肝性脑病大鼠海马齿状回神经元形态及一氧化氮合酶表达的变化

目的:观察肝性脑病模型组大鼠海马齿状回内神经元的变化及一氧化氮合酶(NOS)的表达,探讨海马神经元的形态学改变及一氧化氮(NO)在肝硬化和肝性脑病发病机制中的作用。方法:先对50只雄性大鼠进行Morris水迷宫测试,之后将动物分为正常对照组和实验模型组。9周后建立CCL4肝性脑病模型,分别取两组大鼠肝、海马组织进行HE染色、Nissl染色及NADPH-d染色。结果:(1)肉眼下可见模型组肝脏普遍呈坏死性肝硬化;(2)HE模型组血氨浓度明显高于正常对照组(P<0.05);(3)Nissl染色结果显示实验组大鼠海马神经元数目减少、染色较浅,胞浆内Nissl体减少或消失;(4)NADPH-d染色结果显示实验组可见粗大轴突着色,树突联系广泛;对照组则少有粗大轴突着色,树突间联系不如实验组广泛。实验组一氧化氮合酶(NOS)阳性神经元染色较对照组深,为紫蓝或深蓝色,且阳性神经元的数目较多。结论:(1)血氨增高是肝性脑病发病机制之一;(2)肝性脑病时海马受到损伤,并且一氧化氮(NO)可能介导了神经元的损伤。     

脑室内注射BDNF抗体对大鼠海马NOS表达的影响

探讨脑室内注射脑源性神经营养因子 (BDNF)抗体阻断内源性BDNF对大鼠海马一氧化氮合酶 (NOS)阳性神经元的影响。脑室内注射BDNF抗体一周后 ,采用Morris水迷宫进行行为检测 ;并用NADPH 黄递酶组化染色方法观察海马NOS阳性神经元数目的变化。与对照组相比 ,实验组大鼠空间学习和记忆能力明显下降 (P <0 0 1) ;实验组大鼠海马CA1区NOS阳性神经元数目 (38 37± 5 2 3)明显少于对照组 (4 9 5 3± 5 74 ) (P <0 0 1) ;实验组DG区NOS阳性神经元数目 (4 8 77± 5 5 1)明显少于对照组 (6 0 4 0± 7 39) (P <0 0 1)。脑室内注射BDNF抗体可导致大鼠空间学习记忆能力下降 ,海马NOS阳性神经元数目减少 ,提示BDNF对学习和记忆的影响可能与海马NOS阳性神经元数目的变化有关     

急性缺氧小鼠海马CA1区NOS活力和nNOS蛋白表达的时程变化

目的观察急性缺氧小鼠海马CA1区一氧化氮合酶(NOS)和神经元型一氧化氮合酶(nNOS)阳性神经元的时程变化,探讨NO在脑缺氧中的作用并为抗脑缺氧提供依据.方法复制小鼠急性缺氧模型,采用NADPH-d组织化学和nNOS免疫组织化学方法,研究急性缺氧后不同时程点小鼠海马CA1区NADPH-d和nNOS阳性神经元数量的变化.结果与正常对照组相比较,急性缺氧后0.5h组小鼠海马CA1区NADPH-d和nNOS阳性神经元的数量无明显变化,差异无显著性(P＞0.05),3h、6h和12h组逐渐增多并于12h升高达到最高峰,差异有显著性(P＜0.05),而于24h后开始降低,48h恢复正常.结论急性缺氧后早期海马CA1区NOS和nNOS水平明显增多,NO在缺氧所致早期脑损伤中起重要作用.     

大鼠端脑内一氧化氮合酶阳性神经元的发育

目的研究大鼠胚胎时期及生后早期一氧化氮合酶（NOS）阳性神经元在端脑的分布，探讨一氧化氮（NO）在脑发育过程中的作用。方法应用还原型尼克酰胺腺嘌呤二核苷酸磷酸脱氢酶（NADPH-d）组织化学方法观察孕14d起至生后14d大鼠端脑内NOS阳性神经元的形态和分布。结果孕14d没有观察到阳性神经元。孕15d纹状体腹外侧已有NOS阳性表达。孕17d在大脑皮质、梨状皮质观察到NOS阳性神经元，但胞体小，树突短，且分支少。随着年龄的增长神经元的胞体数目增多、染色增强或维持一定的水平。到孕20d，NOS阳性神经元分布广泛，梨状皮质、纹状体腹外侧及终纹床核均有大量NOS阳性神经元，其胞体明显增大，树突分支复杂化，长度增加。在生后，除上述脑区的阳性神经元进一步发育分化，大脑皮质和纹状体的NOS阳性纤维相互编织成疏密不等的纤维网外，在胼胝体、海马也观察到NOS阳性神经元。到生后14d，NOS阳性神经元的分布模式总体上已与成年大鼠相似。结论NOS阳性神经元在端脑独特的表达模式提示NO在脑发育和成熟过程中扮演重要角色。     

