尿液气味刺激后棕色田鼠主嗅球和副嗅球的神经元活动

本实验利用免疫组织化学方法,以FOS阳性反应作为神经元活动的标志,研究了棕色田鼠在受到同性和异性尿液刺激后主嗅球和副嗅球的神经元活动,表明两大嗅觉系统均有感知社会性化学信号的功能.通过比较棕色田鼠在同性和异性尿液刺激后副嗅球和主嗅球的嗅小球细胞层(GL)、僧帽细胞层(MIT)、颗粒细胞层(GRL)中FOS阳性神经元数量,发现不同性别尿液刺激后棕色田鼠的副嗅球各细胞层FOS阳性神经元数量比对照组明显增加;棕色田鼠受到异性尿液刺激后其副嗅球各细胞层的FOS阳性神经元数量均多于同性尿液刺激组.不同性别尿液刺激后棕色田鼠的主嗅球各细胞层FOS阳性神经元数量相较于对照组有增加或增加显著;异性尿液刺激组主嗅球各细胞层的FOS阳性神经元数量均多于同性尿液刺激组.说明棕色田鼠的副嗅球和主嗅球均参与了通过尿液介导的性别个体的识别.     

棕色田鼠雄性幼体不同发育期犁鼻器和副嗅球的组织结构

通过对出生后不同发育时期雄性棕色田鼠犁鼻器和副嗅球进行组织学观察, 探讨棕色田鼠出生后犁鼻器和副嗅球的发育规律。实验以出生后当天(0 日龄) , 5 日龄, 15 日龄, 25 日龄以及成年棕色田鼠为研究对象,副嗅球采用Pischinger 氏染色法染色, 犁鼻器用H. E. 染色法染色后进行组织学观察。结果显示, 棕色田鼠出生时, 犁鼻器和副嗅球就已具有成体的基本结构, 随着动物个体的发育, 犁鼻上皮逐渐增厚, 犁鼻管变长, 犁鼻上皮中神经元密度增加; 腺体逐渐增大, 犁鼻管腔填充物增多, 犁鼻管背外侧的静脉血管逐日增大, 管腔周围出现越来越多的血管; 副嗅球长宽都增加, 僧帽细胞层和颗粒细胞层逐渐增长, 各层细胞密度变化稍有不同;出生后15 日内, 僧帽细胞层细胞密度增加, 15 日龄以后又开始降低, 25 日龄及成体的僧帽细胞层细胞密度与5日龄的相似; 颗粒细胞层细胞密度持续增高。实验结果提示, 棕色田鼠5 日龄时, 犁鼻器和副嗅球已具有了完整的结构, 到25 日龄时可能达到了功能上的成熟。     

达乌尔黄鼠犁鼻器和副嗅球的组织结构及嗅球c-Fos表达的季节变化

啮齿动物的犁鼻器和副嗅球与社会通讯和生殖行为有关,主嗅球影响其觅食行为。达乌尔黄鼠（Spermophilus dauricus）是一种具有较低社会行为的储脂类冬眠动物。本研究用组织学和免疫组织化学方法探究了其犁鼻器和副嗅球的结构特点及嗅球神经元活动对季节变化的适应。结果发现,达乌尔黄鼠犁鼻器具有较大的血管,犁鼻器管腔外侧为非感觉性的呼吸上皮（Respiratory epithelium,RE）,内侧为感觉上皮（Sensory epithelium,SE）,RE较SE薄,靠近管腔处为假复层柱状上皮。选取犁鼻器中间部位比较,发现SE的厚度、长度及感觉细胞密度均无性别差异。副嗅球位于主嗅球后方背内侧,由6层细胞构成。侧嗅束穿过副嗅球,位于颗粒细胞层之上。雄性达乌尔黄鼠较雌性有更长的僧帽细胞层和颗粒细胞层。春季（3月）和冬季（1月）达乌尔黄鼠主嗅球的嗅小球层、僧帽细胞层和颗粒细胞层的c-Fos-ir神经元密度显著低于夏季（7月）和秋季（10月）,且冬季外网织层的c-Fos-ir神经元密度显著低于夏季和秋季,说明达乌尔黄鼠在冬季和春季的嗅觉神经活动较弱,呈现出对冬眠的生理性适应。这些结果丰富了动物犁鼻器和副嗅球的形态学资料,并有助于理解冬眠动物嗅觉系统对季节变化和冬眠的适应。     

