Origin of basal activity in mammalian olfactory receptor neurons

Mammalian odorant receptors form a large, diverse group of G protein-coupled receptors that determine the sensitivity and response profile of olfactory receptor neurons. But little is known if odorant receptors control basal and also stimulus-induced cellular properties of olfactory receptor neurons other than ligand specificity. This study demonstrates that different odorant receptors have varying degrees of basal activity, which drives concomitant receptor current fluctuations and basal action potential firing. This basal activity can be suppressed by odorants functioning as inverse agonists. Furthermore, odorant-stimulated olfactory receptor neurons expressing different odorant receptors can have strikingly different response patterns in the later phases of prolonged stimulation. Thus, the influence of odorant receptor choice on response characteristics is much more complex than previously thought, which has important consequences on odor coding and odor information transfer to the brain.     

Adjusting neurophysiological computations in the adult olfactory bulb

The olfactory bulb receives signals from olfactory sensory neurons and conveys them to higher centers. The mapping of the sensory inputs generates a reproducible spatial pattern in the glomerular layer of the olfactory bulb for each odorant. Then, this restricted activation is transformed into highly distributed patterns by lateral interactions between relay neurons and local interneurons. Thus, odor information processing requires the spatial patterning of both sensory inputs and synaptic interactions. In other words, odor representation is highly dynamic and temporally orchestrated. Here, we describe how the local inhibitory network shapes the global oscillations and the precise synchronization of relay neurons. We discuss how local inhibitory interneurons transpose the spatial dimension into temporal patterning. Remarkably, this transposition is not fixed but highly flexible to continuously optimize olfactory information processing.     

Olfaction and odor discrimination are mediated by the C. elegans guanylyl cyclase ODR-1

Animals in complex environments must discriminate between salient and uninformative sensory cues. Caenorhabditis elegans uses one pair of olfactory neurons called AWC to sense many different odorants, yet the animal can distinguish each odorant from the others in discrimination assays. We demonstrate that the transmembrane guanylyl cyclase ODR-1 is essential for responses to all AWC-sensed odorants. ODR-1 appears to be a shared signaling component downstream of odorant receptors. Overexpression of ODR-1 protein indicates that ODR-1 can influence odor discrimination and adaptation as well as olfaction. Adaptation to one odorant, butanone, is disrupted by ODR-1 overexpression. Olfactory discrimination is also disrupted by ODR-1 overexpression, probably by overproduction of the shared second messenger cGMP. We propose that AWC odorant signaling pathways are insulated to permit odor discrimination.     

BIG-2 mediates olfactory axon convergence to target glomeruli

Olfactory sensory neurons expressing a given odorant receptor converge axons onto a few topographically fixed glomeruli in the olfactory bulb, leading to establishment of the odor map. Here, we report that BIG-2/contactin-4, an axonal glycoprotein belonging to the immunoglobulin superfamily, is expressed in a subpopulation of mouse olfactory sensory neurons. A mosaic pattern of glomerular arrangement is observed with strongly BIG-2-positive, weakly positive, and negative axon terminals in the olfactory bulb, which is overlapping but not identical with those of Kirrel2 and ephrin-A5. There is a close correlation between the BIG-2 expression level and the odorant receptor choice in individual sensory neurons. In BIG-2-deficient mice, olfactory sensory neurons expressing a given odorant receptor frequently innervate multiple glomeruli at ectopic locations. These results suggest that BIG-2 is one of the axon guidance molecules crucial for the formation and maintenance of functional odor map in the olfactory bulb.     

All in a sniff: olfaction as a model for active sensing

Sensation is an active process involving the sampling and central processing of external stimuli selectively in space and time. Olfaction in particular depends strongly on active sensing due to the fact that-at least in mammals-inhalation of air into the nasal cavity is required for odor detection. This seemingly simple first step in odor sensation profoundly shapes nearly all aspects of olfactory system function, from the distribution of odorant receptors to the functional organization of central processing to the perception of odors. The dependence of olfaction on inhalation also allows for profound modulation of olfactory processing by changes in odor sampling strategies in coordination with attentional state and sensory demands. This review discusses the role of active sensing in shaping olfactory system function at multiple levels and draws parallels with other sensory modalities to highlight the importance of an active sensing perspective in understanding how sensory systems work in the behaving animal.     

