两种阿尔茨海默病转基因小鼠模型脑新皮质区A(beta)分布的比较研究

目的:比较Aβ在两种阿尔茨海默病转基因小鼠模型脑新皮质区分布的差异。方法:采用18月龄雄性APP/PSl双转基因(2×Tg-AD)小鼠与同龄同性别APP/PSl/tau三转基因(3×Tg-AD)小鼠,取新皮质区脑组织行6E10单克隆抗体免疫组化染色等方法显示Aβ阳性神经元及斑块,观察其分布与形态等的差异,图像分析系统定量比较其量的变化。结果:在新皮质区2×Tg-AD组Aβ阳性产物主要位于细胞外即细胞外Aβ(e Aβ),形成大量的老年斑,细胞内阳性产物少;而3×Tg-AD组Aβ阳性产物主要位于神经元细胞内即细胞内Aβ(i Aβ),但老年斑少见。结论:2×Tg-AD组与3×Tg-AD组Aβ阳性产物在新皮质区分布的差异可能反映了两种AD小鼠模型神经病理等改变的不同。     

两种阿尔茨海默病转基因小鼠模型脑新皮质区Aβ分布的比较研究

目的:比较Aβ在两种阿尔茨海默病转基因小鼠模型脑新皮质区分布的差异。方法:采用18月龄雄性APP/PSl双转基因(2×Tg-AD)小鼠与同龄同性别APP/PSl/tau三转基因(3×Tg-AD)小鼠,取新皮质区脑组织行6E10单克隆抗体免疫组化染色等方法显示Aβ阳性神经元及斑块,观察其分布与形态等的差异,图像分析系统定量比较其量的变化。结果:在新皮质区2×Tg-AD组Aβ阳性产物主要位于细胞外即细胞外Aβ(e Aβ),形成大量的老年斑,细胞内阳性产物少;而3×Tg-AD组Aβ阳性产物主要位于神经元细胞内即细胞内Aβ(i Aβ),但老年斑少见。结论:2×Tg-AD组与3×Tg-AD组Aβ阳性产物在新皮质区分布的差异可能反映了两种AD小鼠模型神经病理等改变的不同。     

BDNF和NT—3在鸡胚脊髓发育中的表达——免疫组织化学研究

探讨脑源性神经营养因子（BDNF）和神经营养因子3（NT－3）在脊髓发育中的表达。取Hamburger 30期和40期鸡胚腰段脊髓制作20μm厚冰冻切片,分别用BDNF和NT－3抗体行ABC免疫组化染色。观察BDNF、NT－3样免疫阳性反应物（BDNF－IR、NT－3－IR）在两时相脊髓的分布。结果显示：30期组,BDNF－IR主要分布于脊髓腰段腹角神经元的核周质及神经突起内。NT－3－IR则仅限于腹角前群神经元,胞核、胞浆均梁色;40期组,BDNF－IR除腹角染色外,还扩展至整个脊髓肖解神经元及神经突起。另外,亦见一些BDNF－IR的胶质细胞。与之类似,NT－3－IR除分布于腹角前群神经元外,中间带、背角内亦出现了些NT－3阳性神经元。结果表明：BDNF和NT－3在脊髓的表达随发育进程由腹角向背角扩展。提示：BDNF和NT－3的生理作用与脊髓的发育有关。     

猫外侧膝状体年龄相关性形态学变化

目的比较青年猫与老年猫外侧膝状体(lateral geniculate nucleus,LGN)神经元及γ-氨基丁酸(gama-aminobutyric acid,GABA)能神经元的年龄相关性变化,探讨老年个体视觉功能衰退的相关神经机理。方法Nissl染色示猫外侧膝状体分层结构(A、A1、C3层)及神经元,免疫组织化学法示GABA免疫阳性神经元。光镜下观察、拍照,Nissl染色切片测量外侧膝状体各层厚度、神经元胞体直径并计数神经元数量;免疫组化染色切片测量外侧膝状体各层中GABA阳性神经元胞体直径并计数GABA阳性神经元数量。结果青年猫及老年猫外侧膝状体各层厚度、神经元数量及胞体直径无明显改变(P>0.05);与青年猫相比,老年猫外侧膝状体各层中GABA阳性神经元数量及胞体直径均有不同程度的显著下降(P<0.01),且GABA免疫阳性反应减弱。结论在动物个体衰老进程中,外侧膝状体总体神经元保持相对稳定可能对老年个体维持视觉功能具有一定意义;老年个体外侧膝状体GABA能神经元对视觉信息传递及整合过程的抑制性调节功能削弱,可能是外侧膝状体水平上导致老年个体视觉功能衰退的原因之一。     