棕色田鼠与沼泽田鼠犁鼻器和副嗅球的组织结构

用组织学方法研究了棕色田鼠(Microtus mandarinus)、沼泽田鼠(M．fastis)副嗅球和犁鼻器的结构及其在两种鼠间的差异，以此探讨两种田鼠的进化机制与适应功能。两种田鼠的犁鼻器位于鼻腔前端鼻中隔基部的两侧，呈管状结构；沿着犁鼻器的长轴犁鼻管呈现不同的形态学特征，犁鼻管直接开口于鼻腔，从前向后沿着长轴旋转，中间管壁(犁鼻粘膜)变成底部，侧面管壁(假覆层上皮)变成犁鼻管顶壁，最终犁鼻管变小成为一个腺体的分支，不同部位具有不同的组织学特征。通过选取中间相似部位对两种田鼠进行比较研究，发现棕色田鼠犁鼻粘膜比沼泽田鼠厚，而其长度却短于沼泽田鼠。棕色田鼠副嗅球颗粒细胞带宽和僧帽细胞带宽均大于沼泽田鼠，而带长却小于沼泽田鼠。相关分析发现，犁鼻器和副嗅球形态有一定的对应关系，这可能和两个结构之间存在着神经投射有关。棕色田鼠幼体的犁鼻粘膜、神经细胞核、假覆层上皮、血管面积均小于成体[动物学报49(2)：248—255，2003]。     

小鼠副嗅球编码信息素信号

许多哺乳动物依靠信息素 (同类的其它动物产生的半化学物质 )启动神经内分泌反应和社会群居行为。啮齿类动物通过两个不同的神经回路———主嗅系和副嗅系探查化学信号。在主嗅系 ,通过被动呼吸或主动吸气 ,嗅上皮的嗅感受神经元可被微量的挥发性物质激活 ,嗅感受神经元投射至主嗅球 ,后者的僧帽 /簇状细胞再投射到嗅皮质。在副嗅系 ,位于鼻中隔基部的犁鼻器 (VNO)主动张缩 ,使其感受神经元接受非挥发性物质的刺激 ,然后将兴奋传至副嗅球 (AOB) ,后者的僧帽细胞再通过内侧杏仁核投射到下丘脑。主嗅系大面积毁损动物的生殖行为保存完好 ,…     

