豚鼠小肠神经节丛的NADPH—黄递酶组织化学观察

目前已知,ＮＡＤＰＨ－－黄递酶组化法可选择性地显示－－氧化氮合成酶（ＮＯｓｙｎｔｈａｓｅ,ＮＯＳ）神经元。因此,我们以ＮＡＤＰＨ－黄递酶组化法,观察了豚鼠小肠肌间神经丛和粘膜下神经丛的神经网格以及ＮＯＳ神经元。结果表明,三段小肠肌间神经丛的神经网眼大小和形态有明显差异,与对应的粘膜下神经丛相比,差异更显著。在肌间神经丛中,ＮＡＤＰＨ－黄递酶阳性神经元胞体大小不等;其长突起伸入节间束,而短突起较多,并可见短突起彼此连接．构成节内偶见的局部神经元回路。从小肠上段到下段,ＮＯＳ神经元数量呈下降趋势。在粘膜下神经丛,我们也观察到少数ＮＯＳ神经元。     

人小肠粘膜内神经元的组织学与组织化学观察

一般认为,胃肠道粘膜内不存在神经元,粘膜神经丛仅由神经纤维组成。本文使用常规组织学方法和 NADH 黄递酶组化法证明在人小肠粘膜内可见到单个的神经元或由2～5个神经元组成的神经节。神经元胞体大小不等,形态多样,除梭形、卵圆形和不规则形外,还有呈扁平形的。粘膜内神经元多数位于固有层内,常紧贴粘膜肌;粘膜肌层内也可见到神经元。我们认为,小肠粘膜内神经元是肠神经系中粘膜神经丛的一部分。     

大鼠胃肌间神经丛中NOS阳性神经元的组织化学研究

采用ＮＡＤＰＨ┐黄递酶（ＮＤＰ）组织化学技术在整装铺片上对大鼠胃肌间神经丛中一氧化氮合酶（ＮＯＳ）阳性神经元进行定量和定位研究。结果显示：大鼠胃肌间神经丛中ＮＯＳ阳性神经元密度（个／ｍｍ２）分别为：６２．１±３７．７（幽门部）、４３．４±３１．７（胃体）、３１．６±２７．８（胃底）。胃底部神经节内神经元数量较少,神经元胞体较大,染色较深;幽门部神经节内神经元数量较多,胞体大小不等,染色深浅不一;胃体部的则介于两者之间,呈过渡态。结果表明：胃各部肌间神经丛中ＮＯＳ阳性神经元的差异可能与胃的生理功能密切相关     

扬子鳄小肠壁神经丛内含AChE及SOM神经元的组织化学和免疫组织化学观察

本实验分别应用显示乙酰胆碱酯酶（ＡＣｈＥ）和生长抑素（ＳＯＭ）的组织化学和免疫组织化学方法,对扬子鳄小肠神经丛进行了研究。结果显示;ＡＣｈＥ阳性神经纤维广泛分布于肠壁神经丛内;粘膜下丛和肌间丛可见到许多ＡＣｈＥ阳性神经元,胞体大小不等形态不一。生长抑素免疫反应（ＳＯＭ－ＩＲ）纤维主要分布在肌间神经丛,ＳＯＭ－ＩＲ纤维大体呈两种类型：即较多的膨体状神经纤维和少量的细长神经纤维。粘膜下丛和肌间丛可见ＳＯＭ－ＩＲ阳性神经元,有时尚可见突起与周围神经纤维联系。提示低等脊柱动物两栖爬行类一扬子鳄内脏已存在比较完善的神经调节。     

大鼠食管胸段，腹段乙酰胆碱酯酶阳性神经的配布

大鼠食管胸段和腹段壁内乙酰胆碱酯酶（ＡＣｈＥ）阳性神经存在于神经束和分支的粗细神经纤维内,也见于外膜丛,肌间丛,粘膜下丛和粘膜肌内。食管肌层内ＡＣｈＥ阳性神经纤维多而密集,而食管腹段肌内尤为丰富,肌间神经纤维末梢分布于肌束表面,可能与控制肌纤维活动有关;分布于肌内,粘膜下层和上皮基部的ＡＣｈＥ阳性神经中,尚含有内脏感觉神经纤维。食管壁的肌间丛和粘膜下丛内散在有多极形和卵园形的ＡＣｈＥ阳性神经元,在食管腹段内数多,而以中小型神经元为主。     

