昆虫嗅觉系统结构与功能研究进展

昆虫的脑由前脑、中脑和后脑组成,其中前脑含有高级感觉中枢,如蘑菇体和中央复合体,控制昆虫的学习、记忆和运动等高级神经活动;中脑包含触角叶,是嗅觉神经中心;而后脑则通常不发达,主要包括内分泌神经元和控制进食与消化的运动神经元。不同于其他物种,昆虫由于其特殊的生活习性,听觉和视觉系统相对退化,主要依赖嗅觉来捕食、交流和求偶,因此嗅觉系统尤其发达。本文综述了目前对昆虫的脑部主要神经结构和功能(中央复合体、蕈形体和触角叶结构)以及昆虫脑部结构遗传变异(性别异构,不同发育时期、不同昆虫以及昆虫与其他动物的脑部结构差异)的研究进展,并总结了目前昆虫脑对信号的加工处理和识别机制的研究结果。     

Anatomical and physiological characteristics of individual neurones in the central antennal pathway of insects

Signals of tens up to hundreds of thousands of (mostly olfactory) receptor cells on an insect antenna are switched to a comparatively low number of neurones in the antennal lobe of the deutocerebrum in circumscribed units of neuropile, the glomeruli. Each glomerulus is connected via its output neurone to two separate neuropiles (calyces of mushroom body, and lateral lobe) of the protocerebrum. Local interneurones interconnect between the glomeruli. Certain modes of convergence between receptors and central neurones provide for a very high sensitivity of the latter to certain odours and their sensitivity for complex odour stimuli, and in many cases for a marked multimodality. Anatomical and physiological data are given especially for pheromone sensitive neurones and their projections.     

Physiological mismatching between neurons innervating olfactory glomeruli in a moth

The primary olfactory centres of most vertebrates and most neopteran insects are characterized by the presence of spherical neuropils, glomeruli, where synaptic interactions between olfactory receptor neurons and second-order neurons take place. In the neopteran insect taxa investigated so far, receptor neurons of a specific physiological identity target one glomerulus and thus bestow a functional identity on the glomerulus. In moths, input from pheromone-specific receptor neurons is received in a male-specific structure of the antennal lobe, called the macroglomerular complex (MGC), which consists of a number of specialized glomeruli. Each glomerulus of the complex receives a set of peripheral sensory afferents that encode one of several compounds involved in sexual communication. The complex is also innervated by dendritic branches of antennal lobe output neurons called projection neurons, which transfer information from the antennal lobe to higher centres of the brain. A hypothesis stemming from earlier work on moths claims that the receptor neuron innervation pattern of the MGC should be reflected in the pattern of dendrites of projection neurons invading the different MGC glomeruli. In this study we show that in the noctuid moth Trichoplusia ni, as in several other noctuid moth species, this hypothesis does not hold. The degree of matching between axon terminals of receptor neurons and the dendritic branches of identified projection neurons that express similar physiological specificity is very low.     

Olfactory perception and learning in the honey bee (Apis mellifera): calcium imaging in the antenna lobe

Honey bees are a key-model in the study of learning and memory, because they show considerable learning abilities, their brain is well described and is accessible to a wide range of physiological recordings and treatments. We use in vivo calcium imaging to study olfactory perception in the bee brain, and combine this method to appetitive olfactory conditioning to unravel the neural substrates of olfactory learning. Odours are detected by receptor neurons on the antennae. Each receptor neuron projects to the first-order neuropile of the olfactory pathway, the antennal lobe, connecting to projection neurons in one of its 160 functional units, the glomeruli. In calcium imaging experiments, each odour elicits a particular activity pattern of antennal lobe glomeruli, according to a code conserved between individuals. The antennal lobe is also a site where the olfactory memory is formed. Using optical imaging, two studies have shown modulations of odour representation in the antennal lobe after learning, with different effects depending on the type of conditioning used. While simple differential conditioning (A + B- training) showed an increased calcium response to the reinforced odour, side-specific conditioning (A + B-/B + A- training) decorrelated the calcium responses of odours between brain sides. This difference may owe to the formation of different memories, which will be addressed in future work. By specifically staining antennal lobe neuronal subpopulations, we hope to be able in the future to study synaptic plasticity in the honey bee.     

Odour perception in honeybees: coding information in glomerular patterns

Major advances have been made during the past two years in understanding how honeybees process olfactory input at the level of their first brain structure dealing with odours, the antennal lobe (the insect analogue of the mammalian olfactory bulb). It is now possible to map physiological responses to morphologically identified olfactory glomeruli, allowing for the creation of a functional atlas of the antennal lobe. Furthermore, the measurement of odour-evoked activity patterns has now been combined with studies of appetitive odour learning. The results show that both genetically determined components and learning-related plasticity shape olfactory processing in the antennal lobe.     

