内脏伤害性刺激后Fos在大鼠脑内NOS阳性神经元内的表达

目的：观察一氧化氮合酶(NOS)阳性神经元在内脏伤害性信息传递通路上的分布。方法：给予大鼠内脏伤害性刺激后，采用Fos免疫组织化学(ABC法)和还原型尼克酰胺腺嘌呤二核苷酸脱氢酶(NADPH-d)组织化学双重染色的方法，观察脑内NOS和Fos阳性神经元的分布。结果：脑内Fos／NOS双标阳性神经元主要分布在孤束核，中缝背核，丘脑室旁核，下丘脑室旁核、室周核、背内侧核，中脑导水管周围灰质腹外侧部、背外侧部，臂旁内侧核，内侧缰核，杏仁复合体内侧部等部位。结论：NO是内脏伤害性信息传递和调控通路上的神经递质之一。     

大鼠束缚后脑内Fos蛋白的表达

目的探讨大鼠束缚后对大鼠脑内Fos蛋白表达的影响。方法将大鼠束缚于小的塑料桶内1、3或6h,于解束后30min处死,脑组织进行Fos蛋白免疫组织化学染色。结果Fos阳性神经元表达于1．前脑：扣带回、新皮质(尤其是第3和5层)、外侧隔核、杏仁中央核;2．间脑：下丘脑视前区、下丘脑外侧区、视上核、室旁核、第三脑室室周区、弓状核、丘脑室旁核、外侧膝状体、内侧膝状体;3．脑干：中脑的上丘视性层、中脑导水管周围灰质、下丘的皮质部;脑桥的臂旁外侧核、蓝斑、A5区;延髓的耳蜗核、延髓内脏带(MVZ)等处。Fos表达的时问规律是束缚1h最高,3h次之,6h最少。结论大鼠被束缚后全脑多处核团的神经元发生不同程度的Fos反应,随着束缚时间的延长,动物产生适应性,Fos表达减少。     

束缚应激大鼠下丘脑内一氧化氮合酶与c-fos蛋白的共存

为了探讨下丘脑内一氧化氮与应激之间的关系，本实验应用NADPH-d组化法和C-fos免疫组化技术相结合的方法，对束缚应激大鼠下丘脑内一氧化氮合酶（NOS）和c-fos蛋白的分布以及两者的共存关系进行了观察。结果表明，大鼠在束缚应激4h后，（1）室旁核大细胞部和视上核可见密集深染的NOS阳性大细胞；（2）室旁核小细胞部、背内侧核、穹窿周核、环状核、腹内侧核腹外侧部、结节内侧核、外侧区、室周区、乳头体前核和内侧核的外侧区等核团出现疏密不等和深浅不一的NOS阳性细胞；（3）C-fos蛋白以室旁核表达最为强烈，视前区、背内侧核、弓状核和外侧区亦有较强的表达；（4）在外侧区、室旁核小细胞部及其附近的室周区、背内侧核及乳头体前核腹侧部约有10％～15％的中、小型NOS阳性细胞同时表达C-fos蛋白，室旁核大细胞部则仅有较少的NOS阳性大细胞表达C－fos蛋白，在结节区、结节内侧核和视上核视交叉后部则偶见双标细胞。结果提示上述下丘脑核团内的部分一氧化氮能神经元与束缚应激反应有关。     

脊髓横断伤后Fos在脑内与内脏和心血管相关核团中的表达

目的为阐明急性脊髓损伤对内脏及心血管活动的影响机制提供形态学资料。方法脊髓T4节段横断术后3h,用免疫组织化学方法观察脑与脊髓内脏和心血管相关核团内Fos表达。结果T4水平损伤3h后,中央杏仁核、下丘脑室旁核、中缝背核、导水管周围灰质、臂旁核、蓝斑、孤束核、延髓腹外侧网状核与脊髓中间带外侧核等核团中Fos阳性神经元数目较假手术组显著增加(P<0.01)。结论急性脊髓损伤可引起中枢神经系统的内脏和心血管相关核团内神经元产生特异性反应,但反应的机制不同。     

