Understanding Odor Information Segregation in the Olfactory Bulb by Means of Mitral and Tufted Cells

Odor identification is one of the main tasks of the olfactory system. It is performed almost independently from the concentration of the odor providing a robust recognition. This capacity to ignore concentration information does not preclude the olfactory system from estimating concentration itself. Significant experimental evidence has indicated that the olfactory system is able to infer simultaneously odor identity and intensity. However, it is still unclear at what level or levels of the olfactory pathway this segregation of information occurs. In this work, we study whether this odor information segregation is performed at the input stage of the olfactory bulb: the glomerular layer. To this end, we built a detailed neural model of the glomerular layer based on its known anatomical connections and conducted two simulated odor experiments. In the first experiment, the model was exposed to an odor stimulus dataset composed of six different odorants, each one dosed at six different concentrations. In the second experiment, we conducted an odor morphing experiment where a sequence of binary mixtures going from one odor to another through intermediate mixtures was presented to the model. The results of the experiments were visualized using principal components analysis and analyzed with hierarchical clustering to unveil the structure of the high-dimensional output space. Additionally, Fisher''s discriminant ratio and Pearson''s correlation coefficient were used to quantify odor identity and odor concentration information respectively. Our results showed that the architecture of the glomerular layer was able to mediate the segregation of odor information obtaining output spiking sequences of the principal neurons, namely the mitral and external tufted cells, strongly correlated with odor identity and concentration, respectively. An important conclusion is also that the morphological difference between the principal neurons is not key to achieve odor information segregation.     

嗅球僧帽细胞具有编码气味空间信息的能力

刺激源的方位是刺激的重要特性之一.行为学的研究发现,动物能够利用气味到达左右鼻腔的时间差和强度差信息对气味方位进行感知,但作为嗅觉系统第一神经中枢的嗅球,是否具有利用两侧鼻间差信息对气味方位进行编码的能力一直受到质疑.为探讨该问题,在本研究中通过比较嗅球中84个僧帽细胞对同侧气味刺激、对侧气味刺激以及对侧气味刺激略先于同侧气味刺激时的反应,发现有29个僧帽细胞可被同侧气味所兴奋,其中18个虽然对对侧气味刺激不反应,但对侧气味的存在却能显著降低其对同侧气味刺激的反应.另外,50个僧帽细胞在只给予同侧或对侧气味刺激时不反应,但其中11个在对侧刺激略先于同侧刺激的方式给出气味时,表现出明显的兴奋性反应.我们的研究结果一方面提示僧帽细胞具有编码气味到达两个鼻腔的时间差,或气味源位置信息的能力;另一方面也表明对侧刺激不仅能对同侧嗅球僧帽细胞产生抑制效应,还可能存在目前还不明确的机制而产生兴奋效应.     

Specific Entrainment of Mitral Cells during Gamma Oscillation in the Rat Olfactory Bulb

Local field potential (LFP) oscillations are often accompanied by synchronization of activity within a widespread cerebral area. Thus, the LFP and neuronal coherence appear to be the result of a common mechanism that underlies neuronal assembly formation. We used the olfactory bulb as a model to investigate: (1) the extent to which unitary dynamics and LFP oscillations can be correlated and (2) the precision with which a model of the hypothesized underlying mechanisms can accurately explain the experimental data. For this purpose, we analyzed simultaneous recordings of mitral cell (MC) activity and LFPs in anesthetized and freely breathing rats in response to odorant stimulation. Spike trains were found to be phase-locked to the gamma oscillation at specific firing rates and to form odor-specific temporal patterns. The use of a conductance-based MC model driven by an approximately balanced excitatory-inhibitory input conductance and a relatively small inhibitory conductance that oscillated at the gamma frequency allowed us to provide one explanation of the experimental data via a mode-locking mechanism. This work sheds light on the way network and intrinsic MC properties participate in the locking of MCs to the gamma oscillation in a realistic physiological context and may result in a particular time-locked assembly. Finally, we discuss how a self-synchronization process with such entrainment properties can explain, under experimental conditions: (1) why the gamma bursts emerge transiently with a maximal amplitude position relative to the stimulus time course; (2) why the oscillations are prominent at a specific gamma frequency; and (3) why the oscillation amplitude depends on specific stimulus properties. We also discuss information processing and functional consequences derived from this mechanism.     

