横断大鼠穹窿-海马伞对其海马CA3区突触形态的影响

目的：观察横断大鼠穹窿-海马伞对其海马突触形态的影响。方法：横断大鼠双侧穹窿-海马伞（FF）建立动物模型,于手术前、后对大鼠进行迷宫检查,重点对海马CA3区多形层突触界面的结构参数进行定量分析。结果：突触界面曲率减小,突触间隙宽度加大,突触后膜致密物质厚度明显变薄,穿孔性突触的比例也有不同程度降低。结论：横断穹窿-海马伞引起海马CA3区突触形态明显改变,推测海马内Ach的正常水平对维持海马CA3区突触界面超微结构有重要作用。     

一次性电击引起大鼠脑内突触结构可塑性变化的定量观察

运用电镜,对一次性电击引起大鼠脑内Ｇｒａｙ Ⅰ型突触界面某些结构的变化进行了定量观察。在海马ＣＡ３区,突触后膜致密物质显著增厚（Ｐ〈０．０５）,突触间隙宽度极显著增宽（Ｐ〈０．０１）;在大脑皮层感觉运动区,突触界面曲率显著变大（Ｐ〈０．０５）。突触界面弯曲类型无显著性差异。结果提示：一次性电击可以引起大鼠脑内突触界面结构发生可塑性变化。     

产前束缚应激子代大鼠海马神经颗粒素表达降低

神经颗粒素（neurogranin,NG）是脑特异性突触后蛋白,参与在学习记忆功能中起核心作用的信号转导通路及突触可塑性。本研究旨在探讨产前束缚应激对子代大鼠海马NG表达的影响。连续7d对孕晚期大鼠进行束缚应激,建立产前束缚应激模型,分为对照雌、雄组,应激雌、雄组。采用免疫组化方法观察NG在产前束缚应激子代大鼠海马不同亚区的分布特点;采用蛋白免疫印迹方法检测产前束缚应激子代大鼠海马NG蛋白的表达。结果显示：各组子代大鼠海马各区均有NG蛋白表达,CA1和CA3区表达高于齿状回（dentate gyrus,DG）;应激组雌、雄子代大鼠海马NG的表达明显低于对照组（P〈0．01）,应激组雌性子代比雄性子代减少更显著,对照组雌、雄子代之间无差异。免疫组化与蛋白免疫印迹方法所得结果一致。上述结果表明,NG在产前束缚应激子代大鼠海马表达降低,并且雌性比雄性降低明显,NG对产前束缚应激子代大鼠有差异性调制,NG表达减少可能与产前束缚应激子代大鼠学习记忆能力下降有关。     

浅析BDNF改善痴呆老龄鼠记忆障碍CA1区GrayⅠ型突触结构相关参数的变化

目的探讨BDNF（Brain derived neurotrophic factor）改善痴呆老龄鼠记忆障碍的机制。方法利用Morris迷宫试验观察脑内微量注射BDNF对痴呆小鼠自发活动和记忆巩固过程的影响,应用透射电镜和形态计量学分析痴呆老龄鼠海马GrayⅠ型突触的突出结构参数的变化。结果BDNF使痴呆小鼠在新异环境中的自发活动和探究行为明显增多;并显著延长电击后24 h的步入潜伏期（STL）;BDNF使痴呆老龄鼠海马CA1区GrayⅠ型突触的体积密度、面积密度、比表面和面数密度较治疗前增大,突触平均面积增大;突出界面曲率、突出间隙宽度、突出后致密物质均较治疗前增大,而较正常对照组减小。结论突触结构的变化和突出数量的减少是痴呆发病的病理机制之一;BDNF能够促进突触重建,改善痴呆老龄鼠的学习记忆。     