Subunit contributions to insect olfactory receptor function: channel block and odorant recognition

Insect olfactory receptors are heteromeric ligand-gated ion channels composed of at least one common subunit (Orco) and at least one subunit that confers odorant specificity. Little is known about how individual subunits contribute to the structure and function of the olfactory receptor complex. We expressed insect olfactory receptors in Xenopus oocytes to investigate 2 functional features, ion channel block and odorant recognition. The sensitivity of Drosophila olfactory receptors to inhibition by ruthenium red, a cation channel blocker, varied widely when different specificity subunits were present, suggesting that the specificity subunits contribute to the structure of the ion pore. Olfactory receptors formed by Dmel\Or35a and Orco subunits from several different species displayed highly similar odorant response profiles, suggesting that the Orco subunit does not contribute to the structure of the odorant-binding site. We further explored odorant recognition by conducting a detailed examination of the odorant specificity Dmel\Or67a + Dmel\Orco, a receptor that responds to aromatic structures. This screen identified agonists, partial agonists, and an antagonist of Dmel\Or67a + Dmel\Orco. Our findings favor specific subunit arrangements within the olfactory receptor complex and provide a preliminary odorophore for an olfactory receptor, offering a useful foundation for future exploration of insect olfactory receptor structure.     

Ligand-specific dose-response of heterologously expressed olfactory receptors.

Primary olfactory neuronal cultures exposed to odorant stimulation have previously exhibited concentration-related effects in terms of intracellular cAMP levels and adenylate cyclase activity [Ronnett, G.V., Parfitt, D.J., Hester, L.D. & Snyder, S.H. (1991) PNAS88, 2366-2369]. Maximal stimulation occurred for intermediate concentrations, whereas AC activity declined for both low and high odorant concentrations. We suspected that this behavior might be ascribed to the intrinsic response of the first molecular species concerned by odorant detection, i.e. the olfactory receptor itself. In order to check this hypothesis, we developed an heterologous expression system in mammalian cells to characterize the functional response of receptors to odorants. Two mammalian olfactory receptors were used to initiate the study, the rat I7 olfactory receptor and the human OR17-40 olfactory receptor. The cellular response of transfected cells to an odorant stimulation was tested by a spectrofluorimetric intracellular calcium assay, and proved in all cases to be dose-dependent for the known ligands of these receptors, with an optimal response for intermediate concentrations. Further experiments were carried out with the rat I7 olfactory receptor, for which the sensitivity to an odorant, indicated by the concentration yielding the optimal calcium response, depended on the carbon chain length of the aldehydic odorant. The response is thus both ligand-specific and dose-dependent. We thus demonstrate that a differential dose-response originates from the olfactory receptor itself, which is thus capable of efficient discrimination between closely related agonists.     

The mouse eugenol odorant receptor: structural and functional plasticity of a broadly tuned odorant binding pocket

Molecular interactions of odorants with their olfactory receptors (ORs) are of central importance for the ability of the mammalian olfactory system to detect and discriminate a vast variety of odors with a limited set of receptors. How a particular OR binds and distinguishes different odorant molecules remains largely unknown on a structural basis. Here we investigated this question for the mouse eugenol receptor (mOR-EG). By screening a large odorant library, we discovered a wide range of chemical structures activating the receptor in heterologous mammalian cells. Potent agonists comprise (i) benzene, (ii) cyclohexane, or (iii) polycyclic structures substituted with alcohol, aldehyde, keto, ether, or esterified carboxylic groups. To detect those amino acids within the receptor that are in contact with a particular bound odorant molecule, we investigated how distinct mOR-EG point mutants were activated by the different odorant agonists found for the wild-type receptor. We identified 11 amino acids as a part of the receptor's ligand binding pocket. Molecular modeling predicted 10 of these residues in transmembrane helices TM3-TM6 and one in the extracellular loop between TM2 and TM3. These amino acids participate in odorant binding with variable importance depending on the type of odorant, revealing functional "fingerprints" of ligand-receptor interactions.     