大鼠内侧隔核、斜角带中NOS与ChAT、NGF-R、AChE的共存神经元

用酶组织化学和免疫组织化学双标技术,观察了正常ＳＤ大鼠基底前脑内侧隔核（ＭＳ）、斜角带垂直支（ＶＤＢ）和水平支（ＨＤＢ）中ＮＯＳ阳性神经元的形态和分布及ＮＯＳ与胆碱能神经元标志物ＣｈＡＴ、ＮＧＦ受体（ＮＧＦ－Ｒ）和ＡＣｈＥ之间的共存关系。结果发现,ＭＳ、ＶＤＢ和ＨＤＢ的头端ＮＯＳ阳性神经元较多、胞体较大、突起多,尾端ＮＯＳ阳性神经元数目较少、胞体较小、突起少而短。ＮＯＳ＋ＣｈＡＴ双标神经元占ＮＯＳ阳性神经元总数的９０％,占ＣｈＡＴ阳性神经元总数的３９％;ＮＯＳ＋ＮＧＦ－Ｒ双标神经元占ＮＯＳ阳性神经元总数的８３％,占ＮＧＦ－Ｒ阳性神经元总数的４０％;ＮＯＳ＋ＡＣｈＥ双标神经元占ＮＯＳ阳性神经元总数的９６％,占ＡＣｈＥ阳性神经元总数的３９％。这些结果为研究Ａｌｚｈｅｉｍｅｒ＇ｓｄｉｓｅａｓｅ病理过程中基底前脑隔区胆碱能神经元退变与ＮＯ的关系提供了形态学依据。     

棕色田鼠小肠肌间神经丛NOS的组织化学观察

用NDP-黄递酶组织化学法研究了NOS在棕色田鼠（Microtus mandarinus)小肠肌间神经丛的分布,同样的方法对大白鼠进行了实验比较。结果发现,棕色田鼠小肠肌间祖辈 经丛NOS阳性神经元形态各异,大小悬殊数倍;在神经节和阳性神经纤维中可见阳性神经元呈“串珠”状和“U”型排列。10倍镜下记数100个视野,观察到阳性神经元平均密度为42.8个/mm^2,每个视野内平均有神经节12.4个,每个神经节内平均有NOS阳性神经元8.5个,占其神经节内神经元总数的27.2%,棕色田鼠为野生草食性动物,其NOS在小肠肌间神经丛的分布与其他杂食性啮齿类有相同之处,但也存在着种属差异。     

先天性巨结肠病NOS阳性神经和VIP能神经的异常改变──酶组织化学与免疫组织化学联合法的研究

本文应用还原型辅酶Ⅱ（ＮＡＤＰＨ－Ｏ）－黄递酶组织化学和血管活性肠肽（ＶＩＰ）的免疫组织化学联合染色法,对１０例先天性巨结肠病扩张段和狭窄段结肠及４例“正常对照”结肠进行观察,结果发现：（１）狭窄段结肠壁丛内均缺失含一氧化氮合酶（ＮＯＳ）和ＶＩＰ神经元胞体。（２）狭窄段结肠肌层内ＮＯＳ和ＶＩＰ阳性纤维比“正常”结肠明显减少,酶活性或免疫反应性弱。结果表明,二种神经在病变肠段痉挛狭窄的神经病理机制诸因素中起重要作用。     

国人睾丸、附睾和输精管的氧化还原酶的组织化学观察

本文收集了１９—３８岁国人正常男性新鲜睾丸、附睾和输精管１３例,进行了氧化还原酶组织化学染色、光镜定位及定性观察。结果表明：睾丸曲细精管和输出小管上皮的ＧＤＨ,ＮＡＤＨＤ,ＮＡＤＰＨＤ,ＳＤＨ,ＧＰＤＨ,ＩＣＤＨ,ＭＤＨ,ＬＤＨ和Ｇ－６－ＰＤＨ９种酶;睾丸间质细胞和附睾管上皮的ＮＡＤＨＤ,ＮＡＤＰＨＤ,ＳＤＨ,ＩＣＤＨ,ＭＤＨ,ＧＤＨ,ＬＤＨ和Ｇ－６－ＰＤＨ８种酶;输精管的ＮＡＤＨＤ,ＮＡＤＰＨＤ,ＩＣＤＨ和ＧＤＨ４种酶的酶活性呈强阳性或极强阳性。提示输出小管和头部附睾管含有的多种氧化还原酶对精子功能成熟有极重要作用。     