大鼠肠道内NOS与AChE、VIP阳性神经元的分布关系研究

应用一氧化氮合酶 (NOS)、乙酰胆碱酯酶 (ACh E)组织化学及血管活性肠肽 (VIP)免疫组织化学方法 ,光镜下比较观察大鼠肠道内 NOS、ACh E、VIP阳性神经元的形态学特征。结果显示 ,肠肌间丛 NOS阳性神经元胞体大小不等 ,形态不一 ,NOS、ACh E和 VIP阳性神经元的分布密度为 ACh E>NOS>VIP,在不同的肠段和层次分布密度有差异 ,NOS与 ACh E存在共染。在肌间丛和粘膜下丛 ,少数 VIP与 NOS共染。在粘膜下丛 ,三种阳性神经元的分布密度为 ACh E>VIP>NOS。在肌间丛和粘膜下丛 ,可见 VIP阳性末梢环抱 NOS阳性神经元胞体 ,两者呈终扣样接触。上述结果提示 NOS阳性神经元与 ACh E、 VIP阳性神经元有密切的形态学联系。在消化道功能调节上 ,它们可能起协调作用。     

猫小脑皮质GABA能神经元年龄相关性变化

为探讨青年猫和老年猫小脑皮质GABA能神经元及其表达的年龄相关性变化,利用Nissl染色显示小脑皮质结构及神经元,免疫组织化学ABC法标记GABA免疫阳性神经元。光镜下观察,采集图像,并利用图像分析软件对分子层、蒲肯野细胞层和颗粒层神经元及GABA免疫阳性神经元及其灰度值进行分析统计。结果显示,GABA免疫阳性神经元、阳性纤维及终末在青年猫和老年猫小脑皮质各层均有分布。与青年猫相比,老年猫分子层、蒲肯野细胞层神经元和GABA免疫阳性神经元密度及其GABA免疫阳性反应强度均显著下降(P<0.01),颗粒层神经元密度和GABA免疫阳性强度也显著下降(P<0.01),但其GABA免疫阳性神经元密度无显著变化(P>0.05);蒲肯野细胞的胞体萎缩,阳性树突分枝减少。因此认为,衰老过程中猫小脑皮质GABA能神经元的丢失和GABA表达的下降,可能是老年个体运动协调、精确调速和运动学习等能力下降的重要原因之一。     

慢性间断性低氧诱导大鼠前包钦格复合体磷酸化PKC底物表达增加

延髓腹外侧前包钦格复合体(pre-B?tzinger complex, pre-B?tC)被认为是呼吸节律产生中枢。间断性低氧可诱导呼吸长时程易化(long-term facilitation, LTF),是呼吸可塑性的电生理特征。本研究组前期研究显示慢性间断性低氧(chronic intermittent hypoxia,CIH)诱导大鼠pre-B?tC磷酸化蛋白激酶Cθ(phospho-protein kinase Cθ, P-PKCθ)表达上调。本研究旨在探讨磷酸化-蛋白激酶C底物(P-PKC substrates, P-PKCsub)在大鼠pre-B?tC的超微结构分布以及CIH干预后的表达变化。应用神经激肽1受体(neurokinin-1 receptor, NK1R)免疫反应(immunoreactive, ir)产物作为pre-B?tC神经元的标志,并用抗P-PKCsub抗体双标记进行免疫荧光和免疫电镜观察,用Western blot分析延髓腹外侧区(包含pre-B?tC) P-PKCsub蛋白的表达变化。结果显示,光镜下,NK1R-ir标记主要沿pre-B?tC神经元胞膜分布,清晰勾勒胞体和突起。P-PKCsub-ir标记多呈点状分布在胞体和突起,膜下亦有分布。大部分P-PKCsub-ir神经元共表达NK1R。CIH干预诱导延髓腹外侧区P-PKCsub蛋白表达水平上调。电镜下,NK1R-ir产物主要分布在pre-B?tC神经元胞体和树突的胞膜内表面。P-PKCsub-ir金颗粒分布在pre-B?tC神经元胞体和树突,在细胞膜下有较多分布,内质网和突触后致密体亦可见。以上结果提示,CIH干预可能通过激活PKCθ,上调P-PKCsub蛋白表达,参与pre-B?tC的呼吸可塑性调控。     