Neural circuit mechanisms for pattern detection and feature combination in olfactory cortex

Odors are initially encoded in the brain as a set of distinct physicochemical characteristics but are ultimately perceived as a unified sensory object--a "smell." It remains unclear how chemical features encoded by diverse odorant receptors and segregated glomeruli in the main olfactory bulb (MOB) are assembled into integrated cortical representations. Combining patterned optical microstimulation of MOB with in vivo electrophysiological recordings in anterior piriform cortex (PCx), we assessed how cortical neurons decode complex activity patterns distributed across MOB glomeruli. PCx firing was insensitive to single-glomerulus photostimulation. Instead, individual cells reported higher-order combinations of coactive glomeruli resembling odor-evoked sensory maps. Intracellular recordings revealed a corresponding circuit architecture providing each cortical neuron with weak synaptic input from a distinct subpopulation of MOB glomeruli. PCx neurons thus detect specific glomerular ensembles, providing an explicit neural representation of chemical feature combinations that are the hallmark of complex odor stimuli.     

Molecular determinants of odorant receptor function in insects

The olfactory system of Drosophila melanogaster provides a powerful model to study molecular and cellular mechanisms underlying function of a sensory system. In the 1970s Siddiqi and colleagues pioneered the application of genetics to olfactory research and isolated several mutant Drosophila with odorant-specific defects in olfactory behaviour, suggesting that odorants are detected differentially by the olfactory system. Since then basic principles of olfactory system function and development have emerged using Drosophila as a model. Nearly four decades later we can add computational methods to further our understanding of how specific odorants are detected by receptors. Using a comparative approach we identify two categories of short amino acid sequence motifs: ones that are conserved family-wide predominantly in the C-terminal half of most receptors, and ones that are present in receptors that detect a specific odorant, 4-methylphenol, found predominantly in the N-terminal half. The odorant-specific sequence motifs are predictors of phenol detection in Anopheles gambiae and other insects, suggesting they are likely to participate in odorant binding. Conversely, the family-wide motifs are expected to participate in shared functions across all receptors and a mutation in the most conserved motif leads to a reduction in odor response. These findings lay a foundation for investigating functional domains within odorant receptors that can lead to a molecular understanding of odor detection.     

Speed-accuracy tradeoff in olfaction

The basic psychophysical principle of speed-accuracy tradeoff (SAT) has been used to understand key aspects of neuronal information processing in vision and audition, but the principle of SAT is still debated in olfaction. In this study we present the direct observation of SAT in olfaction. We developed a behavioral paradigm for mice in which both the duration of odorant sampling and the difficulty of the odor discrimination task were controlled by the experimenter. We observed that the accuracy of odor discrimination increases with the duration of imposed odorant sampling, and that the rate of this increase is slower for harder tasks. We also present a unifying picture of two previous, seemingly disparate experiments on timing of odorant sampling in odor discrimination tasks. The presence of SAT in olfaction provides strong evidence for temporal integration in olfaction and puts a constraint on models of olfactory processing.     

昆虫气味受体研究进展

嗅觉在昆虫的多种行为中发挥关键作用。气味分子与嗅觉神经元树突上气味受体的结合,参与了昆虫嗅觉识别的初始过程。昆虫的嗅觉神经元表达两类气味受体： 一是传统气味受体,该类受体同源性较低,在少部分嗅觉神经元中表达; 二是Or83b家族受体,该类受体不感受气味,在不同昆虫间较为保守且在大多数嗅觉神经元中表达。目前,对于单个传统气味受体的气味分子配体特异性所知甚少; 对于Or83b家族受体,一般认为其可能具有将传统气味受体运送至嗅觉神经元树突膜上的功能。此外,有一些实验证据不支持昆虫气味受体为G蛋白偶联受体的观点。     

Odorant representations are modulated by intra- but not interglomerular presynaptic inhibition of olfactory sensory neurons

