慢性应激抑郁大鼠海马CA1神经元突触可塑性研究

目的 探讨慢性轻度不可预见应激(chronic unpredictable mild stress,CUMS)抑郁模型大鼠海马CA1区神经元的突触可塑性改变.方法 将20只雄性Sprague-Dawley (SD)大鼠随机等分为CUMS组和对照组,前者连续28天每天随机接受不同的应激,对照组同样条件下饲养但不给应激,至第28天进行行为测评后处死,在日立(H7500)透射电镜下测量海马CA1神经元突触界面结构参数.结果 CUMS抑郁大鼠海马CA1神经元突触活性区长度(216.64±20.19 nm)及突触后致密物厚度(42.4±5.23 nm)显著小于对照组(321.58±12.27nm,69.6±4.77 nm),差异有统计学意义(P＜0.05),突触界面曲率及宽度与对照组差异无统计学意义(P＞0.05).结论 慢性应激性抑郁大鼠存在海马CA1区神经元突触可塑性的改变.这提示抑郁症的发病机制可能与海马神经元突触可塑性相关.     

慢性吗啡处理大鼠伏隔核、海马CA1区神经元突触界面结构改变

为探讨慢性吗啡处理导致的神经元突触可塑性改变，采用透射电镜技术测量慢性吗啡处理大鼠伏隔核及海马CA1区神经元突触界面结构参数，并与对照组进行比较。     

反义Noggin基因对成年大鼠海马内Nestin及GFAP表达的影响

目的探讨Noggin基因对成年大鼠海马内Nestin及GFAP表达的影响。方法反义寡核苷酸技术封闭内源性Noggin基因的表达,免疫组化法检测成年大鼠海马内Nestin与GFAP的表达。结果侧脑室连续4d注射Noggin基因的反义寡核苷酸后,可见海马齿状回(dentate gyrus,DG)内Nestin阳性细胞数与GFAP阳性细胞数较对照组显著增加;室下区GFAP阳性细胞数亦明显增加。结论Noggin对成年海马干细胞的分化有重要作用,内源性Noggin基因的表达可使神经干细胞向神经元方向分化。     

颅脑创伤后大鼠海马CA1锥体细胞内在电生理和兴奋性突触传递特性的变化

目的 在细胞及突触水平探讨外伤后癫痫的发病机制.方法 自由落体致伤法制备大鼠颅脑创伤模型,采用膜片钳技术监测海马CA1区锥体细胞内在电生理特性和局部突触兴奋性的变化.结果 颅脑创伤后,大鼠CA1锥体细胞膜输入阻抗和时间常数增加,动作电位的阈电流降低;给予配对刺激后,海马CA1区兴奋性突触后电流表现为配对脉冲比率的降低及配对脉冲易化向配对脉冲抑制的转变.结论 颅脑创伤后海马CA1区神经元内在兴奋性和突触传递功能增强,这些改变可能是外伤后癫痫发病的重要原因.     

三磷酸胞苷二钠对脑缺血大鼠海马CA3区突触体素表达的影响

目的　研究局灶性脑缺血后海马 CA3 区突触体素的动态表达及其三磷酸胞苷二钠对其干预的影响。方法　选取 SD大鼠 60只 ,随机分为脑缺血后自然恢复组、药物干预组和假手术对照组。采用线栓法建立大脑中动脉脑缺血大鼠模型 ,应用免疫组化技术观察海马 CA3 区突触体素的表达。结果　脑缺血后自然恢复组大鼠突触体素的表达较对照组明显降低 (P<0 .0 1) ;但 7～ 2 1d突触体素的表达逐渐上调 (P<0 .0 1)。应用三磷酸胞苷二钠干预后 ,突触体素表达与自然恢复组相比明显升高。结论　脑缺血损伤后海马 CA3 区突触体素表达减少 ,但机体自身存在着神经元的修复和再生 ;三磷酸胞苷二钠可上调突触体素的表达 ,具有促进缺血神经元的修复、再生及突触重塑作用。     