刺猬嗅球冬眠期与非冬眠期c-Fos 表达的差异

利用免疫组化法检测c-Fos 蛋白在不同季节刺猬嗅球各层次的表达差异,探讨c-Fos、嗅觉、冬眠三者的关系。分别选取春、夏、秋、冬四个季节各6 只野生健康刺猬,固定剥离嗅球,石蜡切片,免疫组化显色,拍片,载入Motic Images Advanced 3.2 软件,测量四个季节刺猬嗅球各层次c-Fos 的表达率,将结果载入GraphPadPrism4 软件分析,Microsoft Excel 作图。结果表明:c-Fos 蛋白在成年刺猬嗅球各层均有不同程度的表达,阴性对照不着色,且表现出明显的季节性差异。1)与秋季相比,冬眠期c-Fos 蛋白在刺猬嗅球各层次的表达均有极显著的降低(P <0.01);2)与夏季相比,冬眠期c-Fos 蛋白在外网丛层、僧帽细胞层、颗粒细胞层的表达有极显著降低(P < 0. 01),在嗅神经层、嗅小球层、室管膜层的表达也有显著降低(P < 0. 05);3)与冬眠期相比,春季c-Fos 蛋白在嗅小球层、僧帽细胞层、颗粒细胞层的表达有极显著的升高(P <0. 01),在嗅神经层、外网丛层、室管膜层的表达却没有显著变化(P ﹥ 0.05);4)嗅神经层c-Fos 的表达在春季显著低于秋季,夏季与秋季没有显著差异。颗粒细胞层夏季显著低于秋季(P < 0.05)。秋季c-Fos 在其余各层次的表达与春季、夏季相比都有极显著的提高(P <0.01)。结论:秋季刺猬嗅球神经元最活跃,嗅觉最灵敏,冬眠期刺猬嗅球活跃性大大降低,嗅觉系统最迟钝。c-Fos 在刺猬嗅球中的强表达表明其在嗅觉信息的传递中可能发挥一定作用,c-Fos 表达率的显著季节性差异揭示了刺猬嗅球的活跃性与其冬眠具有一定的相关性。     

家犬嗅球组织学结构的性别和年龄差异

用组织学方法研究家犬嗅球的结构,观察家犬嗅球内结构的性别和年龄差异,依据常规HE染色法及数理统计学原理对家犬嗅球各层宽度,主要细胞的数量进行比较统计学分析,探讨嗅球内部结构的发育过程以及性别差异对雌雄动物嗅觉差异的影响。结果表明：雌雄幼年家犬嗅球内各层结构差异不显著;成年家犬也表现出同样的结果,但是成年动物的僧帽细胞形态、数量差异极显著。分析发现,幼年家犬嗅球各层结构都已比较明显,成年家犬嗅球体积和重量明显增加,各层宽度明显变宽,各层细胞密度显著降低,说明嗅球也处在不断的发育完善过程之中。同时僧帽细胞的差异可能是造成雌雄动物嗅觉差别的原因之一。     

初孵扬子鳄嗅球的超微结构研究

用电镜研究初孵扬子鳄的嗅球⒚嗅球的外颗粒层具有明、暗两种细胞⒚僧帽细胞层细胞排列紧密、规则,细胞之间无任何连接结构⒚内颗粒层见有 ３～５ 个细胞聚集成群,并有个别细胞出现胞质降解现象⒚除内颗粒层部分细胞外,其他各层细胞仍处于较幼稚阶段⒚胶质细胞已发生,外网状层中有薄薄的髓鞘出现⒚突触处于不同的发育阶段,大多为不对称型⒚     

棕色田鼠和沼泽田鼠雄性社会行为与嗅觉相关脑区性激素受体表达之间的关系

应用行为聚焦取样观察和免疫组织化学相结合的方法 ,比较研究了棕色田鼠 (Microtusmandarinus)(n =15 )和沼泽田鼠 (M .fostis) (n =15 )在同种雄雄交往中的行为差异 ,及在雄雄交往前后雌激素 β受体(ERβ)和雄激素受体 (AR)表达的差异。在 2h的雄雄交往中 ,前 1h棕色田鼠对同性入侵者有较多的攻击和防御行为 ,后 1h攻击行为较少 ,沼泽田鼠前后 1h差异不大 ,整个 2h期间 ,棕色田鼠较沼泽田鼠对同性入侵者有较多的攻击、防御行为 ,较少的非社会行为。经过免疫组织化学检测 ,没有社会交往时棕色田鼠主嗅球系统投射区和犁鼻系统投射区ERβ免疫阳性细胞 (ERβ IRs)明显少于沼泽田鼠 ,且显色淡 ,AR免疫阳性细胞(AR IRs)在两种鼠间差异不大 ,且都明显少于各自的ERβ IRs。 2h交往后 ,棕色田鼠主嗅球投射区和犁鼻系统投射区的ERβ IRs细胞数明显少于交往前 ,AR IRs细胞数明显多于交往前 ;沼泽田鼠交往前与交往后ERβ IRs和AR IRs细胞数均无显著差异 ,且显著多于交往后棕色田鼠ERβ IRs细胞数 ,显著少于交往后棕色田鼠AR IRs细胞数。以上结果表明 :两种田鼠在社会交往中社会行为不同 ;ERβ的减少和AR的增多可能在社会识别及攻击行为中均起一定的作用 ,可能也是引起两种田鼠社会行为发生差异的原因之一     