代谢型谷氨酸受体五种亚型在猫脊髓内的分布—免疫组织化学方法研究

为研究代谢型谷氨酸受体各亚型在猫脊髓内的定位分布特征,本实验采用免疫组织化学方法调查了代谢型谷氨酸受体五种亚型（ｍＧｌｕＲ１,ｍＧｌｕＲ２,ｍＧｌｕＲ３,ｍＧｌｕｒ５及ｍＧｌｕＲ７）在猫脊髓（颈２、颈６、胸５、腰６、骶１－２）内的分布状况。结果如下：（１）在背角深层（Ⅲ－Ⅳ层）和腹角内分布有大量中小型（＜３０μｍ）ｍＧｌｕＲ１样免疫反应阳性神经元,背角浅层（Ⅰ、Ⅱ层）为阴性;（２）ｍＧｌｕＲ２／３样免疫反应阳性产物仅见于背角Ⅱ层内侧部,其它部位为阴性;（３）致密的ｍＧｌｕＲ５样免疫反应阳性产物主要分布于猫脊髓背角Ⅱ层的神经毯内,背角Ⅰ层及深层呈中等密度染色;（４）致密的ｍＧｌｕＲ７样免疫反应阳性产物分布于背角Ⅱ层的神经毯内,背角Ⅰ层及深层（Ⅲ－Ⅳ层）呈中等密度染色。此外,腹角运动神经元和骶髓副交感核内的神经元亦呈现ｍＧｌｕＲ７样免疫反应阳性。单侧切断背根,发现术侧背角浅层内ｍＧｌｕＲ７样免疫反应阳性产物密度略有降低,而ｍＧｌｕＲ５样免疫反应阳性产物密度与对侧基本相同。本研究结果显示代谢型谷氨酸受体五种亚型在所调查的猫脊髓各节段内（颈２、颈６、胸５、腰６、骶１－２）的分布存在差异,提示它们在介导谷氨酸传递的信     

Ⅱ型囊泡膜谷氨酸转运体阳性终末与γ-氨基丁酸阳性神经元在小鼠腰髓背角的分布及联系

目的 研究Ⅱ型囊泡膜谷氨酸转运体(vesicular glutamate transporter 2,VgluT2)阳性终末与γ-氨基丁酸(γ-aminobutyric acid,GABA)阳性神经元在小鼠腰髓背角的分布和联系。方法采用免疫组织化学方法研究VgluT2阳性终末与GABA阳性神经元在小鼠腰髓背角的分布;采用免疫荧光组织化学双重标记方法研究VgluT2阳性终末与GABA阳性神经元在小鼠腰髓背角的联系。结果VgluT2阳性终末与GABA阳性神经元在小鼠腰髓背角各层均有分布,特别是在Ⅱ层内侧部二分布都较为密集,免疫荧光双重标记后在激光共聚焦显微镜下可见GABA阳性神经元周围有许多VGluT2阳性终末与其胞体或突起密切接触。结论小鼠腰髓背角Ⅱ层内侧部GABA阳性神经元直接接受兴奋性传入。     