大鼠海马和颞叶一氧化氮合酶阳性神经元在学习记忆时的表达

目的　对获得空间辨别性学习记忆大鼠的海马结构和颞叶皮质内一氧化氮合酶 (NOS)免疫阳性神经元系统的观察 ,为一氧化氮 (NO)在学习记忆活动中的作用提供形态学依据。方法　以经水迷宫训练获得空间辨别性学习记忆的大鼠为模型组 ,建立游水对照组和空白对照组。用免疫组化的方法观察 3组大鼠海马和颞叶的NOS阳性神经元的形态学特征及测定免疫反应阳性产物的光密度值 (OD )。结果　 (1)NOS阳性神经元在 3组大鼠海马和颞叶呈散在分布。在海马CA1、CA2、CA3、CA4及龄状回 (DG)都有少量分布 ,颞叶以Ⅱ、Ⅲ、Ⅴ、Ⅵ层分布为主。神经元胞体形态有锥体形 ,圆形或椭圆形 ,梭形3种。 (2 )模型组大鼠海马和颞叶皮质NOS阳性神经元较对照组明显增多。 (3)其免疫反应阳性产物的OD值也较对照组明显增大。结论　NOS阳性神经元形态发生了可塑性变化 ,参与了学习记忆     

急性缺氧小鼠海马CAl区NOS活力和nNOS蛋白表达的时程变化

目的:观察急性缺氧小鼠海马CAl区一氧化氮合酶(NOS)和神经元型一氧化氮合酶(nNOS) 阳性神经元的时程变化,探讨NO在脑缺氧中的作用并为抗脑缺氧提供依据。方法:复制小鼠急性缺氧模型,采用NADPH-d组织化学和nNOS免疫组织化学方法,研究急性缺氧后不同时程点小鼠海马CAl区NADPH-d 和nNOS阳性神经元数量的变化。结果:与正常对照组相比较,急性缺氧后0.5h组小鼠海马CAl区NADPH-d 和nNOS阳性神经元的数量无明显变化,差异无显著性(P>0.05),3h、6h和12h组逐渐增多并于12h升高达到最高峰,差异有显著性(P<0.05),而于24h后开始降低,48h恢复正常。结论:急性缺氧后早期海马CAl区NOS和nNOS水平明显增多,NO在缺氧所致早期脑损伤中起重要作用。     

L-精氨酸甲酯对严重烫伤大鼠回肠NOS阳性神经元的影响

目的 了解一氧化氮合酶抑制剂L-精氨酸甲酯(L-NAME)对大鼠严重烫伤后回肠肌间神经丛内NOS阳性神经元的影响。方法50只SD大鼠随机分为正常组、烫伤组和烫伤+L-NAME组,应用NADPH-d酶组织化学染色观察回肠肌间神经内NOS阳性神经元数量和阳性反应面积。结果大鼠体表烫伤后3d,回肠NOS阳性神经元数目明显增加,染色呈强阳性,阳性反应面积增加。L-NAME可降低回肠肌问神经丛内NOS阳性神经元数目,阳性反应面积减少,染色较淡。结论烫伤可引起大鼠回肠内NOS活性增加,L-NAME可能通过降低NOS活性,从而减轻烫伤引起的神经元损伤。     

一氧化氮与利血平致大鼠胃粘膜损伤的关系

目的: 探讨一氧化氮(NO)与利血平致大鼠胃粘膜损伤机制的关系。方法: 从大鼠腹腔内注入利血平复制成胃粘膜损伤模型,并随机分为实验组和对照组。用镉还原加格氏显色法和改良硫代巴比妥酸法分别测定两组大鼠血浆及胃粘膜中NO和丙二醛(MDA)含量,用还原型辅酶Ⅱ组织化学方法结合图像分析对两组大鼠胃壁内一氧化氮合酶(NOS)进行定位、定量研究。结果: 实验组大鼠血浆、胃粘膜中NO含量显著低于对照组(P＜0.01),而其MDA含量则显著高于对照组(P＜0.05,P＜0.01);实验组大鼠胃壁内NOS阳性神经细胞、阳性神经纤维密度及其着色深度均显著低于对照组(P＜0.05,P＜0.01)。结论: 利血平致大鼠胃粘膜损伤的发生可能与大鼠胃壁内NOS阳性神经成分减少,体内NO不足使其对胃粘膜的保护作用减弱有一定关系。     