Prolonged Intracellular Na+ Dynamics Govern Electrical Activity in Accessory Olfactory Bulb Mitral Cells

Persistent activity has been reported in many brain areas and is hypothesized to mediate working memory and emotional brain states and to rely upon network or biophysical feedback. Here, we demonstrate a novel mechanism by which persistent neuronal activity can be generated without feedback, relying instead on the slow removal of Na⁺ from neurons following bursts of activity. We show that mitral cells in the accessory olfactory bulb (AOB), which plays a major role in mammalian social behavior, may respond to a brief sensory stimulation with persistent firing. By combining electrical recordings, Ca²⁺ and Na⁺ imaging, and realistic computational modeling, we explored the mechanisms underlying the persistent activity in AOB mitral cells. We found that the exceptionally slow inward current that underlies this activity is governed by prolonged dynamics of intracellular Na⁺ ([Na⁺]_i), which affects neuronal electrical activity via several pathways. Specifically, elevated dendritic [Na⁺]_i reverses the Na⁺-Ca²⁺ exchanger activity, thus modifying the [Ca²⁺]_i set-point. This process, which relies on ubiquitous membrane mechanisms, is likely to play a role in other neuronal types in various brain regions.     

Influence of Olfactory Epithelium on Mitral/Tufted Cell Dendritic Outgrowth

Stereotypical connections between olfactory sensory neuron axons and mitral cell dendrites in the olfactory bulb establish the first synaptic relay for olfactory perception. While mechanisms of olfactory sensory axon targeting are reported, molecular regulation of mitral cell dendritic growth and refinement are unclear. During embryonic development, mitral cell dendritic distribution overlaps with olfactory sensory axon terminals in the olfactory bulb. In this study, we investigate whether olfactory sensory neurons in the olfactory epithelium influence mitral cell dendritic outgrowth in vitro. We report a soluble trophic activity in the olfactory epithelium conditioned medium which promotes mitral/tufted cell neurite outgrowth. While the trophic activity is present in both embryonic and postnatal olfactory epithelia, only embryonic but not postnatal mitral/tufted cells respond to this activity. We show that BMP2, 5 and 7 promote mitral/tufted cells neurite outgrowth. However, the BMP antagonist, Noggin, fails to neutralize the olfactory epithelium derived neurite growth promoting activity. We provide evidence that olfactory epithelium derived activity is a protein factor with molecular weight between 50–100 kD. We also observed that Follistatin can effectively neutralize the olfactory epithelium derived activity, suggesting that TGF-beta family proteins are involved to promote mitral/tufted dendritic elaboration.     

Transsynaptic Regulation of Olfactory Bulb Catecholamines in Mice and Rats

Norepinephrine (NE), dopamine (DA), 3,4-dihydroxyphenylalanine (DOPA), and 3,4-dihydroxyphenylacetic acid (DOPAC) were measured simultaneously by high performance liquid chromatography with electrochemical detection in extracts of olfactory bulbs at various intervals after chemical or surgical deafferentation. Chemical deafferentation of mice by intranasal irrigation with Triton X-100 or of rats by olfactory axotomy resulted in a rapid progressive decline of DA and DOPAC and an associated rise in NE in the olfactory bulb. However, after several weeks, these values returned to prelesion levels concomitant with reinnervation of the bulb by the afferent neurons. In contrast, deafferentation by procedures known to prevent reinnervation of the bulb by the afferent chemoreceptor neurons (i.e., a ZnSo4 solution in mice or a surgical procedure in rats) completely blocked the return to pre-lesion values of DA, DOPAC, and NE. The specificity of these effects was demonstrated by the inability of intranasal administration of the neurotoxin 6-hydroxydopamine to alter DA levels, resulting instead in a significant decline in olfactory bulb NE content. These data demonstrate that the DA content of the olfactory bulb can be influenced by either chemical or surgical modulation of the afferent pathway in two different species. This offers additional support for our hypothesis of transsynaptic regulation of intrinsic DA neurons of the bulb by the afferent olfactory chemoreceptor neurons.     