背根切断对脊髓后角Ⅱ板层BDNF,NT-3表达的影响——免疫组织化学研究

目的　采用免疫组织化学技术探讨切断背根 (L1)后脊髓Ⅱ板层脑源性神经营养因子 (BDNF)和神经营养因子 3 (NT 3)表达的变化。方法　将成年雄猫 5只行单侧L1背根切断术 (对侧为非手术侧 )。术后 5天取L1脊髓制作2 0 μm厚冰冻切片 ,用BDNF及NT 3抗体分别进行免疫组化染色。观察BDNF、NT 3免疫阳性反应物在脊髓的分布 ,计数单位面积内Ⅱ板层BDNF阳性膨体密度及NT 3阳性细胞数。结果用t检验进行统计分析。结果　BDNF样免疫反应物在Ⅱ板层主要分布于神经膨体 ,NT 3样免疫反应物在神经元及胶质细胞均有分布。背根切断后 ,手术侧Ⅱ板层BDNF阳性膨体数量明显较非手术侧者减少 (P <0 0 1)。而手术侧Ⅱ板层NT 3阳性神经元及胶质细胞数量则较非手术侧者明显增加(P <0 0 1)。结论　背根切断后脊髓Ⅱ板层BDNF ,NT 3的表达发生不同变化。BDNF减少 ,而NT 3表达增多。提示BD NF和NT 3在脊髓损伤修复中的不同作用。     

Relationships between neurokinin receptor-expressing interstitial cells of Cajal and tachykininergic nerves in the gut

The so-called interstitial cells of Cajal (ICC) are distributed throughout the muscle coat of the alimentary tract with characteristic intramural location and species-variations in structure and staining. Several ICC sub-types have been identified: ICC-DMP, ICC-MP, ICC-IM, ICC-SM. Gut motility is regulated by ICC and each sub-type is responsible for the electrical activities typical of each gut region and/or muscle layer. The interstitial position of the ICC between nerve endings and smooth muscle cells has been extensively considered. Some of these nerve endings contain tachykinins. Three distinct tachykinin receptors (NK1r, NK2r and NK3r) have been demonstrated by molecular biology. Each of them binds with different affinities to a series of tachykinins (SP, NKA and NKB). In the ileum, SP-immunoreactive (SP-IR) nerve fibers form a rich plexus at the deep muscular plexus (DMP), distributed around SP-negative cells, and ICC-DMP intensely express the SP-preferred receptor NK1r; conversely a faint NK1r-IR is detected on the ICC-MP and mainly after receptor internalization was induced by agonists. ICC-IM are never stained in laboratory mammals, while those of the human antrum are NK1r- IR. RT-PCR conducted on isolated ileal ICC-MP and gastric ICC-IM showed that these cells express NK1r and NK3r. Colonic ICC, except those in humans, do not express NK1r-IR, at least in resting conditions. Outside the gut, NK1r-IR cells were seen in the arterial wall and exocrine pancreas. In the mouse gut only, NK1r-IR is present in non-neuronal cells located within the intestinal villi, so-called myoid cells, which are c-kit-negative and alpha-smooth muscle actin-positive. Immunohistochemistry and functional studies confirmed that ICC receive input from SP-IR terminals, with differences between ICC sub-types. In the rat, very early after birth, NK1r is expressed by the ICC-DMP and SP by the related nerve varicosities. Studies on pathological conditions are few and those on mutant strains practically absent. It has only been reported that in the inflamed ileum of rats the NK1r-IR ICC-DMP disappear and that at the peak of inflammatory conditions ICC-MP are NK1r-IR. In the ileum of mice with a mutation in the W locus, ICC-DMP were seen to express c-kit-IR but not NK1-IR, and SP-IR innervation seems unchanged. In summary, there are distinct ICC populations, each of them under a different tachykininergic control and, likely, having different functions. Further studies are recommended at the aim of understanding ICC involvement in modulating/transmitting tachykininergic inputs.     

Co-localization of Pirt protein and P2X2 receptors in the mouse enteric nervous system

P2X2 receptors, with other P2X receptor subtypes, have an important role mediating synaptic transmission in regulating the functions of the gastrointestinal tract. Our recent work has found a new regulator of P2X receptor function, called phosphoinositide-interacting regulator of transient receptor potential channels (Pirt). In the present work, we have shown that Pirt immunoreactivity was localized in nerve cell bodies and nerve fibers in the myenteric plexus of the stomach, ileum, proximal, and distal colon and in the submucosal plexus of the jejunum, ileum, proximal, and distal colon. Almost all the Pirt-immunoreactive (ir) neurons were also P2X2-ir, and co-immunoprecipitation experiments have shown that Pirt co-precipitated with the anti-P2X2 antibody. This work provides detailed information about the expression of Pirt in the gut and its co-localization with P2X2, indicating its potential role in influencing P2X2 receptor function.     