昆虫气味认知的嗅觉神经结构及分子机制

大多数昆虫主要通过气味认知感知外界环境的变化,维持生命活动。探究昆虫气味认知的嗅觉系统神经结构及分子机制,对于完善气味认知神经生物学理论及利用其原理进行仿生学研究等有重要的科学意义。近年,关于昆虫气味认知科学研究有了很大的进展。本文从昆虫神经生物学的视角详细综述了近年关于昆虫气味认知的嗅觉神经结构、分子机制及气味信号的神经传导途径等方面的基本理论及最新研究成果。综述结果显示:昆虫对气味的认知是通过嗅觉神经系统的触角感器、触角叶(AL)、蕈形体(MB)等脑内多层信号处理神经结构来实现的。当外界气味分子进入触角感器内后,由感器内特定的气味识别蛋白(OBP)将气味分子运载到达嗅觉感受神经元(ORN)树突膜上的受体位点,气味分子与表达特定气味的受体(OR)结合产生电信号,并以动作电位的形式通过ORN的轴突传到脑内的触角叶。在触角叶经过嗅觉纤维球对气味信息选择性加工处理,再由投射神经元(PNs)将初步的识别和分类的气味信息传到蕈形体和外侧角(LH)等神经中枢,实现对气味的识别和认知。虽然,近年昆虫气味认知神经生物学的研究有了很大的进步,但是,我们认为目前的研究成果还不能完全阐明昆虫气味认知的神经机制,还有很多问题,例如,触角叶上众多的嗅觉纤维球是如何对嗅觉感受神经元传入的气味信息进行编码处理的?等有待进一步深入研究。为了搞清这些疑难问题,我们认为需要提高现有的实验技术水平,加强电生理学和分子神经生物学相结合的实验研究,从分子水平探究气味认知的神经机制可能是未来研究的热点。     

昆虫触角叶的结构

触角叶是昆虫脑内初级嗅觉中心,通过触角神经与触角联系。触角叶主要由嗅觉受体神经元、局域中间神经元、投射神经元和远心神经元构成。这些神经元的形态多样,其形态变化与其功能和昆虫嗅觉行为相关。这些神经元在触角叶内交织形成神经纤维网,在突触联系紧密的地方形成纤维球,纤维球通常排列在触角叶外周。通常,昆虫触角叶内纤维球的数量、大小和位置相对固定,并且几乎每个小球都可以被识别和命名。不同种类、性别和品级的昆虫中,纤维球的数量、大小和排列方式各不相同。触角叶结构神经元组成和纤维球的多样性,与各种昆虫嗅觉行为的特异性相对应。     

Chemosensory processing in the fruit fly,Drosophila melanogaster: generalization of a feeding response reveals overlapping odour representations

Insects are capable of detecting, and discriminating between, a very large number of odours. The biological relevance of many of those odours, particularly those related to food, must first be learned. Given that the number of sensory receptors and antennal lobe (AL) glomeruli is limited relative to the number of odours that must be detectable, this ability implies that the olfactory system makes use of a combinatorial coding scheme whereby each sensory cell or AL projection neuron can participate in coding for several different odours. An important step in understanding this coding scheme is to behaviourally quantify the degree to which sets of odours are discriminable. Here we evaluate odour discriminability in the fruit fly,Drosophila melanogaster, by first conditioning individual flies to not respond to any of several odourants using a nonassociative conditioning protocol (habituation). We show that flies habituate unconditioned leg movement responses to both mechanosensory and olfactory stimulation over 25 unreinforceed trials. Habituation is retained for at least 2 h and is subject to dishabituation. Finally, we test the degree to which the conditioned response generalizes to other odourants based on molecular features of the odourants (e.g. carbon chain length and the presence of a target functional group). These tests reveal predictable generalization gradients across these molecular features. These data substantiate the claim that these features are relevant coding dimensions in the fruit fly olfactory system, as has been shown for other insect and vertebrate species.     