胸腺及其投射神经元一氧化氮的分布

本实验用 NADPH-d组织化学方法及 n NOS免疫细胞化学方法 ,对大鼠胸腺内一氧化氮 (NO)阳性细胞的分布进行了研究。采用 CB-HRP逆行追踪结合 n NOS免疫细胞化学双重反应技术 ,观察了大鼠胸腺投射神经元 n NOS的分布。结果显示 :(1)在脑干的疑核、面后核内有 CB-HRP与 n NOS双重阳性细胞 ;(2 )在胸腺内有多种 NADPH-d和 n NOS阳性细胞 ,按其形态可分为 :髓质上皮样细胞、胸腺树突样细胞、神经元样细胞、胸腺细胞样细胞及胸腺小体 ;(3 )在胸腺的被膜下、小梁内、皮髓质交界处、小血管的周围有丰富的 n NOS阳性纤维。提示 ,胸腺内 NO的来源不同 ,其在调节胸腺的各种活动中可能发挥的作用也不同     

长时间应用L-精氨酸增强大鼠学习记忆功能和大脑皮质、海马nNOS或c-fos的表达及神经元数目的增加(英文)

为探讨长时间应用一氧化氮(NO)对学习记忆功能和神经系统可塑性的影响,以及NO与c-fos基因表达的关系,分别给初断乳的大鼠灌胃NO前体左旋-精氨酸(L-Arg)或一氧化氮合酶抑制剂L-NAME,用Morris水迷宫检测大鼠的学习记忆功能,用免疫组化技术和HE染色检测大脑皮质和海马CA1、CA3、DG区的神经元型一氧化氮合酶(nNOS)和c-fos基因的表达以及神经细胞数目的变化。将大鼠随机分成L-Arg组、L-NAME组和对照组。大鼠断乳后分别每天用L-Arg[200mg/(kg.d)]、L-NAME[50mg/(kg.d)]和等剂量的蒸馏水灌胃,持续3个月。结果显示:长期应用NO前体L-Arg可明显缩短大鼠的寻台潜伏期,促进nNOS和c-fos基因的表达,同时大脑皮质和海马CA1、CA3、DG区的神经元数目增加;而长期应用一氧化氮合酶抑制剂L-NAME可抑制大鼠寻台潜伏期的缩短和nNOS、c-fos基因的表达,同时大脑皮质和海马CA1、CA3、DG区的神经元数目减少。因此我们认为长时间应用NO可促进神经系统c-fos基因的表达和幼鼠神经系统的发育,即可塑性的变化,继而影响大鼠的学习记忆功能。     

Ultrastructural localisation of NADPH-d/nNOS expression in the superior cervical ganglion of the hamster

This study examined NADPH-d and nNOS expression in the SCG of hamsters. By light microscopy, numerous NADPH-d/NOS positive processes were widely distributed in the ganglion. Ultrastructurally, the NADPH-d reaction product was associated with the membranous organelles of neuronal soma, dendrites, myelinated fibres, small granular cells, and axon profiles bearing agranular vesicles. The NOS immunoreaction product, on the other hand, was localised in the cytoplasm of principal neurons and dendrites. Some of the NADPH-d/NOS labelled processes formed junctional contacts including synapses or zonulae adherentia. Compared with the neurons, the nonneuronal cells in the ganglion, namely, macrophages, satellite cells and endothelial cells were labelled by NADPH-d but devoid of nNOS immunoreaction product. The results suggest that the NADPH-d/NOS positive fibres in the SCG originate not only from the projecting fibres of the lateral horns of thoracic spinal cord, but also from the principal neurons and small granular cells; some may represent visceral afferent fibres. Electron microscopic morphometry has shown that about 67% of the principal neurons contain NADPH-d reaction product, and that the majority were small to medium sized neurons based on cross-sectional areas in image analysis. On the basis of the present morphological study, it is concluded NO is produced by some local neurons and possibly some nonneuronal cells in the SCG as well as some fibres of extrinsic origin. In this connection, NO may serve either as a neurotransmitter or neuromodulator.     