大熊猫胃肠道神经肽Y和长型瘦素受体的表达

为了观察神经肽Y(NPY)和瘦素长型受体(OB-Rb)在大熊猫胃肠道的表达分布,并探讨其功能。本实验选用中国保护大熊猫研究中心濒危动物繁殖与保护遗传四川省重点实验室提供的3例大熊猫胃肠组织样品,采用HE和免疫组化SABC法进行组织学、NPY及OB-Rb蛋白的表达研究。HE染色结果显示3例大熊猫胃肠道组织学结构完整,主要表现在单细胞和多细胞黏液腺丰富、小肠绒毛较长,肠道黏膜肌层与肌肉层较厚等。免疫组化方法观察到NPY和OB-Rb阳性产物广泛分布于大熊猫的胃肠道中。NPY阳性神经纤维呈串珠状或点状排列,主要位于黏膜下神经丛和肌间神经丛内,前者阳性神经纤维数量较多。NPY阳性细胞主要分布于黏膜层和肠腺,多呈椭圆形、多边形等。OB-Rb阳性产物主要分布在黏膜层,且在固有层内有大量阳性细胞分布。表明NPY和OB-Rb在大熊猫肠道中的广泛表达,为研究NPY和OB-Rb影响肠道的生长发育、消化吸收、免疫等多种功能奠定了基础。     

A neurochemical characterisation of the golden hamster myenteric plexus

The neurochemical composition of nerve fibres and cell bodies in the myenteric plexus of the proventriculus, stomach and small and large intestines of the golden hamster was investigated by using immunohistochemical and histochemical techniques. In addition, the procedures for localising nitric-oxide-utilising neurones by histochemical (NADPH-diaphorase) and immunohistochemical (nitric oxide synthase) methods were compared. The co-localisation of vasoactive intestinal polypeptide and nitric oxide synthase in the myenteric plexus of all regions of the gut was also assessed. The results demonstrated the presence of nerve fibres and nerve cell bodies immunoreactive to protein gene product, vasoactive intestinal polypeptide, substance P, calcitonin gene-related peptide, tyrosine hydroxylase, 5-hydroxytryptamine and nitric oxide synthase in all regions of the gastrointestinal tract examined. The pattern of distribution of immunoreactive nerve fibres and nerve cell bodies containing the above markers was found to vary in different regions of the gut. Myenteric neurones and nerve fibres containing immunoreactivity to nitric oxide synthase and NADPH-diaphorase reactivity, however, were shown to have an identical distribution throughout the gut. In contrast to some studies on the guinea-pig and rat, the co-existence of vasoactive intestinal polypeptide and nitric oxide synthase was seen in only a small population of myenteric neurones.     

Substance P and Neurokinin 1 receptor - expression is affected in the ileum of mice with mutation in the W locus

The tachykinin substance P (SP) acts on the gut muscle coat via its preferred receptor, neurokinin 1 (NK1r). In the mouse ileum, NK1r-immunoreactivity (NK1r-IR) was detected in neurons, in the interstitial cells of Cajal at the deep muscular plexus (ICC-DMP) and the myoid cells of the villi. SP-IR was detected in neurons and varicose nerve fibers, which were especially numerous at the DMP and closely associated with the ICC-DMP. In mice with a mutation in the W locus (ckit mutant animals), innervation is suggested to be normal although few studies have actually tested this hypothesis. Indeed, studies demonstrating ICC-DMP integrity are lacking and whether SP- and NK1r-IR are normal in these animals has not been investigated. Our aim was to perform an immunohistochemical study on the ileum of a strain of heterozygous mice with a mutation in the W locus, the W(e/+) mice, to test this hypothesis. SP-IR nerve fibers were significantly more numerous than in wild type mice; NK1r-IR was clustered on the plasma membrane and also intracytoplasmatic in the neurons, but absent in the ICC-DMP. The richness in SP-IR nerve fibers and the NK1r-IR distribution in the neurons, similar to that of activated cells, might be attempts to compensate for the SP preferred receptor absence at the ICC-DMP. In conclusion, SP content and NK1r expression are noticeably different in c-kit mutants with respect to wild type mice, and probably causing an anomalous tachykininergic control of intestinal motility. Physiological studies on Wmutant mice have to take into account that innervation in this animal model is affected by the c-kit mutation.     