大白鼠导水管周围灰质的传入性联系研究

应用辣根过氧化物酶微量及离子透入法注射在大白鼠的中脑导水管周围灰质的背侧区、外侧区、腹外侧区以及间脑与中脑交界处第Ⅲ脑室室周灰质部位。在导水管周围灰质的背侧区注射后,标记细胞可在:室旁核、下丘脑腹内侧核、下丘脑背内侧核、下丘脑后核、乳头体内侧核、暖昧带、导水管周围灰质腹外侧核和外侧核、上丘、下丘、中缝背核、第Ⅳ脑室室周灰质中出现。在导水管周围灰质的外侧区注射后,标记细胞则可在终纹床核、室旁核、下丘脑前核、下丘脑腹内侧核、丘脑内侧核、后联合巨细胞间质核、暖味带、导水管周围灰质腹外侧核、黑质网带、中缝背核、第Ⅲ及第Ⅳ脑室室周灰质中出现。在导水管周围灰质的腹外侧区注射后,始终可有标记的核群是。视上核、室旁核、第Ⅲ脑室室周灰质、导水管周围灰质外侧核、黑质网带、中缝背核、蓝斑。在间脑与中脑交界处第Ⅲ脑室室周灰质注射后,有标记的核群是:终纹床核、视上核、室旁核、下丘脑腹内侧核、丘脑内侧核、暖昧带、导水管周围灰质外侧核、黑质网带、中缝背核和蓝斑。     

卵蛋白诱发哮喘发作时豚鼠大脑和肺内Fos蛋白的表达

为了解支气管哮喘后豚鼠大脑和肺组织c-fos基因表达的变化，探讨c-fos表达在豚鼠哮喘发病中的可能意义。我们复制卵蛋白致敏哮喘豚鼠模型，采用免疫组织化学ABC方法，对Fos蛋白在大脑和肺脏内的分布情况进行观察。结果发现：哮喘组豚鼠大脑和肺内c-fos表达较对照组明显增加，其Fos阳性产物在大脑主要分布于额顶皮质、边缘前脑（扣带皮质、梨状皮质和中央杏仁核等）、丘脑室旁核、下丘脑室旁核、视上核、下丘脑外侧区、下丘脑室周核、孤束核、最后区和延髓腹外侧区内，小脑内无明显Fos分布密集区。原癌基因c-fos的表达增强在豚鼠支气管哮喘发病过程中可能起一定作用。     

大鼠延髓至下丘脑室旁核的儿茶酚胺能投射神经元在胃伤害性刺激后的FOS表达

用ＨＲＰ注入下丘脑室旁核逆行追踪与抗Ｆｏｓ和抗酪氨酸羟化酶（ＴＨ）双重免疫细胞化学相结合的三重标记方法，对大鼠孤束核和延髓腹外侧区至下丘脑室旁核的儿茶酚胺能投射神经元对胃伤害性刺激后的ｃ－ｆｏｓ表达进行了观察，发现孤束核和延髓腹外侧区有７种不同的标记细胞：ＨＲＰ、Ｆｏｓ、ＴＨ单标细胞，Ｆｏｓ／ＨＲＰ、Ｆｏｓ／ＴＨ、ＨＲＰ／ＴＨ双标细胞，Ｆｏｓ／ＨＲＰ／ＴＨ三标细胞。上述７种标记细胞主要分布在延髓中、尾段孤束核的内侧亚核、连合亚核和延髓腹外侧区以及两者之间的网状结构。ＨＲＰ标记细胞以注射侧为主，对侧有少量分布。本文结果证明，大鼠孤束核和延髓腹外侧区至下丘脑室旁核投射的部分儿茶酚胺能神经元可能参与胃伤害性刺激的传导和调控。     

全脑暂时性缺血诱导c—fos癌基因蛋白(FoS)在大鼠下丘脑的表达

本文用免疫组化(ABC)方法探查了大鼠全脑暂时性缺血诱导的c—fos癌基因蛋白(Fos)在下丘脑表达的分布以及时间过程。结果如下:四管阻塞全脑缺血后15分钟至12小时内,Fos在下丘脑广泛区域和核团出现短暂而快速的表达。Fos免疫阳性(Fos—IR)产物定位于神经元的胞核内。缺血再循环后15分钟开始表达,3小时达到高峰,12小时消退。最早和最强的表达出现在室周区(特别是视上核和室旁核),其次是内侧区和外侧区。结果提示全脑缺血诱导的Fos在下丘脑短暂而快速的表达与调节缺血引起的内分泌、自主功能、情感和行为反应和维持机体内环境的稳定有关。     

大鼠斜角带核垂直支(VDB)的传入联系

用HRP法研究了VDB的传入联系。结果表明VDB的背、腹侧部均接受下列结构的传入:斜角带核水平支、内侧隔核、扣带皮质、梨状皮质、内嗅皮质、海马、下丘脑外侧核、下丘脑后核、乳头体上核、杏仁内侧核、杏仁皮质核、杏仁基底核内侧及外侧部、室周核、中央灰质腹外侧部、中缝背核、中缝正中核、脚间核、臂旁背侧核、被盖腹侧区及蓝斑。此外,VDB背侧部还接受杏仁外侧核后部、被盖背核的腹侧部及外侧部、嗅前核、未定带及连接核的投射。偶见屏状核投射到VDB背侧部;VDB腹侧部还接受外侧缰核内侧部及杏仁前区的少量投射。     