大鼠海马神经元突触超微结构的增龄变化

目的为研究脑老化过程中学习、记忆功能减退的神经结构基础提供实验依据。方法应用透射电子显微镜,观察比较从出生1 d至24月龄（1 d、1月龄、3月龄、6月龄、18月龄、24月龄）的Sprague Dawley大鼠海马神经元突触超微结构的随增龄变化,同时观察与脑老化密切相关的指标脂褐素沉积。结果在大鼠6月龄之前,随着月龄的增加,海马神经元突触超微结构的发育逐渐完善,至6月龄大鼠突触数量明显增多;此后突触数量逐渐减少,至24月龄大鼠神经元突触数量最少。从1月龄开始海马神经元内即可见少量脂褐素颗粒沉积,随着月龄的逐渐增加,至24月龄时脂褐素颗粒沉积显著。结论青年期大鼠的海马神经元突触发育最好,进入老年期后,突触结构受损,老年期损伤最为严重,同时伴有大量的脂褐素颗粒沉积。     

学习训练对谷氨酸单钠所致大鼠海马损伤的神经保护作用

目的：探讨学习训练对谷氨酸神经毒性的保护作用。方法：在SD大鼠生后第3～9d腹腔注射谷氨酸单钠复制谷氨酸毒性模型,在1月龄和2月龄时训练大鼠学会以明暗辨别来获得食物,3月龄时取脑,在光镜下计数海马内存活神经元数,电镜下观察海马CA1区的超微结构,并计数突触数,测量突触活性带长度。结果：学习训练组海马CA3区和CA4区内的存活神经元数、海马CA1区内的突触数和突触活性带长度均大于非学习组,结论：结果提示学习训练可在一定程度上减轻MSG对海马的损伤。     

孕期地塞米松暴露对大鼠子代海马神经元增殖以及突触可塑性的影响

地塞米松是一种糖皮质激素药物，具有抗炎、抑制免疫等多种药理作用，广泛应用于治疗多种疾病。临床上常使用地塞米松来促进早产儿的肺成熟以及预防胎儿呼吸窘迫综合征。目前的流行病学以及试验研究表明，地塞米松孕期暴露会增加子代患软骨病、肾脏损伤等疾病的风险。为了探究孕期地塞米松暴露（prenatal dexamethasone exposure, PDE）对大鼠子代胎鼠海马神经元增殖发育以及胎鼠海马突触可塑性形成的影响，对孕中晚期Wistar大鼠皮下注射地塞米松(0.2 mg·kg^-1·d^-1)，对照组注射等剂量0.9%氯化钠溶液。收集GD20子代海马，采用实时荧光定量PCR以及Western blot法对海马神经增殖、突触可塑性形成和APPL1(adaptor protein containing pH domain, PTB domain and leucine zipper motif 1)进行相关功能检测,并进一步使用投射电镜观察海马突触超微结构。结果显示，与空白对照组相比，PDE胎海马Ki67、增殖细胞核抗原（proliferating cell nuclear antigen, PCNA）、突触融合蛋白(syntaxin)、25 kD突触相关蛋白（SNAP25）等特异性指标显著降低，提示PDE对胎海马神经增殖、突触发育具有不同程度的抑制作用。Western blot 结果显示，PDE组突触后致密蛋白95（PSD95）显著下调，进一步电镜结果证实PDE可导致子代海马突触可塑性受损。APPL1检测结果显示，PDE子代海马APPL1表达水平下调，故而推测PDE可能是通过调节APPL1表达改变子代海马突触可塑性。研究结果揭示PDE具有海马发育毒性并可导致海马神经元增殖减少以及突触可塑性发育损伤。研究结果为指导孕期合理用药和有效评估胎儿海马发育毒性风险提供了实验与理论依据。     

围产期慢性尼古丁摄入对子代小鼠学习与记忆的影响

尼古丁对学习记忆间接作用的研究鲜有报道。昆明小鼠母鼠受孕后随机分为对照组(CON)和尼古丁组(NIC)。CON组母鼠自由饮水,NIC组母鼠饮水中给予浓度为50μg/mL的尼古丁。子代小鼠60日龄时,进行Morris水迷宫实验,之后在体记录海马区穿通纤维通路(perforant pathway,PP)至齿状回(dentate gyrus,DG)的长时程增强(long-term potentiation,LTP)。结果显示,NIC组仔鼠的逃避潜伏期从第3天开始明显大于CON组,目标象限停留时间所占百分比和穿越平台次数均低于CON组,LTP群峰电位幅值和场兴奋性突触后电位斜率也都显著低于CON组。说明由母体摄入的尼古丁,可经胎盘和乳汁明显作用于其子代,导致子代学习记忆功能的明显损伤,其可能机制是因为海马神经元突触传递可塑性的效率显著降低。     