Ghrelin-producing cells exist as two types of cells, closed- and opened-type cells, in the rat gastrointestinal tract

Ghrelin was recently isolated from the rat stomach as an endogenous ligand for the growth-hormone secretagogue receptor (GHS-R) and is known to exist in the gastrointestinal tract and hypothalamus. In this study, we investigated in detail the distribution and morphologic characteristics of ghrelin-containing cells (ghrelin cells) in the gastrointestinal tract by immunohistochemistry and in situ hybridization. Ghrelin cells were found to be localized in the mucous membrane of the stomach, duodenum, ileum, cecum and colon but not in myenteric plexus, and they can be classified into open- and closed-type cells. The greatest number of ghrelin cells was found in the stomach, and it was found that the number of the opened-type cells gradually increased in the direction from stomach to the lower gastrointestinal tract. These results suggest that the two types of ghrelin cells may be distinctly regulated and play different physiological roles in various regions of the gastrointestinal tract.     

Substance P and substance K receptor binding sites in the human gastrointestinal tract: Localization by autoradiography

Quantitative receptor autoradiography was used to localize and quantify the distribution of binding sites for ¹²⁵I-radiolabeled substance P (SP), substance K (SK) and neuromedin K (NK) in the human GI tract using histologically normal tissue obtained from uninvolved margins of resections for carcinoma. The distribution of SP and SK binding sites is different for each gastrointestinal (GI) segment examined. Specific SP binding sites are expressed by arterioles and venules, myenteric plexus, external circular muscle, external longitudinal muscle, muscularis mucosa, epithelial cells of the mucosa, and the germinal centers of lymph nodules. SK binding sites are distributed in a pattern distinct from SP binding sites and are localized to the external circular muscle, external longitudinal muscle, and the muscularis mucosa. Binding sites for NK were not detected in any part of the human GI tract. These results demonstrate that: 1) surgical specimens from the human GI tract can be effectively processed for quantitative receptor autoradiography; 2) of the three mammalian tachykinins tested, SP and SK, but not NK binding sites are expressed in detectable levels in the human GI tract; 3) whereas SK receptor binding sites are expressed almost exclusively by smooth muscle, SP binding sites are expressed by smooth muscle cells, arterioles, venules, epithelial cells of the mucosa and cells associated with lymph nodules; and 4) both SP and SK binding sites expressed by smooth muscle are more stable than SP binding sites expressed by blood vessels, lymph nodules, and mucosal cells.     

Distribution of neurokinin-2 receptors in the guinea-pig gastrointestinal tract

The distribution of neurokinin-2 (NK2) tachykinin receptors was investigated by immunohistochemistry in the guinea-pig oesophagus, stomach, small and large intestine. Receptor immunoreactivity occurred at the surfaces of smooth muscle cells throughout the digestive tract. Nerve fibre varicosities in enteric ganglia were also immunoreactive. In myenteric ganglia, these varicosities were most numerous in the ileum, frequent, but less dense, in the proximal colon and caecum, rare in the distal colon, extremely infrequent in the rectum and duodenum, and absent from the stomach and oesophagus. Reactive varicosities were rare in the submucous ganglia. Reactive nerve fibres in the mucosa were only found in the caecum and proximal colon. Strong NK2 receptor immunoreactivity was also found on the surfaces of enterocytes at the bases of mucosal glands in the proximal colon. Receptors were not detectable on the surfaces of nerve cells or on non-terminal axons. Reactivity did not occur on nerve fibres innervating the muscle. Denervation studies showed that the immunoreactive varicosities in the myenteric plexus of the ileum were the terminals of descending interneurons. Immunoreactivity for nitric oxide synthase was colocalised with NK2 receptor (NK-R) immunoreactivity in about 70% of the myenteric varicosities in the small intestine. Bombesin immunoreactivity occurred in about 30% of NK2-R immunoreactive varicosities in the small intestine. Received: 10 April 1996 / Accepted: 13 May 1996     