大白鼠上丘水平导水管周灰质的传入联系——HRP法研究

本实验在45只大白鼠上丘水平导水管周灰质(PAG)内输入30％HRP(Sigma Ⅵ)0.05～0.2μl,分别用DAB、BDHC、O-D及TMB四种组化反应方法处理,以资对照。按注射中心部位及反应范围大小选择29例并分成三组:1.注射中心区在PAG外侧区内,反应范围小,局限于PAG范围内11例。本组仅在同侧PAG、顶盖、邻近网状结构及中缝背核观察到标记细胞。2.注射中心区在PAG背外侧区,反应范围超出PAG,向背外侧扩散到邻近网状结构及顶盖9例。本组除与第一组有共同标记区外,尚在缰核、束旁核、下丘脑(前核、后核、腹内侧核、背内侧核、外侧核)、丘脑(外侧核、网状核、腹核内侧部)、室旁灰质、视束核、后连合核、旁二叠体、蓝斑、外侧膝状体腹核、脚内核见到标记细胞。3.注射中心区在PAG外侧区或腹外侧区,反应区向外侧或腹外侧扩大到邻近网状结构、被盖及部分顶盖9例。本组除与第一组有共同标记区外,尚在黑质、H_1、H_2、未定带、缰核、束旁核、后连合核、旁二叠体、蓝斑、盖前区及下丘脑诸核(同第二组)见到标记细胞。其中3例应用TMB法,除上述标记细胞外,尚在大脑皮质扣带回前分(23、24区)深层、中缝大核、巨细胞网状核、第四脑室室底灰质、小脑齿状核、脊髓颈段(Ⅴ～Ⅶ层)(其他节段未观察)、视前区、视上区、交叉上核、乳头体前核、杏仁内侧核、三叉神经脊束核见到标记细胞。     

内脏伤害性刺激诱导的大鼠延髓内Fos表达

用抗Ｆｏｓ蛋白和酷氨酸羟化酶（ＴＨ）的免疫组织化学方法，对大鼠两种内脏伤害性刺激诱导的大鼠延髓Ｆｏｓ表达情况及其与儿茶酚胺能神经元的关系进行了观察。提示延髓内脏带半数以上的儿茶酚胺神经元参与对内脏伤害性刺激的反应。     

大鼠延髓内神经激肽B受体神经元对腹膜伤害性刺激的Fos表达——免疫组织化学双重染色研究

为探讨神经激肽Ｂ受体（ｎｅｕｒｏｋｉｎｉｎｂｒｅｃｅｐｔｏｒ，ＮＫＲ）神经元在大鼠延髓的定位及其在腹膜伤害性传入信息整合中的作用。应用免疫组织化学双重染色方法，显示ＮＫＲ神经元的分布并观察它们对腹膜化学伤害性刺激的Ｆｏｓ表达。结果表明：ＮＫＲ样免疫反应（ＮＫＲ－ＬＩ）神经元主要分布在孤束核（Ｓｏｌ）、延髓腹外侧区（ＶＬＭ）、三叉神经脊束核尾侧亚核（Ｓｐ５Ｃ）浅层和三叉旁核（Ｐａ５）。腹膜伤害性刺激激诱导的Ｆｏｓ表达神经元多数分布在Ｓｏｌ、ＶＬＭ、Ｐａ５和最后区。Ｓｏｌ、ＶＬＭ和Ｐａ５中约３０．１％的ＮＫＲ－ＬＩ神经元同时呈Ｆｏｓ样免疫反应。本研究提示：延髓内上述核团ＮＫＲ－ＬＩ神经元与腹膜伤害性初级传入信息的整合有关，可能参与内脏伤害性刺激的抗伤害性感受（ａｎｔｉ－ｎｏｃｅｃｉｐｔｉｏｎ）过程。     