产前850-1900MHz手机暴露对子代大鼠齿状回PCNA和DCX表达的影响

目的：探讨产前手机暴露对子代大鼠海马齿状回增殖细胞核抗原（PCNA）和双皮质素（DCX）表达的影响。方法：构建孕鼠手机射频暴露模型,分为对照组、短时暴露组和长时暴露组（n=6）,短时和长时暴露组于孕第1-17天分别给予6 h/d和24 h/d的手机通话暴露,观察孕鼠的孕期长短、孕期体重增长和各组的胎儿数、胎儿出生体重。1月龄子代大鼠行焦油紫染色观察海马齿状回细胞形态,免疫组化观察齿状回PCNA和DCX表达,Western blot检测DCX和脑源性神经营养因子（BDNF）表达。结果：各组孕鼠的孕期、妊娠期体重增长和各组的胎儿数、胎儿出生体重无显著差异,长时暴露组子代大鼠的齿状回多形细胞层锥形细胞和DCX阳性细胞出现形态改变。与对照组、短时暴露组比较,长时暴露组子代大鼠齿状回PCNA阳性细胞和DCX、BDNF表达均明显减少（P<0.05）。结论：产前长时手机暴露可能通过改变子代大鼠海马BDNF而影响齿状回的PCNA和DCX表达。     

突触前α7烟碱受体对海马神经元兴奋性突触传递的调控

采用盲法膜片钳技术观察突触前烟碱受体(nicotinic acetylcholinel receptors,nAChRs)对海马脑片CAl区锥体神经元兴奋性突触传递的调控作用。结果显示,nAChRs激动剂碘化二甲基苯基哌嗪(dimethylphenyl—piperazinium iodide,DMPP)不能在CAl区锥体神经元上诱发出烟碱电流。DMPP对CAl区锥体神经元自发兴奋性突触后电流(spontaneous excitatory postsynaptic current,sEPSC)具有明显的增频和增幅作用,并呈现明显的浓度依赖关系。DMPP对微小兴奋性突触后电流(miniature excitatory postsynaptic current,mEPSC)具有增频作用,但不具有增幅作用。上述DMPP增强突触传递的作用不能被nAChRs拮抗剂美加明、六烃季铵和双氢-β-刺桐丁所阻断,但可被α-银环蛇毒素阻断。上述结果提示,海马脑片CAl区锥体神经元兴奋性突触前nAChRs含有对α-银环蛇毒素敏感的胡亚单位,其激活可增强海马CAl区锥体神经元突触前递质谷氨酸的释放,从而对兴奋性突触传递发挥调控作用。     

视皮层LTP维持阶段的突触形态计量学研究

本实验使用１８～２０ｄ的幼年大鼠视皮层脑片标本,在ＬＴＰ出现后３ｈ取局部微脑片固定进行ＬＴＰ维持阶段超微结构的研究。分别与孵育相同时间而未予任何刺激的脑片和仅给予测试刺激的脑片作比较。运用图像分析仪分别对三组电镜结果进行以下参数的测量：（１）突触间隙的宽度;（２）突触后致密物（ＰＳＤ）的厚度;（３）活性区的长度;和（４）突触界面曲率。用双盲法对突触数目进行计量,并用立体计量学方法对各种突触类型进行定量,所得数据用方差分析进行统计学处理。结果显示：（１）ＬＴＰ形成后１５ｈ左右,其反应达到峰值,然后维持在最高水平一直到３ｈ仍无下降趋势;（２）突触间隙的宽度较两个对照组明显增宽;（３）ＰＳＤ的厚度也明显增厚;（４）活性区的面密度及突触界面曲率明显增加;（５）总突触数目和棘突触数目的数密度较空白对照明显增高;（６）穿孔性突触的数密度与对照组相比明显增加。结果提示：活性区面密度的增加及突触界面曲率的增大可能是ＬＴＰ维持的形态学基础。穿孔性突触的形成与ＬＴＰ的维持密切相关。     