Input to the central nervous system from olfactory sensory neurons (OSNs) is modulated presynaptically. We investigated the functional organization of this inhibition and its role in odor coding by imaging neurotransmitter release from OSNs in slices and in vivo in mice expressing synaptopHluorin, an optical indicator of vesicle exocytosis. Release from OSNs was strongly suppressed by heterosynaptic, intraglomerular inhibition. In contrast, inhibitory connections between glomeruli mediated only weak lateral inhibition of OSN inputs in slices and did not do so in response to odorant stimulation in vivo. Blocking presynaptic inhibition in vivo increased the amplitude of odorant-evoked input to glomeruli but had little effect on spatial patterns of glomerular input. Thus, intraglomerular inhibition limits the strength of olfactory input to the CNS, whereas interglomerular inhibition plays little or no role. This organization allows for control of input sensitivity while maintaining the spatial maps of glomerular activity thought to encode odorant identity.     

Genetic and functional subdivision of the Drosophila antennal lobe

Olfactory systems confer the recognition and discrimination of a large number of structurally distinct odor molecules. Recent molecular analysis of odorant receptor (OR) genes and circuits has led to a model of odor coding in which a population of olfactory sensory neurons (OSNs) expressing a single OR converges upon a unique olfactory glomerulus. Activation of the OR can thus be read out by the activation of its cognate glomerulus. Drosophila is a powerful system in which to test this model because the entire repertoire of 62 ORs can be manipulated genetically. However, a complete understanding of how fly olfactory circuits are organized is lacking. Here, we present a nearly complete map of OR projections from OSNs to the antennal lobe (AL) in the fly brain. Four populations of OSNs coexpress two ORs along with Or83b, and a fifth expresses one OR and one gustatory receptor (GR) along with Or83b. One glomerulus receives coconvergent input from two separate populations of OSNs. Three ORs label sexually dimorphic glomeruli implicated in sexual courtship and are thus candidate Drosophila pheromone receptors. This olfactory sensory map provides an experimental framework for relating ORs to glomeruli and ultimately behavior.     

Learned odor discrimination in Drosophila without combinatorial odor maps in the antennal lobe

A unifying feature of mammalian and insect olfactory systems is that olfactory sensory neurons (OSNs) expressing the same unique odorant-receptor gene converge onto the same glomeruli in the brain [1-7]. Most odorants activate a combination of receptors and thus distinct patterns of glomeruli, forming a proposed combinatorial spatial code that could support discrimination between a large number of odorants [8-11]. OSNs also exhibit odor-evoked responses with complex temporal dynamics [11], but the contribution of this activity to behavioral odor discrimination has received little attention [12]. Here, we investigated the importance of spatial encoding in the relatively simple Drosophila antennal lobe. We show that Drosophila can learn to discriminate between two odorants with one functional class of Or83b-expressing OSNs. Furthermore, these flies encode one odorant from a mixture and cross-adapt to odorants that activate the relevant OSN class, demonstrating that they discriminate odorants by using the same OSNs. Lastly, flies with a single class of Or83b-expressing OSNs recognize a specific odorant across a range of concentration, indicating that they encode odorant identity. Therefore, flies can distinguish odorants without discrete spatial codes in the antennal lobe, implying an important role for odorant-evoked temporal dynamics in behavioral odorant discrimination.     

Computational Biology of Olfactory Receptors

Olfactory receptors, in addition to being involved in first step of the physiological processes that leads to olfaction, occupy an important place in mammalian genomes. ORs constitute super families in these genomes. Elucidating ol-factory receptor function at a molecular level can be aided by a computationally derived structure and an understanding of its interactions with odor molecules. Experimental functional analyses of olfactory receptors in conjunction with computational studies serve to validate findings and generate hypotheses. We present here a review of the research efforts in: creating computational models of olfactory receptors, identifying binding strategies for these receptors with odorant molecules, performing medium to long range simulation studies of odor ligands in the receptor binding region, and identifying amino acid positions within the receptor that are responsible for ligand-binding and olfactory receptor activation. Written as a primer and a teaching tool, this review will help researchers extend the methodologies described herein to other GPCRs.     

