻神经元无明显空泡，细胞超微结构改变较轻微，脑组织LDH活性高于对照组。结论神经肽对神经组织具有营养作用，并对急性脑缺血也具有保护作用。     

P物质对大鼠新鲜分离DRG神经元NMDA及GABA激活电流的调制作用

实验应用全细胞膜片钳技术在大鼠新鲜分离的背根神经节（ＤＲＧ）神经元胞体膜上观察到Ｐ物质（ＳＰ）对ＮＭＤＡ和ＧＡＢＡ激活电流有调制作用。单独给予ＳＰ（１０－８～１０－６ｍｏｌ／Ｌ）可在ＤＲＧ神经元记录到一幅值较小的，浓度依赖性的无明显去敏感之内向电流。预加ＳＰ３０ｓ后，其对ＮＭＤＡ和ＧＡＢＡ激活电流分别具有明显的增强和抑制作用，而且ＳＰ对ＮＭＤＡ和ＧＡＢＡ激活电流的增强和抑制作用也是剂量依赖性的。如ＳＰ浓度为１０－７ｍｏｌ／Ｌ时，可使ＮＭＤＡ激活电流较之对照增强４６．３±７．２％（ｘ±ｓ，ｎ＝８）；使ＧＡＢＡ激活电流比对照减小３８．９±７．８％（ｘ±ｓ，ｎ＝６）．ＳＰ的此种调制作用可以在同一ＤＲＧ神经元上观察到。     

Mechanisms of the suppression of the nociceptive reflex of the opening of the mouth during stimulation of the structures of the limbic brain

The comparative effectiveness of the inhibitory influence of tetanic stimulation of hypothalamus, amygdala and limbic cortex on EMG-response of m. digastricus evoked by electrical stimulation of tooth pulp nociceptive afferents was studied in cats anesthetized with a mixture of chloralose and nembutal. It was found that inhibition of the EMG-component of the jaw-opening reflex is most pronounced in case of stimulation of medial and lateral region of the hypothalamus, the inhibitory effect of central and medial nuclei of the amygdala is less pronounced and the effect of the limbic cortex is the weakest. It was shown that the mechanism of the antinociceptive effect of tetanic stimulation of the hypothalamus is not related to the concomitant increase of the blood pressure. After stabilization of the blood pressure the suppressive effect of the hypothalamus remains without changes, that points out to a direct, primary, not baro-afferent mechanism of the inhibition of the activity of nociceptive neurons of the trigeminal sensory nuclei. Noradrenaline, injected intravenously, induced a large increase of the blood pressure accompanied by a pronounced inhibition of the pain reflex. Angiotensin causes the same degree of blood pressure elevation without changes in the amplitude of the EMG-response of the pain reflex. Hypothalamic and noradrenergic mechanisms for control of pain sensitivity are discussed.     

药物治疗与合并认知行为治疗对强迫症疗效的比较

目的探讨认知行为心理治疗(CBT)在强迫症(OCD)患者各亚型治疗中的有效性和规律性。方法本研究为临床对照研究。符合入组标准的强迫症患者按患者自愿原则分为两组,治疗观察3、6、12个月。疗效评定分别运用Yale-Brown强迫量表,自拟的自评好转程度量表和临床疗效评定。结果认知行为心理治疗合并药物治疗组31例,临床有效率70.9%,其中治愈率1.8%。单纯药物治疗组24例,临床有效率33.3%。Yale-Brown强迫量表和自评量表得分在6个月和12个月两组有显著差异(P<0.05)。其中强迫症亚型(怕脏型、反复检查型和反复担心型)的疗效比较,怕脏型在治疗3个月末两组间自评量表评分有显著性差异(P<0.05);反复担心型在治疗6个月末两组间Yale-Brown强迫量表总分有显著性差异(P<0.05);反复检查型两组间无统计学差异。结论认知行为心理治疗合并药物治疗强迫症的疗效明显优于单纯药物治疗。强迫症的亚型在治疗中的有效性次序为:反复担心型>怕脏型>反复检查型。     