Nitrergic neurons during early postnatal development of the prefrontal cortex in the rat: Histochemical study

The presence of nitrergic cells in the prefrontal cortex has been confirmed, however little is known about the postnatal development of these cells. Nitrergic neurons were studied histochemically by using NADPH-diaphorase staining in the prefrontal cortex of male Wistar rats from postnatal day 7–21 (P7–21). Neuronal NADPH-diaphorase is a nitric oxide synthase that provides a specific histochemical marker for neurons producing nitric oxide (NO). NO acts as a neurotransmitter and intracellular signaling molecule in the nervous system. We observed in 7 day old rats NADPH-d containing neurons that were intensely stained. These neurons were bipolar with a short dendrite with average length of 23 μm. During the second postnatal week, the neurons were mainly bipolar and were rarely multipolar. By P14 the cells were located primarily in cortical layers III–VI. Nitrergic neurons of the 21 day old rats were histochemically identified as multipolar cells with long radial extending dendrites. Dendrites of neurons in 14 and 21 day old rats were a similar length with an average of 57 μm. These results suggest that nitrergic neurons differentiate during a relatively short period of time and reach their structural maturity by the end of the second week of postnatal development.     

PKC抑制剂灯盏花素乙对甲醛炎性痛大鼠脊髓NOS表达及NO含量的影响

目的：观察蛋白激酶C（PKC）选择性抑制剂灯盏花素乙（CH）对甲醛炎性痛时大鼠自发痛反应、脊髓一氧化氮合酶（NOS）表达和一氧化氮（NO）含量的影响，探讨炎性痛时脊髓内NO产生是否受PKC调控。 方法： 采用右后掌足底注射甲醛复制炎性痛模型；计数缩足反射次数反映自发痛程度；应用NADPH-d组织化学法测定脊髓NOS表达；硝酸/亚硝酸还原法测定脊髓腰膨大部位NO2-/NO3-含量。 结果： 甲醛炎性痛大鼠L5脊髓后角浅层和中央管周围灰质NADPH-d阳性细胞的数目、阳性细胞胞体及纤维的染色深度均明显大于正常对照组，脊髓腰膨大部位 NO2-/NO3-含量明显增高。预先鞘内给予CH，可明显抑制甲醛炎性痛诱导的大鼠第二相自发痛反应以及脊髓NOS表达和NO2-/NO3-的含量。 结论： 炎性痛时，脊髓伤害性感受神经元内PKC激活可以促进NOS表达和NO的产生。     

大鼠创伤性脑损伤后细胞凋亡及NOS阳性细胞的变化

目的：探讨大鼠创伤性脑损伤后不同时相皮质、海马、隔区细胞凋亡及NOS、ChAT阳性细胞的变化。方法：采用大鼠自由落体脑损伤模型，伤后1、2、3、4、5、7、10d取脑切片，经Nissl染色，用TUNEL法检测细胞凋亡，NADPH—d组化染色观察NOS阳性细胞，ChAT免疫组化染色观察隔区ChAT阳性细胞。结果：Nissl染色可见损伤侧海马CA2、CA3区锥体细胞层细胞消失或紊乱。损伤区周围皮质凋亡细胞伤后3d达到高峰；损伤侧海马凋亡细胞伤后5d达到高峰；损伤侧隔区凋亡细胞7d达到高峰。正常侧上述脑区各时相点均未见到凋亡细胞。损伤区周围皮质、损伤侧海马和隔区iNOS阳性细胞数量明显增加。损伤侧隔区ChAT阳性神经元也明显减少。结论：大鼠创伤性脑损伤后损伤区周围皮质和损伤侧海马、隔区细胞凋亡数量的变化与伤后时程有关。伤后细胞iNOS表达增加是导致细胞凋亡的因素。     

Mild hypoxic preconditioning attenuates injury-induced NADPH-d/nNOS expression in brainstem motor neurons of adult rats

Excessive production of nitric oxide (NO) might have detrimental effects on the hypoxia-related neuropathology. This study aimed to test if mild hypoxic preconditioning (MHPC) would attenuate the pathological changes in the brainstem motoneurons having a different functional component after peripheral nerve crush injury (PNCI). Prior to PNCI treatment, young adult rats were caged in the mild hypoxic altitude chamber with 79Torr of the partial oxygen concentration ( pO(2)) (i.e., 0.5atm at 5500m in height) for 4 weeks to adapt the environmental changes. After that, all the animals having successfully crushed both the hypoglossal and vagus nerves (left-side) were allowed to survive for 3, 7, 14, 30 and 60 successive days in normoxic condition. Nicotinamine adenine dinucleotide phosphate-diaphorase (NADPH-d) histochemistry and neuronal nitric oxide synthase (nNOS) immunohistochemistry revealed that MHPC reduces NADPH-d/nNOS expression in the hypoglossal nucleus (HN) and the dorsal motor nucleus of the vagus (DMN) at different time points after PNCI. The morphological findings were further ascertained by Western blot analysis of nNOS and nitrite assay for NO production. Both the morphological and quantitative results peaked at 7 days in HN, whereas for those in DMN were progressively increased up to 60 days following PNCI. The staining intensity of NADPH-d/nNOS(+) neurons, expression of nNOS protein, NO production levels as well as the neuronal loss in HN and DMN of MHPC rats following PNCI were attenuated, especially for those having a longer survival period over 14 days. The MHPC treatment might induce minute amounts of NO to alter the state of milieu of the experimental animals to protect against the PNCI.     