亚低温对大鼠弥漫性脑损伤后海马CA3区HSP70表达及细胞凋亡的影响

目的 观察亚低温对大鼠弥漫性脑损伤(DBI)后海马CA3区HSP70在蛋白质和mRNA水平的表达及细胞凋亡上的影响,探讨亚低温脑保护分子生物机制。方法 将大鼠随机分成空白对照、假手术、单纯DBI和DBI后亚低温治疗四组,按Marmarou氏方法制作大鼠DBI模型,采用免疫组化法、逆转录聚合酶链反应(RT-PCR)及流式细胞仪(FCM),分别观察各组动物脑海马CA3区HSP70在蛋白质和mRNA水平的表达及细胞凋亡率。结果 与对照组相比,大鼠DBI后海马CA3区HSP70表达水平及细胞凋亡率均升高(P<0.05);亚低温治疗后,大鼠脑海马CA3区HSP70表达水平较单纯DBI组显著增高(P<0.01),而细胞凋亡率则明显降低(P<0.05)。结论 亚低温对创伤性脑损伤的脑保护机制可能与促进HSP70表达,并减少神经细胞凋亡有关。     

16Hz次声作用对大鼠海马TRPV4、GFAP和fos蛋白表达的影响

目的研究16Hz，90dB和130dB次声作用后，大鼠海马瞬时感受电位香草酸家族4（TRPV4）通道蛋白、胶质纤维酸性蛋白（GFAP）和fos蛋白的表达情况。方法16Hz，90dB和130dB次声作用于大鼠，2h／d，作用7d后采用免疫组织化学染色方法，观察大鼠海马中TRPV4蛋白、GFAP和fos蛋白表达的情况。结果16Hz，130dB次声作用7d后，与对照组相比较大鼠海马中显著表达TRPV4阳性神经元，GFAP阳性星形胶质细胞和fos阳性神经元（P〈0．05），三者分布一致，关系密切；90dB组大鼠的上述三种蛋白表达均较130dB组弱（P〈0．05）。结论16Hz，90dB和130dB次声作用可以引起大鼠海马TRPV4阳性细胞表达增多，且能够激活神经元和星形胶质细胞。     

ACh对正常大鼠和吗啡成瘾大鼠海马CA1区痛反应电活动的影响

目的 研究ACh对正常大鼠和吗啡成瘾大鼠海马CA1区痛兴奋神经元(pain-excitation neurons,PEN)和痛抑制神经元(pain-inhibitation neurons,PIN)电活动的影响,进一步探讨ACh对正常和吗啡成瘾状态下CA1区痛觉调制的作用及机制.方法 电刺激坐骨神经作为伤害性电刺激,在细胞外用玻璃微电极记录CA1区PEN和PIN的放电,观察ACh对正常大鼠和吗啡成瘾大鼠CA1区PEN和PIN电活动的影响.结果 伤害性刺激能够增强PEN的电活动,而减弱PIN的电活动.正常大鼠中,ACh使PEN的痛诱发放电频率降低,PIN的放电频率增加;ACh的作用在注射后4 min达到峰值.吗啡成瘾大鼠中,ACh同样也抑制了PEN的电活动,兴奋PIN的电活动,但是作用的高峰出现在注射后6min.胆碱能受体拮抗剂阿托品可阻断ACh的作用.结论 海马CA1区内的胆碱能神经元和毒蕈碱受体参与了伤害性信息的处理,并且起到了镇痛作用.吗啡成瘾可以降低CA1区痛反应神经元对伤害性刺激的敏感性.     