大鼠“延髓内脏带”向杏仁核投射的儿茶酚胺神经元对内脏伤害性刺激的FOS表达

本文通过建立ＨＲＰ逆行追踪结合抗ＦＯＳ和抗酪氢酸羟化酶（ＴＨ）的免疫组织化学三重标记技术,在延髓见到以下七种标记细胞：ＦＯＳ,ＨＲＰ,ＴＨ单标细胞,ＦＯＳ／ＨＲＰ,ＦＯＳ／ＴＨ,ＨＲＰ／ＴＨ双标细胞,ＦＯＳ／ＨＲＰ／ＴＨ三标细胞。这些细胞主要分布于延髓中、尾段由孤束核、腹外侧区以及在两者之间的网状结构共同组成的弧形带状区──“延髓内脏带”。本文结果表明大鼠“延髓内脏带”向杏仁核直接投射的儿茶酚胺能神经元中约有４８％对内脏伤害性刺激起反应,提示“延髓内脏带”及其向杏仁核的儿茶酚胺能投射通路参与机体应激反应的调控过程。     

一氧化氮合酶阳性结构在大鼠三叉神经感觉核簇内的分布及其与面部伤害性刺激诱发的FOS蛋白表达的关系

用还原型尼克酰胺腺嘌呤二核苷酸脱氢酶组织化学技术与ＦＯＳ免疫组化（ＡＢＣ法）相结合的方法，观察了大鼠三叉神经感觉核簇内一氧化氮合酶阳性神经元和纤维的分布及其与面部伤害性刺激诱发的ｃ－ｆｏｓ原癌基因蛋白表达的关系．证明，浓密的一氧化氮合酶阳性纤维和终末分布于三叉神经脊束核尾侧亚核的Ⅰ和Ⅱ层，阳性成分呈蓝黑色的带状分布，而在三叉神经感觉核簇的其余核区内很稀疏．一氧化氮合酶阳性神经元胞体与面部伤害性刺激诱发的ｃ－ｆｏｓ原癌基因蛋白表达的阳性神经元在此核簇内的分布节段非常相似，都主要存在于与伤害性传入传递和调控有关的三叉神经脊束核尾侧亚核，并且二者都主要分布于与痛觉调控有关的浅层．约有１０～１５％的一氧化氮合酶阳性神经元同时呈ｃ－ｆｏｓ原癌基因蛋白表达．在三叉脊束核尾侧亚核的Ⅲ～Ⅴ层仅出现散在的阳性细胞，除在吻侧亚核的背内缘与孤束核交界处有少量一氧化氮合酶同性神经元外，在三叉神经感觉核簇的其余核区两者均几无表达．本文及文献结果提示，一氧化氮可能对面部伤害性信息的传递和调控起重要作用．     

Neuronal activation at the spinal cord and medullary pain control centers after joint stimulation: a c-fos study in acute and chronic articular inflammation

Chronic inflammatory pain induces short- and long-term central changes, which have been mainly studied at the spinal cord level. Supraspinal pain control centers intrinsically connected with the dorsal horn are also prone to be affected by chronic inflammatory pain. C-fos expression was used as a neuronal activation marker at spinal and supraspinal levels to i) compare acute and chronic articular inflammation, and ii) analyze the effects of brief innocuous or noxious stimulation of a chronically inflamed joint. Acute articular inflammation was induced by an inflammatory soup with prostaglandin E(2) and bradykinin, both at 10(-5) M. Chronic articular inflammation consisted of 14 days of monoarthritis. Early c-fos expression was studied 4 min after inflammatory soup injection or stimulation of the arthritic joint whereas late c-fos expression was evaluated 2 h after those stimuli. At the spinal cord, the analysis was focused on the dorsal horn (laminae I-V) and supraspinally, five major regions of the endogenous pain control system were considered: the caudal ventrolateral medulla (VLM), the dorsal reticular nucleus (DRt), the ventral reticular nucleus (VRt), the nucleus of the solitary tract (Sol) and the rostroventromedial medulla (RVM). Acute articular inflammation induced early and late increases in c-fos expression at the spinal level and late increases supraspinally whereas the effects of monoarthritis were more moderate and restricted to the spinal cord. When monoarthritic animals were subjected to gentle touch or bending of the joint, early increases in c-fos expression were detected supraspinally, but not at the spinal level. In this region, noxious mechanical stimulation induced late increases in non-inflamed animals and both early and late increases in monoarthritic rats. Supraspinally, noxious stimulation induced only late increases in c-fos expression. The present results show complex differences in the patterns of c-fos expression between the spinal cord and medullary areas of the pain control system during articular inflammation, which indicate that the somatosensory system is differentially affected by the installation of chronic pain.     