Distribution of NADPH diaphorase-containing neurons in the pigeon central nervous system

The aim of the present study was to determine the distribution of nitric oxide-synthesizing neurons in the pigeon brain and spinal cord. Tissue sections were stained for reduced nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d). In the telencephalon, intensely stained neurons with dendrites extending distally were seen in most regions. The ectostriatum was characterized by intensely and diffusely stained neuropil. In the diencephalon, intensely positive neurons were seen in the lateral hypothalamic region and lateral mammillary nucleus. In the mesencephalon, intensely stained, multipolar neurons were abundantly scattered in the central gray, nucleus intercollicularis, reticular formation, nucleus tegmenti pedunculo-pontinus, pars compacta, area ventralis of Tsai, and ansa lenticularis. In the rhombencephalon, positively-stained neurons were found in the pontine nuclei and reticular formation. The cerebellar cortex, except for Purkinje cells, was a preferential region for NADPH-d activity. Positive end-bulbs made contact on somata in the nucleus magnocellularis cochlearis. In the spinal cord, NADPH-d positive neurons were seen in layer II and the marginal nucleus. Our results demonstrated that the distribution of NADPH-d-containing neurons in the pigeon brain and spinal cord is more complex than in other avian species. Our findings indicate that NADPH-d-containing neurons are present in several sensory pathways, including olfactory, visual, auditory, and somatosensory tracts, although some nuclei in each system did not show NADPH-d activity. The wide distribution of NADPH-d activity in the pigeon CNS suggests that nitric oxide modulates sensory transmission in avian central nervous system.     

急性乙醇中毒大鼠学习记忆行为的改变与脑组织NO和nNOS含量变化

目的：通过观察急性乙醇中毒对大鼠学习记忆的影响并测定脑组织中一氧化氮（NO）与神经型一氧化氮合酶（nNOS）含量的变化，探讨乙醇中毒影响学习记忆的分子机制。方法:成年SD大鼠随机分为2组，模型组腹腔1次性注射乙醇2.5 g/kg［用生理盐水配成含20%乙醇(W/V)溶液］制备急性乙醇中毒大鼠模型；对照组注射等容量的生理盐水。Y-型迷宫检测大鼠学习记忆成绩、硝酸还原酶法检测鼠脑海马CA1、新纹状体中NO的含量、免疫组化方法检测鼠脑海马CA1、纹状体、小脑中nNOS的含量。结果:（1）模型组大鼠达到学会标准所需要的训练次数（34.33±13.04）明显大于对照组（27.50±8.79），P<0.05；（2）海马CA1区NO的含量在模型组为23.09±9.60，明显高于对照组（8.46±5.67）（P<0.01）；新纹状体（尾壳核）NO的含量在模型组（19.46±8.25）也明显高于对照组（8.22±4.46），P<0.01；（3）海马CA1区nNOS阳性神经元的数量在模型组为18.22±7.47，明显高于对照组（10.15±4.24）（P<0.05）；新纹状体（尾壳核）nNOS阳性神经元的数量在模型组（11.38±5.00）也明显高于对照组（6.15±3.69），P<0.05。结论:乙醇的神经毒性作用可能与脑组织中nNOS和 NO信号通路有关。     

大鼠大脑内血管的氮能神经支配

目的：观察大鼠大脑内氮能神经元及血管的氮能神经支配。阐述一氧化氮在脑内血管中的作用。方法：大鼠经灌流固定后用恒冷切片机漂浮切片。还原型尼克酰胺腺嘌呤二核苷酸脱氢酶（ＮＡＤＰＨ－ｄ）染色。结果：氮能神经元依形态分为双极和多极。见纤细的树突与血管紧密相连及串珠样血管周围神经丛。结论：大脑ＮＡＤＰＨ－ｄ阳性神经元纤细的树突与脑内血管密切相关。提示一氧化氮 控脑血管张力和血液循环方面有重要作用。