棕色田鼠和沼泽田鼠的气味识别

把棕色田鼠或沼泽田鼠暴露于同种个体底物(个体饲养箱内的锯末)1h,不同组别的田鼠在间隔15min、30min、60min后,记录对暴露过的熟悉底物和另一个陌生底物的访问、嗅闻、挖掘等社会探究行为(每组在每一个时间间隔只试验1次),统计分析发现,田鼠在暴露于同种异性底物,然后间隔不同的时间后,总是倾向于选择熟悉同种异性底物;田鼠在暴露于同种同性底物,然后间隔不同的时间后,总是倾向于选择陌生同种同性底物;棕色田鼠在暴露于同性底物1h,然后间隔60min后,对陌生的同性底物有明显多的探究行为,但沼泽田鼠在同种情况下却没有表现出这种选择倾向。在暴露底物1h,然后间隔60min后,雌性田鼠比雄性田鼠表现出更明显的选择倾向。以上结果表明不同种类和性别的田鼠对气味表现出不同的识别模式。这种差异可能表明不同种具有木同的社会组织、生态和生活史对策。     

Calbindin D28k, parvalbumin, and calretinin immunoreactivity in the main and accessory olfactory bulbs of the gray short-tailed opossum, Monodelphis domestica

The vertebrate main and accessory olfactory bulbs (MOB and AOB) are the first synaptic sites in the olfactory pathways. The MOB is a cortical structure phylogenetically well conserved in its laminar structure and overall synaptic organization, while the AOB has significant species variation in size. In order to better understand signal processing in the two olfactory systems and the species differences, immunocytochemical staining and analysis were done of the neuronal expression patterns of the calcium-binding proteins calbindin D28k (CB), parvalbumin (PV), and calretinin (CR) in the MOB and AOB in a marsupial species, the gray short-tailed opossum, Monodelphis domestica. In the MOB, antibody to CB labeled periglomerular cells, superficial short axon cells / Van Gehuchten cells; antibody to PV labeled Van Gehuchten cells; and antibody to CR immunostained periglomerular cells, superficial short axon cells / Van Gehuchten cells, and granule cells. In the AOB, CB immunoreactivity was detected in periglomerular cells and a subpopulation of granule cells; antibody to PV labeled the superficial short axon cells / Van Gehuchten cells and granule cells; and antibody to CR labeled a small number of periglomerular cells, superficial short axon cells / Van Gehuchten cells, and granule cells. These results showed that the patterns of CB, PV, and CR expression differ in the opossum main and accessory olfactory bulbs and differ from that in other animal species. These varying patterns of neuronal immunostaining may be related to the different functions of the main and accessory olfactory bulbs and to the differing signal processing features.     

Postnatal developmental expression of calbindin, calretinin and parvalbumin in mouse main olfactory bulb

The distribution of calbindin, calretinin and parvalbumin during the development of the mouse main olfactory bulb (MOB) was studied using immunohistochemistry techniques. The results are as follows:(1) calbindin-immunoreactive profiles were mainly located in the glomerular layer, and few large calbindin-immunoreactive cells were found in the subependymal layer of postnatal day 10 (P10) to postnatal day 40 (P40) mice; (2) no calbindin was detected in the mitral cell layer at any stage; (3) calretinin-immunoreactive profiles were present in all layers of the main olfactory bulb at all stages, especially in the olfactory nerve layer, glomerular layer and granule cell layer; (4) parvalbumin-immunoreactive profiles were mainly located in the external plexiform layer (except for P10 mice); (5) weakly stained parvalbumin-immunoreactive profiles were present in the glomerular layer at all stages; and (6) no parvalbumin was detected in the mitral cell layer at any stage.     