孕烯醇酮对老年SD大鼠海马区突触素蛋白1表达的影响

目的 观察孕烯醇酮(PREG)慢性干预对老年大鼠海马区突触素蛋白1(SYP1)表达的变化.方法 24月龄雄性SD大鼠40只,随机分为空白对照组、溶剂对照组、小剂量PREG(0.5 mg/kg)干预组、大剂量PR EG(2.0 mg/kg)干预组,隔日腹腔注射干预1个月,通过Western Blot技术和免疫组化技术检测分别检测各组大鼠海马区蛋白表达情况.结果 与对照组相比,免疫组化显示大剂量PREG干预组大鼠海马区SYP1表达明显增加,Western Blot定量检测发现大鼠海马区SYP1表达也显著升高(P＜0.05);而小剂量组SYP1表达无明显增加(P＞0.05).结论 大剂量PREG干预可以显著增加老年大鼠记忆相关的海马区SYP1表达,SYP1表达增加可能明显改善大鼠海马区突触结构可塑性与突触功能,进而有助于改善老年大鼠的学习记忆功能.     

吗啡依赖和戒断大鼠海马CA1区TNF-α和GDNF的表达变化

药物成瘾或药物依赖是一种慢性复发性脑疾病。药物依赖的形成机制目前尚不清楚，已有研究提示胶质源性神经营养因子（GDNF）和肿瘤坏死因子（TNF-α）等多种因子可能参与药物依赖形成过程，但这方面的实验证据尚少，很多环节需进一步研究。     

Effects of diazepam on glutamatergic synaptic transmission in the hippocampal CA1 area of rats with traumatic brain injury

The activity of the Schaffer collaterals of hippocampal CA3 neurons and hippocampal CA1 neurons has been shown to increase after fluid percussion injury. Diazepam can inhibit the hyperexcitability of rat hippocampal neurons after injury, but the mechanism by which it affects excitatory synaptic transmission remains poorly understood. Our results showed that diazepam treatment significantly increased the slope of input-output curves in rat neurons after fluid percussion injury. Diazepam significantly decreased the numbers of spikes evoked by super stimuli in the presence of 15 μmol/L bicuculline, indicating the existence of inhibitory pathways in the injured rat hippocampus. Diazepam effectively increased the paired-pulse facilitation ratio in the hippocampal CA1 region following fluid percussion injury, reduced miniature excitatory postsynaptic potentials, decreased action-potential-dependent glutamine release, and reversed spontaneous glutamine release. These data suggest that diazepam could decrease the fluid percussion injury-induced enhancement of excitatory synaptic transmission in the rat hippocampal CA1 area.     

细胞外三磷酸腺苷对大鼠脊髓损伤后胶质纤维酸性蛋白表达的影响

目的 研究细胞外三磷酸腺苷(ATP)对大鼠脊髓损伤后胶质纤维酸性蛋白(GFAF)表达和运动功能恢复的影响.方法 健康成年Wistar大鼠66只按照随机数字表法取6只作为正常对照组,余60只制作成脊髓打击伤动物模型,并再按照随机数字表法分为两组:ATP组(A组,给予ATP注射)和对照组(B组,给予等量生理盐水注射),每组30只大鼠.伤后1、3、7、14和28 d取材,应用免疫组织化学方法观察GFAP的表达,采用计算机图像分析系统进行半定量分析;并用改良的Tarlov评分观察大鼠脊髓损伤后运动功能的恢复情况.结果 大鼠脊髓损伤后GFAP的表达呈进行性升高,损伤后14 d达高峰;在损伤后7、14和28 d,A组大鼠GFAP的表达明显强于B组;脊髓损伤后14 d和28 d,A组大鼠改良的Tarlov评分明显大于B组;以上差异均有统计学意义(P<0.05).结论 细胞外ATP能促进大鼠损伤脊髓表达GFAP,并有助于大鼠脊髓损伤后运动功能的恢复.     