骤冷与饥饿对小鼠胃影响的实验研究

当机体受饥饿与寒冷等因素刺激后, 由于应激反应, 可导致物质代谢的变化, 并造成机体损害。为探讨饥饿及饥饿与骤冷对动物胃的影响,本实验用健康昆明种小鼠２５ 只,随机分成正常组５ 只,饥饿组１０只, 饥饿后再予冷刺激组１０ 只（下称骤冷组）。采用组织化学及酶组织化学方法观察糖原（ＰＡＳ反应）,ＳＤＨ（琥珀酸脱氢酶）, ＬＤＨ （乳酸脱氢酶）, ＣｈＥ （胆碱酯酶）, Ｍｇ２＋ －ＡＴＰａｓｅ （镁激活三磷酸腺苷酶）, ＭＡＯ（单胺氧化酶）。结果显示： １． 饥饿时胃粘膜层ＰＡＳ反应, ＳＤＨ、ＣｈＥ活性显著下降, 肌间神经丛ＭＡＯ活性增强。２． 饥饿后骤冷时胃粘膜层出血, 胃粘膜ＰＡＳ反应、ＳＤＨ 活性更显著下降, 而肌间神经丛的ＣｈＥ、ＭＡＯ 活性明显增强。     

Nitric oxide synthase immunoreactivity in the enteric nervous system of the developing human digestive tract

We have investigated indirectly the presence of nitric oxide in the enteric nervous system of the digestive tract of human fetuses and newborns by nitric oxide synthase (NOS) immunocytochemistry and nicotinamide adenine dinucleotide phosphate diaphorase (NADPHd) histochemistry. In the stomach, NOS immunoactivity was confined to the myenteric plexus and nerve fibres in the outer smooth musculature; few immunoreactive nerve cell bodies were found in ganglia of the outer submucous plexus. In the pyloric region, a few nitrergic perikarya were seen in the inner submucous plexus and some immunoreactive fibres were found in the muscularis mucosae. In the small intestine, nitrergic neurons clustered just underneath or above the topographical plane formed by the primary nerve strands of the myenteric plexus up to the 26th week of gestation, after which stage, they occurred throughout the ganglia. Many of their processes contributed to the dense fine-meshed tertiary nerve network of the myenteric plexus and the circular smooth muscle layer. NOS-immunoreactive fibres directed to the circular smooth muscle layer originated from a few NOS-containing perikarya located in the outer submucous plexus. In the colon, caecum and rectum, labelled nerve cells and fibres were numerous in the myenteric plexus; they were also found in the outer submucous plexus. The circular muscle layer had a much denser NOS-immunoreactive innervation than the longitudinally oriented taenia. The marked morphological differences observed between nitrergic neurons within the developing human gastrointestinal tract, together with the typical innervation pattern in the ganglionic and aganglionic nerve networks, support the existenc of distinct subpopulations of NOS-containing enterice neurons acting as interneurons or (inhibitory) motor neurons.     

Some morphological and histochemical features of the midgut myenteric plexus of the common European frog, Rana esculenta.

The neuron morphology and distribution of four putative transmitters were investigated in the myenteric plexus of frog (Rana esculenta) midgut. The gross morphology was revealed by NADH-diaphorase histochemistry, and the shape of the neurons by silver impregnation. Nerve cells had heterogeneous distribution: they either formed ganglia or placed as solitary neurons in the duodenum, while in the rest of the midgut only solitary neurons were observed. Three morphologically distinct cell types were revealed by silver impregnation: mainly type I and type II neurons cells were seen in the duodenum, while the rest of the intestine contained type II and III cells. Catecholamine fluorescence was revealed in nerve fibres in the duodenum, while few small nerve cells were observed in the small intestinal region. Acetylcholinesterase histochemistry showed strongly reactive nerve cells that were associated with the main fibre bundles in the duodenum. Only longitudinally oriented fibres and occasionally stained neurons were seen in the small intestine. Substance P immunocytochemistry revealed an extensive plexus, which contained a moderate number of stained perikarya in the full length of the midgut. Gamma-aminobutyric acid showed non-uniform distribution in the two parts of the midgut: a stronger and more regular fibre staining was found in the duodenum then in the rest of the intestine. Ultrastructural observations demonstrated that intrinsic neurons received synaptic inputs from the profiles contained agranular vesicles, while "P"-type profiles established close contacts with neurons. Both profile types formed close contacts with the smooth muscle cells.(ABSTRACT TRUNCATED AT 250 WORDS)     

Distribution, morphology and projections of nitrergic and non-nitrergic submucosal neurons in the pig small intestine