猫嗅球外丛层和僧帽细胞层年龄相关形态学变化

分别用Nissl法及免疫组织化学ABC法标记青、老年猫嗅球中嗅觉二级神经元和外丛层胶质细胞,显微镜下观察其分布并计数,对嗅觉二级神经元胞体直径和外丛层厚度进行测量,比较其年龄相关性变化,研究神经元与胶质细胞之间的关系,探讨老年性嗅觉功能衰退的相关神经机理。结果显示,老年猫嗅觉二级神经元胞体直径和分布密度均有不同程度的显著性下降(P<0.05);外丛层厚度变化不明显(P>0.05);外丛层胶质细胞特别是星形胶质细胞显著性增生(P<0.05)。表明在衰老过程中嗅觉二级神经元有丢失,并呈现功能下降,可能是老年性嗅觉功能衰退的原因之一。同时外丛层胶质细胞增生以进一步保护神经元,延缓其衰老。     

Olfaction in Lamprey Pallium Revisited—Dual Projections of Mitral and Tufted Cells

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Gene Expression Changes in the Olfactory Bulb of Mice Induced by Exposure to Diesel Exhaust Are Dependent on Animal Rearing Environment

There is an emerging concern that particulate air pollution increases the risk of cranial nerve disease onset. Small nanoparticles, mainly derived from diesel exhaust particles reach the olfactory bulb by their nasal depositions. It has been reported that diesel exhaust inhalation causes inflammation of the olfactory bulb and other brain regions. However, these toxicological studies have not evaluated animal rearing environment. We hypothesized that rearing environment can change mice phenotypes and thus might alter toxicological study results. In this study, we exposed mice to diesel exhaust inhalation at 90 µg/m³, 8 hours/day, for 28 consecutive days after rearing in a standard cage or environmental enrichment conditions. Microarray analysis found that expression levels of 112 genes were changed by diesel exhaust inhalation. Functional analysis using Gene Ontology revealed that the dysregulated genes were involved in inflammation and immune response. This result was supported by pathway analysis. Quantitative RT-PCR analysis confirmed 10 genes. Interestingly, background gene expression of the olfactory bulb of mice reared in a standard cage environment was changed by diesel exhaust inhalation, whereas there was no significant effect of diesel exhaust exposure on gene expression levels of mice reared with environmental enrichment. The results indicate for the first time that the effect of diesel exhaust exposure on gene expression of the olfactory bulb was influenced by rearing environment. Rearing environment, such as environmental enrichment, may be an important contributive factor to causation in evaluating still undefined toxic environmental substances such as diesel exhaust.     

Photoperiod Mediated Changes in Olfactory Bulb Neurogenesis and Olfactory Behavior in Male White-Footed Mice (Peromyscus leucopus)

Brain plasticity, in relation to new adult mammalian neurons generated in the subgranular zone of the hippocampus, has been well described. However, the functional outcome of new adult olfactory neurons born in the subventricular zone of the lateral ventricles is not clearly defined, as manipulating neurogenesis through various methods has given inconsistent and conflicting results in lab mice. Several small rodent species, including Peromyscus leucopus, display seasonal (photoperiodic) brain plasticity in brain volume, hippocampal function, and hippocampus-dependent behaviors; plasticity in the olfactory system of photoperiodic rodents remains largely uninvestigated. We exposed adult male P. leucopus to long day lengths (LD) and short day lengths (SD) for 10 to 15 weeks and then examined olfactory bulb cell proliferation and survival using the thymidine analog BrdU, olfactory bulb granule cell morphology using Golgi-Cox staining, and behavioral investigation of same-sex conspecific urine. SD mice did not differ from LD counterparts in granular cell morphology of the dendrites or in dendritic spine density. Although there were no differences due to photoperiod in habituation to water odor, SD mice rapidly habituated to male urine, whereas LD mice did not. In addition, short day induced changes in olfactory behavior were associated with increased neurogenesis in the caudal plexiform and granule cell layers of the olfactory bulb, an area known to preferentially respond to water-soluble odorants. Taken together, these data demonstrate that photoperiod, without altering olfactory bulb neuronal morphology, alters olfactory bulb neurogenesis and olfactory behavior in Peromyscus leucopus.     