Amphetamine administration improves neurochemical outcome of lateral fluid percussion brain injury in the rat

This study examined the effects of the administration of d-amphetamine on the regional accumulation of lactate and free fatty acids (FFAs) after lateral fluid percussion (FP) brain injury in the rat. Rats were subjected to either FP brain injury of moderate severity (1.9 to 2.0 atm) or sham operation. At 5 min after injury, rats were treated with either d-amphetamine (4 mg/kg, i.p.) or saline. At 30 min and 60 min after brain injury, brains were frozen in situ, and cortices and hippocampi were excised at 0°C. In the saline-treated brain injured rats, levels of lactate were increased in the ipsilateral left cortex and hippocampus at 30 min and 60 min after injury. These increases were attenuated by the administration of d-amphetamine at 5 min after lateral FP brain injury. At 30 and 60 min after FP brain injury, increases in the levels of all individual FFAs (palmitic, stearic, oleic and arachidonic acids) and of total FFAs were also observed in the ipsilateral cortex of the saline-treated injured rats. These increases in the ipsilateral cortex and hippocampus were also attenuated by the administration of d-amphetamine. Neither levels of lactate nor levels of FFAs were increased in the contralateral cortex in the saline-treated injured rats at 30 min or 60 min after FP brain injury. The levels of lactate and FFAs in the contralateral cortex were also unaffected by the administration of d-amphetamine. These results suggest that the attenuation of increases in the levels of lactate and FFAs in the ipsilateral cortex and hippocampus may be involved in the amphetamine-induced improvement in behavioral outcome after lateral FP brain injury.     

Chronic failure in the maintenance of long-term potentiation following fluid percussion injury in the rat

Traumatic brain injury (TBI) can produce chronic cognitive learning/memory deficits that are thought to be mediated, in part, by impaired hippocampal function. Experimentally induced TBI is associated with deficits in hippocampal synaptic plasticity (long-term potentiation, or LTP) at acute post-injury intervals but plasticity has not been examined at long-term survival periods. The present study was conducted to assess the temporal profile of LTP after injury and to evaluate the effects of injury severity on plasticity. Separate groups of rats were subjected to mild (1.1-1.4 atm), moderate (1.8-2.1 atm), or severe (2.2-2.7 atm) fluid percussion (FP) injury (or sham surgery) and processed for hippocampal electrophysiology in the first or eighth week after injury. LTP was defined as a lasting increase in field excitatory post-synaptic potential (fEPSP) slope in area CA1 following tetanic stimulation of the Schaffer collaterals. The fEPSP slope was measured for 60 min after tetanus. Assessment of LTP at the acute interval (6 days) revealed modest peak slope potentiation values (129-139%), which declined in each group (including sham) over the hour-long recording session and did not differ between groups. Eight weeks following injury, slices from all groups exhibited robust maximal potentiation (134-147%). Levels of potentiation among groups were similar at the 5-min test interval but differed significantly at the 30- and 60-min test intervals. Whereas sham slices showed stable potentiation for the entire 60-min assessment period, slices in all of the injury groups exhibited a significant decline in potentiation over this period. These experiments reveal a previously unknown effect of TBI whereby experimentally induced injury results in a chronic inability of the CA1 hippocampus to maintain synaptic plasticity. They also provide evidence that sham surgical procedures can significantly influence hippocampal physiology at the acute post-TBI intervals. The results have implications for the mechanisms underlying the impaired synaptic plasticity following TBI.     

锂-匹罗卡品致痫大鼠海马胶质纤维酸性蛋白的表达及意义

目的观察其海马经HE染色后组织病理学、胶质纤维酸性蛋白免疫反应阳性表达细胞在LPS中各观察时间点海马CA1、CA3、齿状回的表达,探讨其致机制。方法锂-匹罗卡品急性诱导SD癫痫持续状态模型鼠形成后,采用免疫组化和图像分析方法观察海马HE染色组织病理学、胶质纤维酸性蛋白免疫反应阳性表达细胞。结果模型组各时间点海马细胞形态出现病理性改变,部分细胞脱失,胞浆浓缩,胞核固缩深染;胶质纤维酸性蛋白免疫反应阳性表达细胞亦显著上调（P〈0.05）。结论Pilo诱导SD大鼠癫痫发作后存在显著的海马神经元结构和胶质细胞的损伤,以胶质细胞损伤更显著,胶质纤维酸性蛋白持续高表达可能是这种功能异常的胶质细胞增生的重要原因,也可能是锂-匹罗卡品致癫痫发作的重要因素之一。     