Different profiles of main and accessory olfactory bulb mitral/tufted cell projections revealed in mice using an anterograde tracer and a whole-mount, flattened cortex preparation

A whole-mount, flattened cortex preparation was developed to compare profiles of axonal projections from main olfactory bulb (MOB) and accessory olfactory bulb (AOB) mitral and tufted (M/T) cells. After injections of the anterograde tracer, Phaseolus vulgaris leucoagglutinin, mapping of labeled axons using a Neurolucida system showed that M/T cells in the AOB sent axons primarily to the medial and posterior lateral cortical amygdala, with minimal branching into the piriform cortex. By contrast, M/T cells in the MOB displayed a network of collaterals that branched off the primary axon at several levels of the lateral olfactory tract (LOT). Collaterals emerging from the LOT into the anterior piriform cortex were often observed crossing into the posterior piriform cortex. M/T cells in the dorsal MOB extended fewer collaterals from the primary axon in the rostral LOT than did M/T cells from the anterior or ventral MOB. MOB M/T cells that projected to the medial amygdala did not do so exclusively, also sending collaterals to the anterior cortical amygdala as well as to olfactory cortical regions. This arrangement may be related to the ability of social experience to modify the response of mice to volatile pheromones detected by the main olfactory system.     

The postnatal development of the main olfactory bulb of the rat

The postnatal development from birth to 1 year of the main olfactory bulb was examined quantitatively. The volume of the main olfactory bulb increased over seven-fold by day 30 and remained unchanged thereafter. During the same period the volume of the granular layer increased 18-fold and the mean areas of the olfactory glomeruli increased seven-fold. The mean areas of mitral cell perikarya doubled between the neonatal and juvenile periods. The total number of the mitral cells, however, declined during the first three postnatal weeks. In the internal granular layer of the main olfactory bulb, 89% of the granule cells were acquired postnatally. Much of the cellular gain occurred during the first 3 weeks, with the period of maximum acquisition between days 8 and 14. The number of subependymal cells, the precursors of granule cells, reached a peak at 12 days and gradually declined. But some primitive cells could still be found at one year of age and there was an increase in the total number of granule cells beyond day 30. The mean nuber of internal granular layer cells in a single main olfactory bulb of adult rats was about 5 X 10(6); the number of mitral cells about 4 X 10(4). In the animals injected with 3H-thymidine on day 20 and killed 2 h after injection a small but significant proportion of cells was labelled in the subependymal layer but few in the internal granular layer. In the animals killed 20 and 40 days after injection there was a 10--11-fold rise in the proportion of labelled internal granular layer cells. The proportion of labelled internal granular layer cells decreased in longer survival groups but the total number of labelled cells remained the same, even in year-old animals. However, the total number of internal granular layer cells in the sections examined increased with age.     

Activity-dependent adjustments of the inhibitory network in the olfactory bulb following early postnatal deprivation

The first-order sensory relay for olfactory processing, the main olfactory bulb (MOB), retains the ability to acquire new interneurons throughout life. It is therefore a particularly appropriate region for studying the role of experience in sculpting neuronal networks. We found that nostril closure decreased the number of newborn granule cells in the MOB, the complexity of their dendritic arborization, and their spine density, without affecting the preexisting population of granule cells. Accordingly, the frequency of miniature synaptic inhibitory events received by mitral cells was reduced. However, due to a compensatory increase in newborn granule cell excitability, action potential-dependent GABA release was dramatically enhanced, thus counteracting the reduction in spine density and leading to an unaltered synchronization of mitral cell firing activity. Together, this study reveals a unique form of adaptive response brought about exclusively by the cohort of newborn cells and used to maintain normal functioning of the MOB.     