 Sequential nitric oxide synthase immunohistochemistry and nicotinamide adenine dinucleotide phosphate diaphorase (NADPHd) histochemistry in pig small intestinal wholemounts revealed a complete colocalisation of the two nitrergic markers in submucous neurons. The external submucous plexus (ESP) contained nitrergic neurons throughout. In the internal submucous plexus (ISP) we found a moderate number of nitrergic neurons in the duodenum, while they were rare in the jejunum and nearly absent in the ileum. Combined NADPHd histochemistry and silver impregnation showed morphological ESP type III and VI neurons to be NADPHd positive whereas ESP type II, IV and V neurons were NADPHd negative. Axons of ESP type III, IV and VI neurons were often observed to enter interconnecting strands directed abluminally. ESP type II neurons projected mainly to the ISP. In special silver-impregnated wholemounts containing both external muscle layers and the abluminal part of the submucous layer, i.e. the myenteric plexus and the ESP, the great majority of impregnated axons within the interconnecting strands were observed to run between both plexuses and did not enter the circular muscle layer. We conclude that ESP type III and VI neurons are nitrergic while ESP type II, IV and V neurons are non-nitrergic. Furthermore, we assume that ESP type III, IV and VI neurons may represent a submucosal input to the myenteric plexus. Accepted: 26 August 1997     

甘肃鼢鼠胃肠道肌间神经丛一氧化氮合酶阳性神经元的分布

用NADPH-黄递酶组织化学法及整装铺片技术,对甘肃鼢鼠(Myospalax cansus)胃肠道肌间神经丛NOS阳性神经元的分布进行研究.结果显示,甘肃鼢鼠胃肠道肌间神经丛NOS阳性神经元分布广泛,形态多样,神经元大小不同,阳性神经节与阳性神经纤维束形成网络;胃肠道不同部位NOS阳性神经元密度有差异,结肠最高,直肠次之.从十二指肠至回肠段,NOS阳性神经元密度整体呈上升趋势;胃与空肠、十二指肠与盲肠间NOS阳性神经元密度无显著差异,其他各段之间差异显著.甘肃鼢鼠胃肠道肌间神经丛NOS阳性神经元分布与其他已研究动物的分布模式基本一致.     

Calbindin neurons of the guinea-pig small intestine: quantitative analysis of their numbers and projections

Summary The distribution of nerve cells with immunoreactivity for the calcium-binding protein, calbindin, has been studied in the small intestine of the guinea-pig, and the projections of these neurons have been analysed by tracing their processes and by examining the consequences of nerve lesions. The immunoreactive neurons were numerous in the myenteric ganglia; there were 3500±100 reactive nerve cells per cm² of undistended intestine, which is 30% of all nerve cells. In contrast, reactive nerve cells were extremely rare in submucous ganglia. The myenteric nerve cells were oval in outline and gave rise to several long processes; this morphology corresponds to Dogiel's type-II classification. Processes from the cell bodies were traced through the circular muscle in perforating nerve fibre bundles. Other processes ran circumferentially in the myenteric plexus. Removal of the myenteric plexus, allowing time for subsequent fibre degeneration, showed that reactive nerve fibres in the submucous ganglia and mucosa came from the myenteric cell bodies. Operations to sever longitudinal or circumferential pathways in the myenteric plexus indicated that most reactive nerve terminals in myenteric ganglia arise from myenteric cell bodies whose processes run circumferentially for 1.5 mm, on average. It is deduced that the calbindin-reactive neurons are multipolar sensory neurons, with the sensitive processes in the mucosa and with other processes innervating neurons of the myenteric plexus.     

Projections of neurochemically specified neurons in the porcine colon

The intramural projections of nerve cells containing serotonin (5-HT), calcitonin gene-related peptide (CGRP), vasoactive intestinal peptide (VIP) and nitric oxide synthase or reduced nicotinamide adenine dinucleotide phosphate diaphorase (NOS/NADPHd) were studied in the ascending colon of 5- to 6-week-old pigs by means of immunocytochemistry and histochemistry in combination with myectomy experiments. In control tissue of untreated animals, positive nerve cells and fibres were common in the myenteric and outer submucous plexus and, except for 5-HT-positive perikarya, immunoreactive cell bodies and fibres were also observed in the inner submucous plexus. VIP- and NOS/NADPHd-positive nerve fibres occurred in the ciruclar muscle layer while VIP was also abundant in nerve fibres of the mucosal layer. 5-HT- and CGRP-positive nerve fibres were virtually absent from the aganglionic nerve networks. In the submucosal layer, numerous paravascular CGRP-immunoreactive (IR) nerve fibres were encountered. Myectomy studies revealed that 5-HT-, CGRP-, VIP- and NOS/NADPHd-positive myenteric neurons all displayed anal projections within the myenteric plexus. In addition, some of the serotonergic myenteric neurons projected anally to the outer submucous plexus, whereas a great number of the VIP-ergic and nitrergic myenteric neurons send their axons towards the circular muscle layer. The possible function of these nerve cells in descending nerve pathways in the porcine colon is discussed in relation to the distribution pattern of their perikarya and processes and some of their morphological characteristics.     