Effects of hemoglobin and its breakdown products on synaptic transmission in rat hippocampal CA1 neurons

During head injuries and hemorrhagic stroke, blood is released into the extravascular space. The pooled erythrocytes get lysed and hemoglobin is released into the intracranial cavities. Therefore, neurons may be exposed to hemoglobin and/or its breakdown products, hemin and iron, for long periods of time. In this study, the electrophysiological actions of these agents on synaptic transmission in rat hippocampal CA1 pyramidal neurons were studied using extracellular field- and whole cell patch-recordings. Previously our laboratory reported that commercially available hemoglobin produced a dose dependent suppression of synaptic transmission in hippocampal CA1 neurons. In the present study, however, we found that this depression was caused by impurities present in the hemoglobin samples. Commercially available hemoglobin and methemoglobin did not have a significant effect on synaptic transmission. Although, reduced-hemoglobin prepared using a method described by Martin et al. [J. Pharm. Exp. Ther. 232 (1985) 708], produced a significant depression of synaptic transients, these effects were due to contamination with bisulfite that was present due to the reducing procedure. Therefore, the technique of Martin et al. was inadequate in removing the reducing agents or their breakdown products. A number of studies in literature used commercial samples of hemoglobin or reduced hemoglobin prepared using the method of Martin et al. Our observations indicate that it would be important to determine if contaminants, rather than hemoglobin, are responsible for the observed effects in these studies. Unlike hemoglobin, its breakdown products, ferrous chloride and hemin, produced an irreversible and significant depression of field excitatory postsynaptic potentials. The relevance of these effects in neurological complications that follow head injuries and hemorrhagic stroke awaits further investigation.     

大鼠海马星形胶质细胞对突触可塑性的影响

目的研究星形胶质细胞在神经系统发育成熟过程中对突触可塑性的调控规律.方法取健康初生、幼年和成年大鼠各10只,每只取脑切片,用免疫组织化学方法观察其海马CA1区的S100、胶质纤维酸性蛋白(GFAP)和P38免疫反应产物强度;HE染色法显示神经元胞体.结果初生大鼠海马CA1区中神经元大量存在,但S100、GFAP和P38表达均少,幼鼠的表达增加,但仍显著少于成鼠的表达(P＜0.01).结论大鼠海马CA1区星形胶质细胞增殖与突触出现时间、突触数目增加及功能成熟有关.     

Tissue tears in the white matter after lateral fluid percussion brain injury in the rat: relevance to human brain injury

A characteristic feature of severe diffuse axonal injury in man is radiological evidence of the “shearing injury triad” represented by lesions, sometimes haemorrhagic, in the corpus callosum, deep white matter and the rostral brain stem. With the exception of studies carried out on the non-human primate, such lesions have not been replicated to date in the multiple and diverse rodent laboratory models of traumatic brain injury. The present report describes tissue tears in the white matter, particularly in the fimbria of Sprague-Dawley rats killed 12, 24, and 48 h and 7 days after lateral fluid percussion brain injury of moderate severity (2.1–2.4 atm). The lesions were most easily seen at 24 h when they appeared as foci of tissue rarefaction in which there were a few polymorphonuclear leucocytes. At the margins of these lesions, large amounts of accumulated amyloid precursor protein (APP) were found in axonal swellings and bulbs. By 1 week post-injury, there was macrophage infiltration with marked astrocytosis and early scar formation. This lesion is considered to be due to severe deformation of white matter and this is the first time that it has been identified reproducibly in a rodent model of head injury under controlled conditions. Received: 25 February 1999 / Revised: 7 June 1999 / Accepted: 22 June 1999