Comparison of Changes in GAD65 and GAD67 Immunoreactivity and Levels in the Gerbil Main Olfactory Bulb Induced by Transient Ischemia

In the present study, we investigated changes in glutamate decarboxylase 65 (GAD65) and GAD67 immunoreactivity and protein levels in the main olfactory bulb (MOB) after 5 min of transient forebrain ischemia in gerbils. GAD65 immunoreactivity in the sham-operated group was shown in neurons and neuropil except for the somata of granule cells. GAD65 immunoreactivity was increased in neurons in the external plexiform layer 60 days after ischemia, and in mitral cells 30 and 60 days after ischemia. GAD67 immunoreactivity in the sham-operated group was shown in periglomerular cells, neuron in the external plexiform layer and granule cells with neuropil. GAD67 immunoreactivity in periglomerular cells was increased 10, 45 and 60 days after ischemia. GAD67 immunoreactivity in neurons in the external plexiform layer was increased 10 and 15 days after ischemia. Mitral cells showed strong GAD67 immunoreactivity 10 days after ischemia. However, GAD67 immunoreactivity in the granule cells was not changed with time after ischemia. In Western blot analysis for GAD65 and GAD67 protein levels in the ischemic gerbil MOB, GAD65 level was not changed after ischemia; GAD67 level was increased 10 days after ischemia. These results suggest that transient ischemia causes changes in GAD65 and GAD67 immunoreactivity in the gerbil MOB, and this change may induce a malfunction in olfaction after an ischemic insult. Ki-Yeon Yoo and In Koo Hwang equally contributed to this article.     

Comparison of Newly Generated Doublecortin-immunoreactive Neuronal Progenitors in the Main Olfactory Bulb among Variously Aged Gerbils

In the present study, we investigated age-related differences in neuronal progenitors in the gerbil main olfactory bulb (MOB) using doublecortin (DCX), a marker for neuronal progenitors which differentiate into neurons in the brain. No difference in the number of neuronal nuclei (NeuN)-immunoreactive neurons was found in the MOB at variously aged gerbils. At postnatal month (PM) 1, DCX immunoreaction was detected in all layers of the MOB except for the olfactory nerve layer. At this time point, DCX-immunoreactive cells (neuronal progenitors) were very abundant; however, they did not have fully developed-processes. From PM 3, the number of DCX-immunoreactive neuronal progenitors was decreased with age. At PM 6, DCX-immunoreactive cells showed very well-developed processes. In western blot analysis, DCX protein level in the MOB was highest at PM 1. Thereafter, levels of DCX protein were decreased with age. In the subventricular zone of the lateral ventricle, the number of Ki-67-immunoractive cells (proliferating cells) was also significantly decreased with age. In addition, increases of α-synuclein-immunoreactive structures were observed in the MOB with age. These results suggest that decrease in DCX-immunoreactive neuronal progenitors and its protein levels in the MOB with age may be associated with reduction of cell proliferation in the SVZ and with an increase in α-synuclein in the MOB.     

Slice culture of the olfactory bulb of Xenopus laevis tadpoles

We report on the development of a slice culture of amphibian brain tissue. In particular, we cultured slices from Xenopus laevis tadpoles that contain the olfactory mucosae, the olfactory nerves, the olfactory bulb and the telencephalon. During 6 days in roller tubes the slices flattened, starting from 250 microm and decreasing to approximately 40 microm, corresponding to about three cell layers. Dendritic processes could be followed over distances as long as 200 microm. Neurons in the cultured slice could be recorded using the patch clamp technique and simultaneously imaged using an inverted laser scanning microscope. We characterized the main neuron types of the olfactory bulb, i.e. mitral cells and granule cells, by correlating their typical morphological features in the acute slice with the electrophysiological properties in both the acute slice and slice culture. This correlation allowed unambiguous identification of mitral cells and granule cells in the slice culture.