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.     

外周输入依赖的嗅球颗粒细胞的突触结构可塑性

活动依赖的突触结构可塑性是学习和记忆的基础.哺乳动物,尤其是啮齿类动物,具有高度发达的嗅觉系统和惊人的气味学习和记忆能力.本研究以CNGA2敲除而导致外周输入缺失的小鼠为模型,研究嗅球内活动依赖的突触结构可塑性.利用特异性的突触前和突触后标记物,发现外周输入缺失减少了突触标记蛋白突触素(synaptophysin)和抑制性突触标记蛋白桥蛋白(gephyrin)在嗅球外网状层和颗粒细胞层中的表达;兴奋性突触标记蛋白囊泡谷氨酸转运蛋白1(VGluT1)的表达水平只在外网状层中有显著下降,而在颗粒细胞层中没有明显变化.进一步通过活体质粒电转标记嗅球颗粒细胞后发现,CNGA2敲除小鼠颗粒细胞上位于外网状层中的远端树突棘密度显著减小,而位于颗粒细胞层中的近端树突棘密度没有明显变化.这些结果表明颗粒细胞上的树-树突触具有对外周活动依赖的结构可塑性,而轴-树突触则无.     

Olfactory Enrichment Influences Adult Neurogenesis Modulating GAD67 and Plasticity-Related Molecules Expression in Newborn Cells of the Olfactory Bulb

The olfactory bulb (OB) is a highly plastic region of the adult mammalian brain characterized by continuous integration of inhibitory interneurons of the granule (GC) and periglomerular cell (PGC) types. Adult-generated OB interneurons are selected to survive in an experience-dependent way but the mechanisms that mediate the effects of experience on OB neurogenesis are unknown. Here we focus on the new-generated PGC population which is composed by multiple subtypes. Using paradigms of olfactory enrichment and/or deprivation combined to BrdU injections and quantitative confocal immunohistochemical analyses, we studied the effects of olfactory experience on adult-generated PGCs at different survival time and compared PGC to GC modulation. We show that olfactory enrichment similarly influences PGCs and GCs, increasing survival of newborn cells and transiently modulating GAD67 and plasticity-related molecules expression. However, PGC maturation appears to be delayed compared to GCs, reflecting a different temporal dynamic of adult generated olfactory interneuron integration. Moreover, olfactory enrichment or deprivation do not selectively modulate the survival of specific PGC phenotypes, supporting the idea that the integration rate of distinct PGC subtypes is independent from olfactory experience.     

Effects of Lead Exposure on Nitric Oxide-associated Gene Expression in the Olfactory Bulb of Mice

Lead (Pb) is known to have toxic effects on the brain; however, data regarding its specific toxic effects on the olfactory bulb are lacking. Therefore, we investigated the relationship between acute Pb exposure and alterations in gene expression associated with the nitric oxide signaling pathway in the olfactory bulb of mice. After administration of Pb (intraperitoneal injections of 1 or 10 mg/kg Pb(CH₃CO₂)₂ · 3H₂O once per day for 4 days), body weight, motor activity, and gene expression in the olfactory bulb of mice were examined. High doses of Pb resulted in significant decreases in body weight, but motor coordination was not significantly altered until 11 days after the end of Pb treatment. The expression patterns of dimethylarginine dimethylaminohydrolase 1 (Ddah1), superoxide dismutase 1 (Sod1), and superoxide dismutase (Ccs) were increased, whereas expression of the Stratifin (Sfn) gene was significantly decreased following treatment with 10 mg/kg Pb. The expression patterns of nitric oxide synthases at the mRNA and protein levels, however, were not significantly altered by treatment with 10 mg/kg Pb. These findings indicate that Pb-induced neurotoxicity may be modulated in part by the expression of Ddah1, Sod1, Ccs, and Sfn in the olfactory bulb.     