Nitric oxide synthase in the spinal cord of the frog, Xenopus laevis

The expression of nitric oxide synthase was investigated in the spinal cord of the South African clawed frog by NADPH diaphorase histochemistry and immunohistochemistry. The dorsal field contained many strongly positive neurons and a dense plexus of processes. Only few nitric oxide synthase-positive cells occurred in the lateral and central field. Motoneurons were negative. A dense accumulation of stained neurons was located dorsal and dorsomedial to the motoneurons. The white matter harbored many positive fibers. These were most abundant in the dorsal funiculus, and obviously consist of nonprimary projections to the brainstem. These results suggest that nitric oxide represents a widely used messenger molecule in the frog spinal cord, in particular with respect to the processing of sensory information.     

Transient expression of the calcitonin receptor by enteric neurons of the embryonic and early post-natal mouse

Calcitonin receptor-immunoreactivity (CTR-ir) was found in enteric neurons of the mouse gastrointestinal tract from embryonic day 13.5 (E13.5) to post-natal day 28 (P28). CTR-ir occurred in cell bodies in ganglia of the myenteric plexus extending from the esophagus to the colon and in nerve cells of the submucosal ganglia of the small and large intestines. CTR-ir was also found in vagal nerve trunks and mesenteric nerves. Counts in the ileal myenteric plexus revealed CTR-ir in 80% of neurons. CTR-ir was clearly evident in the cell bodies of enteric neurons by E15.5. The immunoreactivity reached maximum intensity between P1.5 and P12 but was weaker at P18 and barely detectable at P28. The receptor was detected in nerve processes in the intestine for only a brief period around E17.5, when it was present in one to two axonal processes per villus in the small intestine. In late gestation and soon after birth, CTR-ir was also evident in the mucosal epithelium. The perinatal expression of CTR within the ENS suggests that the calcitonin/CTR system may have a role in the maturation of enteric neurons. Signals may reach enteric neurons in milk, which contains high levels of calcitonin.     

Calcitonin gene-related peptide-containing neurons supplying the rat digestive system: differential distribution and expression pattern.

In the enteric nervous system, calcitonin gene-related peptide (CGRP) immunoreactivity is localized to a substantial number of capsaicin-sensitive afferent fibers and to intrinsic neurons and processes. CGRP immunoreactivity detected by immunohistochemistry represents the expression of two distinct genes, the calcitonin/alpha-CGRP and the beta-CGRP genes, which have different tissue distributions. In the present study, we used (1) in situ hybridization histochemistry and ribonucleic acid (RNA) blot hybridization with RNA probes complementary to the divergent sequences of alpha- and beta-CGRP messenger RNAs (mRNAs) to differentiate which CGRP gene was expressed in enteric and afferent neurons; and (2) axonal transport approaches in combination with CGRP immunohistochemistry to define the location of CGRP-containing afferent neurons supplying the digestive system. In situ hybridization histochemistry with [35S]-labeled RNA probes indicated that in the gastrointestinal tract beta-CGRP mRNA, but not alpha-CGRP mRNA, was expressed in enteric neurons confined to the myenteric and submucous plexuses of the small and large intestine. In dorsal root and vagal sensory ganglia, mRNAs for alpha-CGRP and beta-CGRP were both present in a vast population of neurons, with an overlapping pattern, even though the alpha-CGRP signal appeared more intense. RNA blot hybridization analysis showed a single band of hybridization at 1.2 Kb with the beta-CGRP RNA probe in RNA extracts from muscle layer-myenteric plexus and submucosal layer preparations of the ileum, and from dorsal root ganglia; it also showed a single band at 1.3 Kb with the alpha-CGRP RNA probe in extracts from dorsal root ganglia, but not from the intestine. These findings further support the differential expression of alpha- and beta-CGRP mRNAs. Retrograde transport of fast blue or fluorogold coupled with CGRP immunohistochemistry demonstrated that the vast majority of CGRP-containing afferent neurons supplying the stomach, proximal duodenum, and pancreas were located in dorsal root ganglia at the middle and lower thoracic and at the upper lumbar levels, and represented a major component of the afferent innervation of these viscera (up to 89%). Approximately 50% of CGRP-immunoreactive afferent neurons also expressed tachykinin (TK) immunoreactivity, as shown by triple labeling. Only a minor component of the afferent innervation of the stomach, duodenum, and pancreas derived from vagal CGRP-containing neurons (less than 8%). A large portion of these neurons (an average of 62%) also contained TK immunoreactivity.(ABSTRACT TRUNCATED AT 400 WORDS)