Suppressive effects of Neiting acupuncture on toothache: an experimental analysis on Fos expression evoked by tooth pulp stimulation in the trigeminal subnucleus pars caudalis and the periaqueductal gray of rats

To clarify the antinociceptive mechanism of acupuncture on acute pain, c-fos protein (Fos) expression induced by tooth pulp stimulation was immunohistochemically examined in the spinal trigeminal subnucleus pars caudalis (spVc) and the periaqueductal gray (PAG) of rats with or without Neiting acupuncture. The central projection of trigeminal ganglion neurons innervating in the tooth pulp was examined by tract-tracing method with horseradish peroxidase-conjugated wheat germ agglutinin (WGA-HRP). Central terminals from the first maxillary molar tooth were labeled transganglionically in the dorsomedial part of spVc with WGA-HRP. Numerous numbers of Fos-immunoreactive (Fos-ir) cells were found in the spVc and PAG by stimulation of the tooth pulp with acetic acid or saline. Neiting acupuncture significantly reduced the Fos expression in the spVc induced by tooth pulp stimulation. On the other hand, Neiting acupuncture evoked many Fos-ir cells in the PAG. The present results suggest that Neiting acupuncture activated PAG neurons that sent descending inhibitory fibers to medullo-spinal nociceptive neurons, and reduced the number of Fos-expressed neurons in the trigeminal subnucleus pars caudalis mediating noxious information from teeth to the higher central nervous system.     

The alpha3 subtype of the nicotinic acetylcholine receptor is expressed in airway-related neurons of the nucleus tractus solitarius, but is not essential for reflex bronchoconstriction in ferrets

To assess the role of nicotinic acetylcholine receptors (nACh-R) in the transmission of afferent constricting inputs from bronchopulmonary receptors to the nucleus tractus solitarius (nTS) and in the mediation of reflex airway constriction, we performed a combined immunohistological and functional study. In ferrets, the expression of nAch-R on the nTS neurons activated by histamine stimulation of airway sensory receptors was studied using laser scanning confocal microscopy to co-immunolocalize c-fos encoded protein (cFos) and nACh-R alpha3 subunit. We observed that activation of airway sensory receptors by inhalation of aerosolized histamine, induced cFos expression in a subset of nTS neurons that also expressed the nAch-R alpha3 subtype. Furthermore, activation of nACh-R within the commissural subnucleus by nicotine, increased cholinergic outflow to the airways. These effects were diminished by prior administration of hexamethonium (nACh-R blocker) within the commissural subnucleus of the nTS. However, hexamethonium had no significant effects on airway reflex constrictions induced by lung deflation. These findings indicate that nACh-R are expressed by the nTS neurons receiving inputs from airway sensory receptors, activation of which by nicotine increases cholinergic outflow to the airways, but the nACh-R pathways are not required for reflex bronchoconstriction.     

Collateralized projections from neurons in the rostral medulla to the nucleus locus coeruleus, the nucleus of the solitary tract and the periaqueductal gray.

We have examined collateral projections of locus coeruleus afferent neurons in the rostral medulla to the caudal nucleus of the solitary tract or to the periaqueductal gray using double retrograde labeling techniques in the rat. The present findings confirm previously reported connections to the locus coeruleus, the nucleus of the solitary tract and the lateral periaqueductal gray from the nucleus paragigantocellularis in the rostral ventral medulla. Our results also reveal previously unreported projections from the rostral dorsomedial medulla (in a similar region as locus coeruleus-projecting neurons) to the lateral periaqueductal gray. Following retrograde tracer injections into the nucleus of the solitary tract and the locus coeruleus, doubly labeled neurons were seen in both the nucleus paragigantocellularis and in the rostral dorsomedial medulla. Cell counts revealed that approximately 25% of locus coeruleus-projecting neurons in the nucleus paragigantocellularis, and 12% in the dorsomedial medulla, also innervate the caudal nucleus of the solitary tract. In contrast, no doubly labeled neurons within the rostral ventral medulla were found following injections into the lateral periaqueductal gray and the locus coeruleus, although singly labeled neurons for the two tracers were interdigitated in some regions. Following these injections, numerous neurons were also retrogradely labeled in the dorsomedial medulla in the region of the medial prepositus hypoglossi and the perifascicular reticular formation. A small percentage of locus coeruleus afferents in the dorsal medulla (approximately 10%) also projected to the lateral periaqueductal gray. These results indicate that neurons in both the ventrolateral and dorsomedial rostral medulla frequently send collaterals to both the locus coeruleus and the caudal nucleus of the solitary tract. A small number of neurons in the dorsomedial medulla project to both the locus coeruleus and the lateral periaqueductal gray, but separate populations of neurons project to the locus coeruleus and the lateral periaqueductal gray from the ventrolateral medulla. These results functionally link the locus coeruleus and the nucleus of the solitary tract by virtue of common afferents, and support other studies indicating the importance of central autonomic circuitry in the afferent control of locus coeruleus neurons.