Characteristics of the sympathetic innervation of the nictitating membrane and of the vasculature of the nose and tongue of the cat

Summary Vasomotor responses from the nasal mucosa and tongue, and contractions of the nictitating membrane, were recorded on stimulation of the cervical sympathetic or internal carotid nerves.Preganglionic sympathetic nerve fibres which elicited a membrane response possessed a lower threshold than those which evoked nasal vasoconstriction, while the latter displayed a lower threshold than fibres which evoked tongue vasoconstriction. The sympathetic vasodilator fibres to the tongue, whose activity was revealed after-receptor blockade, had a similar threshold to the vasoconstrictor fibres.Membrane contraction, nasal vasoconstriction and occasionally tongue vasoconstriction could be evoked by stimulating the internal carotid nerve. The postganglionic fibres innervating the nasal mucosa had a similar threshold to those of the nictitating membrane, which may indicate that there are small myelinated fibres innervating the mucosa.The preganglionic compound nerve action potential had four major components, S₁–S₄. S₁, S₂ and usually S₃ fibres were associated with membrane contraction; S₂, S₃ and sometimes S₁ fibres were associated with nasal vasoconstriction; and S₃, usually S₂ and occasionally S₁ fibres were associated with vasoconstriction in the tongue. It is concluded that each of these three groups of nerve fibres, but not S₄ fibres, may include fibres associated functionally with the three effectors.There was a considerable difference between the relative amplitude of the responses of the three effectors elicited by stimulation of the cervical sympathetic nerve at frequencies between 0.2 and 2 Hz. Vasoconstrictor responses were relatively larger than membrane contractions suggesting differences in the mechanisms of neurotransmission at the neuroeffector junctions.     

Development of the projection from the nucleus of the brachium of the inferior colliculus to the superior colliculus in the ferret

Neurons in the deeper layers of the superior colliculus (SC) have spatially tuned receptive fields that are arranged to form a map of auditory space. The spatial tuning of these neurons emerges gradually in an experience-dependent manner after the onset of hearing, but the relative contributions of peripheral and central factors in this process of maturation are unknown. We have studied the postnatal development of the projection to the ferret SC from the nucleus of the brachium of the inferior colliculus (nBIC), its main source of auditory input, to determine whether the emergence of auditory map topography can be attributed to anatomical rewiring of this projection. The pattern of retrograde labeling produced by injections of fluorescent microspheres in the SC on postnatal day (P) 0 and just after the age of hearing onset (P29), showed that the nBIC-SC projection is topographically organized in the rostrocaudal axis, along which sound azimuth is represented, from birth. Injections of biotinylated dextran amine-fluorescein into the nBIC at different ages (P30, 60, and 90) labeled axons with numerous terminals and en passant boutons throughout the deeper layers of the SC. This labeling covered the entire mediolateral extent of the SC, but, in keeping with the pattern of retrograde labeling following microsphere injections in the SC, was more restricted rostrocaudally. No systematic changes were observed with age. The stability of the nBIC-SC projection over this period suggests that developmental changes in auditory spatial tuning involve other processes, rather than a gross refinement of the projection from the nBIC.     

A contribution to the study of the architecture of the autonomic nervous system of the digestive tract of the rat

Summary The distribution of aminergic and non-aminergic nerve fibres to the different constituents of the wall of the digestive tract in various regions is described. Aminergic fibres synapse with all nervous perikarya. Densely interlacing networks of nerve fibres are found in both layers of the tunica muscularis and in the lamina muscularis mucosae. A finely meshed plexus is observed in relation to the wall of the blood vessels in the wall of the gut. There are many fibres connecting the muscular and the vascular plexus. No nerve fibres have been observed in direct relation to the epithelium.The functional implications of these findings are discussed.