Prenatal Nicotine and Maternal Deprivation Stress De-Regulate the Development of CA1, CA3, and Dentate Gyrus Neurons in Hippocampus of Infant Rats

Adverse experiences by the developing fetus and in early childhood are associated with profound effects on learning, emotional behavior, and cognition as a whole. In this study we investigated the effects of prenatal nicotine exposure (NIC), postnatal maternal deprivation (MD) or the combination of the two (NIC+MD) to determine if hippocampal neuron development is modulated by exposure to drugs of abuse and/or stress. Growth of rat offspring exposed to MD alone or NIC+MD was repressed until after weaning. In CA1 but not CA3 of postnatal day 14 (P14) pups, MD increased pyramidal neurons, however, in dentate gyrus (DG), decreased granule neurons. NIC had no effect on neuron number in CA1, CA3 or DG. Unexpectedly, NIC plus MD combined caused a synergistic increase in the number of CA1 or CA3 neurons. Neuron density in CA regions was unaffected by treatment, but in the DG, granule neurons had a looser packing density after NIC, MD or NIC+MD exposure. When septotemporal axes were analyzed, the synergism of stress and drug exposure in CA1 and CA3 was associated with rostral, whereas MD effects were predominantly associated with caudal neurons. TUNEL labeling suggests no active apoptosis at P14, and doublecortin positive neurons and mossy fibers were diminished in NIC+MD relative to controls. The laterality of the effect of nicotine and/or maternal deprivation in right versus left hippocampus was also analyzed and found to be insiginificant. We report for the first time that early life stressors such as postnatal MD and prenatal NIC exposure, when combined, may exhibit synergistic consequences for CA1 and CA3 pyramidal neuron development, and a potential antagonistic influence on developing DG neurons. These results suggest that early stressors may modulate neurogenesis, apoptosis, or maturation of glutamatergic neurons in the hippocampus in a region-specific manner during critical periods of neurodevelopment.     

脊髓背角Ⅱ板层长时程增强诱导及维持过程中的突触形态计量学研究

本研究和体视学方法探讨了在C纤维诱发电位长时程增强(long—-term potentiation,LTP)的诱导及维持过程中的脊髓背角Ⅱ板层的突触形念变化。结果显示(1)在LTP形成后30min,Ⅱ板层内的突触后致密物质(postsynaptic density,PSD)增厚,突触间隙增宽;(2)在LTP形成后3h,PSD厚度、突触间隙宽度及突触界面曲率都有明显增加;(3)在LTP诱导和维持全过程中,总突触的数密度比对照组有明显增高。(4)在LTP形成后3h和5h,穿孔性突触的数密度与对照组比较有明显增高。上述结果显示：PSD增厚是LTP诱导阶段的主要形态学变化。突触界面曲率增人及穿孔突触数目增多是LTP维持阶段的主要形态学基础。     

Prenatal choline supplementation attenuates spatial learning deficits of offspring rats exposed to low-protein diet during fetal period

Prenatal intake of choline has been reported to lead to enhanced cognitive function in offspring, but little is known about the effects on spatial learning deficits. The present study examined the effects of prenatal choline supplementation on developmental low-protein exposure and its potential mechanisms. Pregnant female rats were fed either a normal or low-protein diet containing sufficient choline (1.1 g/kg choline chloride) or supplemented choline (5.0 g/kg choline chloride) until delivery. The Barnes maze test was performed at postnatal days 31–37. Choline and its metabolites, the synaptic structural parameters of the CA1 region in the brain of the newborn rat, were measured. The Barnes maze test demonstrated that prenatal low-protein pups had significantly greater error scale values, hole deviation scores, strategy scores and spatial search strategy and had lesser random search strategy values than normal protein pups (all P<.05). These alterations were significantly reversed by choline supplementation. Choline supplementation increased the brain levels of choline, betaine, phosphatidylethanolamine and phosphatidylcholine of newborns by 51.35% (P<.05), 33.33% (P<.001), 28.68% (P<.01) and 23.58% (P<.05), respectively, compared with the LPD group. Prenatal choline supplementation reversed the increased width of the synaptic cleft (P<.05) and decreased the curvature of the synaptic interface (P<.05) induced by a low-protein diet. Prenatal choline supplementation could attenuate the spatial learning deficits caused by prenatal protein malnutrition by increasing brain choline, betaine and phospholipids and by influencing the hippocampus structure.     