Lack of neurokinin-1 receptor expression affects tissue mast cell numbers but not their spatial relationship with nerves

A spatial association between mast cells and nerves has been described in both the gastrointestinal and genitourinary tracts. However, the factors that influence the anatomic relationship between mast cells and nerves have not been completely defined. It has been suggested that the high-affinity receptor for substance P [neurokinin-1 (NK1)] might modulate this interaction. We therefore assessed mast cell-nerve relationships in tissues isolated from wild-type and NK1 receptor knockout (NK1-/-) mice. We now report that, in the complete absence of NK1 receptor expression, there is a significant increase in the number of mast cells without a change in the anatomic relationship between mast cell and nerves in stomach and bladder tissues at the light microscopic level. We next determined whether transplanted mast cells would maintain their spatial distribution, number, and contact with nerve elements. For this purpose, mast cell-deficient Kit(W)/Kit(W-v) mice were reconstituted with wild-type or NK1-/- bone marrow. No differences in mast cell-nerve contact were observed. These results suggest that NK1 receptor expression is important in the regulation of the number of mast cells but is not important in the interaction between mast cells and nerves. Furthermore, the interaction between mast cells and nerves is not mediated through NK1 receptor expression on the mast cell. Further studies are needed to determine the molecular pathway involved in mast cell migration and interaction with nerve elements, but the model of reconstitution of Kit(W)/Kit(W-v) mice with mast cells derived from different genetically engineered mice is a useful approach to further explore these mechanisms.     

Choline acetyltransferase immunoreactivity of putative intrinsic primary afferent neurons in the rat ileum

The colocalisation of choline acetyltransferase (ChAT) with markers of putative intrinsic primary afferent neurons was determined in whole-mount preparations of the myenteric and submucosal plexuses of the rat ileum. In the myenteric plexus, prepared for the simultaneous localisation of ChAT and nitric oxide synthase (NOS), all nerve cells were immunoreactive (IR) for ChAT or NOS, but seldom for both; only 1.6 +/- 1.8% of ChAT-IR neurons displayed NOS-IR and, conversely, 2.8 +/- 3.3% of NOS-IR neurons were ChAT-IR. In preparations double labelled for NOS-IR and the general nerve cell marker, neuron-specific enolase, 24% of all nerve cells were immunoreactive for NOS, indicating that about 75% of all nerve cells have ChAT-IR. All putative intrinsic primary afferent neurons in the myenteric plexus, identified by immunoreactivity for the neurokinin 1 (NK1) receptor and the neurokinin 3 (NK3) receptor, were ChAT-IR. Conversely, of the ChAT-IR nerve cells, about 45% were putative intrinsic primary afferent neurons (this represents 34% of all nerve cells). The cell bodies of putative intrinsic primary afferent neurons had Dogiel type II morphology and were also immunoreactive for calbindin. All, or nearly all, nerve cells in the submucosal plexus were immunoreactive for ChAT. About 46% of all submucosal nerve cells were immunoreactive for both neuropeptide Y (NPY) and calbindin; 91.8 +/- 10.5% of NPY/calbindin cells were also ChAT-IR and 99.1 +/- 0.7% were NK3 receptor-IR. Of the nerve cells with immunoreactivity for ChAT, 44.3 +/- 3.8% were NPY-IR, indicating that about 55% of submucosal nerve cells had ChAT but not NPY-IR. Only small proportions of the ChAT-IR, non-NPY, nerve cells had NK3 receptor or calbindin-IR. It is concluded that about 45% of submucosal nerve cells are ChAT/calbindin/NPY/VIP/NK3 receptor-IR and are likely to be secretomotor neurons. Most of the remaining submucosal nerve cells are immunoreactive for ChAT, but their functions were not deduced. They may include the cell bodies of intrinsic primary afferent neurons.