The Efferent Connections of the Olfactory Bulb and Accessory Olfactory Bulb in the Snakes,Thamnophis sirtalis and Thamnophis radix

The efferent connections of the olfactory bulb and accessory olfactory bulb of two species of garter snakes, Thamnophis sirtalis and T. radix were studied with experimental anterograde degeneration techniques. Axons of cells located in the olfactory bulb terminate ipsilaterally in all parts of the anterior olfactory nucleus, olfactory tubercle and lateral pallium. In addition, some axons enter the ipsilateral stria medullaris thalami, cross the midline in the habenular commissure, enter the contralateral stria medullaris thalami and terminate in the contralateral lateral pallium. The axons of cells in the accessory olfactory bulb course through the telencephalon completely separated from the fibers of olfactory bulb origin and terminate predominantly in the nucleus sphericus. These results confirm previous reports of the separation between the central projections of the olfactory and vomeronasal systems in a variety of vertebrates. The totality of the separation between these two systems coupled with the extensive development of the vomeronasal-accessory bulb system in these snakes suggests that they may be ideal subjects for further research on the functional significance of the vomeronasal system.     

He—Ne激光照射对离体小鼠巨噬细胞膜电位的影响

低功率Ｈｅ－Ｎｅ激光照射引起细胞膜电位的改变,为激光照射治疗提供理论依据。方法不同能量密度的Ｈｅ－Ｎｅ激光照射离体在噬细胞,用图像分析系统检测Ｃｙａｎｉｎｅ荧光染色的巨噬细胞膜电位的变化。     

The Olfactory Bulb in Newborn Piglet Is a Reservoir of Neural Stem and Progenitor Cells

The olfactory bulb (OB) periventricular zone is an extension of the forebrain subventricular zone (SVZ) and thus is a source of neuroprogenitor cells and neural stem cells. While considerable information is available on the SVZ-OB neural stem cell (NSC)/neuroprogenitor cell (NPC) niche in rodents, less work has been done on this system in large animals. The newborn piglet is used as a preclinical translational model of neonatal hypoxic-ischemic brain damage, but information about the endogenous sources of NSCs/NPCs in piglet is needed to implement endogenous or autologous cell-based therapies in this model. We characterized NSC/NPC niches in piglet forebrain and OB-SVZ using western blotting, histological, and cell culture methods. Immunoblotting revealed nestin, a NSC/NPC marker, in forebrain-SVZ and OB-SVZ in newborn piglet. Several progenitor or newborn neuron markers, including Dlx2, musashi, doublecortin, and polysialated neural cell adhesion molecule were also detected in OB-SVZ by immunoblotting. Immunohistochemistry confirmed the presence of nestin, musashi, and doublecortin in forebrain-SVZ and OB-SVZ. Bromodeoxyuridine (BrdU) labeling showed that the forebrain-SVZ and OB-SVZ accumulate newly replicated cells. BrdU-positive cells were immunolabeled for astroglial, oligodendroglial, and neuronal markers. A lateral migratory pathway for newly born neuron migration to primary olfactory cortex was revealed by BrdU labeling and co-labeling for doublecortin and class III β tubulin. Isolated and cultured forebrain-SVZ and OB-SVZ cells from newborn piglet had the capacity to generate numerous neurospheres. Single cell clonal analysis of neurospheres revealed the capacity for self-renewal and multipotency. Neurosphere-derived cells differentiated into neurons, astrocytes, and oligodendrocytes and were amenable to permanent genetic tagging with lentivirus encoding green fluorescent protein. We conclude that the piglet OB-SVZ is a reservoir of NSCs and NPCs suitable to use in autologous cell therapy in preclinical models of neonatal/pediatric brain injury.