高低频强直刺激诱导大鼠视皮层长持续长时程增强后突触超微结构的变化

在发育期大鼠视皮层上以2与100 Hz强直刺激诱导长持续长时程增强(long-lasting long-term potentiation,L-LTP),然后观察突触超微结构的变化.在L-LTP形成后,运用电子显微镜及图像分析技术分析突触形态的变化.实验中观察到,突触界面曲率、突触数密度以及突触后致密物厚度在2与100 Hz组较对照组均显著增加,而突触间隙宽度减小.在100 Hz强直刺激诱导L-LTP组中,单位体积的活性区面积显著增加.100 Hz强直刺激诱导L-LTP组较2 Hz强直刺激诱导L-LTP组中单个突触活性区的面积大.以上结果表明100 Hz强直刺激诱导L-LTP组中新形成的突触较2 Hz强直刺激诱导L-LTP组中的突触大,提示100 Hz强直刺激引起的L-LTP可能伴随有突触后细胞骨架蛋白重组或合成的增加.     

Effects of estradiol benzoate on learning-memory behavior and synaptic structure in ovariectomized mice

There is increasing evidence that estrogen is involved in CNS activity, particularly memory. Several studies have suggested that estrogen improves memory by altering neuronal plasticity, including increased hippocampus CA1 dendritic spine density and enhanced long-term potentiation (LTP). In the present study, we investigated the effects of estrogen on the ultrastructural modifications in cerebral frontal cortex and hippocampus of female ovariectomized mice. One week after ovariectomy (Ovx), ICR female mice received daily injection of estradiol benzoate (EB, 20, 100, 200 microg/kg, s.c.) for 4-5 weeks. Spatial memory was then tested in the water maze, and the overall locomotor activity was monitored in open field. Synaptic morphologic parameters were examined using a graph analyzer. The results from open field did not show any alterations in locomotor activity following Ovx and EB replacement. Both the latency to find the platform and the distance to reach the platform were significantly reduced in Ovx mice by EB at 20 or 100 microg/kg when compared to vehicle treated Ovx mice. The results from synaptic ultrastructural measurement and analysis did not show any differences in hemispheric or hippocampal volumes, the numeric synaptic density, the length of active zones, or the curvature of synaptic interface among Sham, Ovx, and Ovx plus EB replacement mice. However, EB replacement effectively normalized the changes induced by Ovx, reducing the width of the synaptic cleft, enlarging the thickness of postsynaptic density (PSD), and increasing the number of synaptic vesicles in the presynapse in both cerebral cortex Fr1 and hippocampus CA1 areas. These results suggest that the beneficial effects of EB on improving memory behavior of Ovx female mice are associated with the changes of some subtle structural parameters of synapses, including the width of PSD and synaptic cleft rather than some basic and permanent structure in frontal cortex and hippocampus regions.     

Effects of strong static magnetic fields used in magnetic resonance imaging on insulin-secreting cells

The magnetic flux density of MRI for clinical diagnosis has been steadily increasing. However, there remains very little biological data regarding the effect of strong static magnetic fields (SMFs) on human health. To evaluate the effects of strong SMFs on biological systems, we cultured insulin-secreting cells under exposure to sham and SMF conditions (3-10 T of magnetic flux density, and 0-41.7 T/m of magnetic field gradient) for 0.5 or 1 h, and analyzed insulin secretion, mRNA expression, glucose-stimulated insulin secretion, insulin content, cell proliferation and cell number. Exposure to SMF with a high magnetic field gradient for 1 h significantly increased insulin secretion and insulin 1 mRNA expression. Exposure to SMF with a high magnetic flux density for 0.5 h significantly enhanced responsiveness to glucose stimulation. Exposure to SMF did not affect the insulin content, cell proliferation or cell number. Our results suggested that MRI systems with a higher magnetic flux density might not cause cell proliferative or functional damages on insulin-secreting cells, and that SMF with a high magnetic field gradient might be used clinically after thorough in vivo investigations are conducted.