Retinoic acid receptor and CNGA2 channel signaling are part of a regulatory feedback loop controlling axonal convergence and survival of olfactory sensory neurons

Little is known about the identities and functions of extracellular signaling molecules that work in concert with neuronal activity to regulate refinement and maintenance of the mouse olfactory sensory map. We show that expression of a dominant negative retinoic acid receptor (RAR) in olfactory sensory neurons (OSNs) increased the number of glomeruli that incorrectly contained OSN axons expressing different odorant receptors. This phenotype became apparent postnatally, coincided with increased cell death, and was preceded by increased Neuropilin-1 and reduced Kirrel-2 expressions. Kirrel-2-mediated cell adhesion influences odorant receptor-specific axonal convergence and is regulated by odorant receptor signaling via the olfactory cyclic nucleotide-gated (CNG) ion channel. Accordingly, we found that inhibited RAR function correlated with reduced CNG channel expression. Naris occlusion experiments and analysis of CNG channel-deficient mice further indicated that RAR-regulated CNG channel levels influenced the intrinsic neuronal activity required for cell survival in the absence of odor stimulation. Finally, we showed that CNG channel activity regulated expression of the retinoic acid-degrading enzyme Cyp26B1. Combined, these results identify a novel homeostatic feedback mechanism involving retinoic acid metabolism and CNG channel activity, which influences glomerular homogeneity and maintenance of precisely connected OSNs.     

昆虫非典型嗅觉受体Orco的功能和分子结构研究进展

嗅觉受体是参与昆虫嗅觉识别过程的一类重要蛋白。在昆虫的众多嗅觉受体中, 有一类受体明显不同于其他受体, 被称为Orco。该受体基因在不同昆虫种间高度保守, 且表达广泛。Orco在昆虫嗅觉识别过程中发挥关键作用。采用基因突变或RNAi等技术使Orco基因沉默后, 昆虫会出现严重的嗅觉缺陷, 但Orco本身不与气味配体结合, 它与传统嗅觉受体形成复合体Or-Orco, 促进传统嗅觉受体在神经元树突膜上的定位并维持其稳定性, 提高传统嗅觉受体对气味反应的效率。昆虫嗅觉受体的结构与脊椎动物的G蛋白偶联受体相似, 均有7个跨膜区, 但二者的膜拓扑结构相反, 昆虫嗅觉受体的N末端位于细胞质膜内, C末端在细胞质膜外, Orco与传统嗅觉受体通过保守的C末端区域相互作用形成一种新型的配体门控离子通道--Or-Orco复合体。阐明Orco在昆虫嗅觉识别中的功能机制, 可为开创基于昆虫嗅觉行为干扰的新的害虫防治措施提供基础。     

Concentration and Membrane Fluidity Dependence of Odor Discrimination in the Turtle Olfactory System

In the present study, we examined the concentration dependenceof odor discrimination in turtle olfactory bulbar responsesusing the cross-adaptation technique. In the odorant pairs withdiverse molecular structures, the degree of discrimination wasunchanged or only slightly decreased with an increase in odorantconcentrations, suggesting that odorants are well discriminatedeven at high concentrations. In the odorant pairs with closelyrelated molecular structures, the degree of discrimination wasdecreased with an increase in odorant concentrations. An increasein the temperature of turtle olfactory epithelium also decreasedthe ability to discriminate these odorants. There was a goodcorrelation between changes in the odor discriminating abilityinduced by an increase in odor concentrations and those inducedby a temperature increase. The liposomes were made of lipidsextracted from the turtle olfactory epithelia and changes oftheir membrane fluidity induced by adsorption of odorants weremonitored with DPH. There was a good correlation between a decreasein odor discriminating ability and the membrane fluidity changesinduced by odorants. We suggest that decreases in odor discriminatingability induced either by an increase in odor concentrationor by a temperature increase are ultimately caused by changesin the membrane fluidity. Chem. Senses 22: 553–563, 1997.