Glomerular interactions in olfactory processing channels of the antennal lobes

An open question in olfactory coding is the extent of interglomerular connectivity: do olfactory glomeruli and their neurons regulate the odorant responses of neurons innervating other glomeruli? In the olfactory system of the moth Manduca sexta, the response properties of different types of antennal olfactory receptor cells are known. Likewise, a subset of antennal lobe glomeruli has been functionally characterized and the olfactory tuning of their innervating neurons identified. This provides a unique opportunity to determine functional interactions between glomeruli of known input, specifically, (1) glomeruli processing plant odors and (2) glomeruli activated by antennal stimulation with pheromone components of conspecific females. Several studies describe reciprocal inhibitory effects between different types of pheromone-responsive projection neurons suggesting lateral inhibitory interactions between pheromone component-selective glomerular neural circuits. Furthermore, antennal lobe projection neurons that respond to host plant volatiles and innervate single, ordinary glomeruli are inhibited during antennal stimulation with the female’s sex pheromone. The studies demonstrate the existence of lateral inhibitory effects in response to behaviorally significant odorant stimuli and irrespective of glomerular location in the antennal lobe. Inhibitory interactions are present within and between olfactory subsystems (pheromonal and non-pheromonal subsystems), potentially to enhance contrast and strengthen odorant discrimination.     

Projection pattern of poreplate sensory neurones in honey bee worker,Apis mellifera L. (Hymenoptera : Apidae)

The projections of olfactory receptor cells of the poreplate sensilla were studied in the worker honey bee, Apis mellifera L. (Hymenoptera : Apidae) by filling single sensilla iontophoretically with cobalt chloride. Successful fillings of individual sensilla lead to staining of one to 22 sensory neurones. All stained receptor cell axons are uniglomerular. Seven fillings of poreplates from the 5th flagellar segment in different animals were compared to analyse the distribution patterns of the receptor cell axons in the antennal lobe. The sensory neurones of individual poreplates project to widely distributed glomeruli in the antennal lobe. The projection patterns of different poreplates are not the same, but may be overlapping.     

A candidate olfactory receptor subtype highly conserved across different insect orders

Candidate olfactory receptors of the moth Heliothis virescens were found to be extremely diverse from receptors of the fruitfly Drosophila melanogaster and the mosquito Anopheles gambiae, but there is one exception. The moth receptor type HR2 shares a rather high degree of sequence identity with one olfactory receptor type both from Drosophila (Dor83b) and from Anopheles (AgamGPRor7); moreover, in contrast to all other receptors, this unique receptor type is expressed in numerous antennal neurons. Here we describe the identification of HR2 homologues in two further lepidopteran species, the moths Antheraea pernyi and Bombyx mori, which share 86-88% of their amino acids. In addition, based on RT-PCR experiments HR2 homologues were discovered in antennal cDNA of the honey bee (Apis mellifera; Hymenoptera), the blowfly (Calliphora erythrocephala; Diptera) and the mealworm (Tenebrio molitor; Coleoptera). Comparison of all HR2-related receptors revealed a high degree of sequence conservation across insect orders. In situ hybridization of antennal sections from the bee and the blowfly support the notion that HR2-related receptors are generally expressed in a very large number of antennal cells. This, together with the high degree of conservation suggests that this unique receptor subtype may fulfill a special function in chemosensory neurons of insects.     

Concurrent modulation of neuronal and behavioural olfactory responses to sex and host plant cues in a male moth

Mating has profound effects on animal physiology and behaviour, not only in females but also in males, which we show here for olfactory responses. In cotton leafworm moths, Spodoptera littoralis, odour-mediated attraction to sex pheromone and plant volatiles are modulated after mating, producing a behavioural response that matches the physiological condition of the male insect. Unmated males are attracted by upwind flight to sex pheromone released by calling females, as well as to volatiles of lilac flowers and green leaves of the host plant cotton, signalling adult food and mating sites, respectively. Mating temporarily abolishes male attraction to females and host plant odour, but does not diminish attraction to flowers. This behavioural modulation is correlated with a response modulation in the olfactory system, as shown by electro-physiological recordings from antennae and by functional imaging of the antennal lobe, using natural odours and synthetic compounds. An effect of mating on the olfactory responses to pheromone and cotton plant volatiles but not to lilac flowers indicates the presence of functionally independent neural circuits within the olfactory system. Our results indicate that these circuits interconnect and weigh perception of social and habitat odour signals to generate appropriate behavioural responses according to mating state.     

Cloning and expression pattern of a putative octopamine/tyramine receptor in antennae of the noctuid moth <Emphasis Type="Italic">Mamestra brassicae</Emphasis>

In insects, biogenic amines have been shown to play an important role in olfactory plasticity. In a first attempt to decipher the underlying molecular mechanisms, we report the molecular cloning and precise expression pattern of a newly identified octopamine/tyramine-receptor-encoding gene in the antennae of the noctuid moth Mamestra brassicae (MbraOAR/TAR). A full-length cDNA has been obtained through homology cloning in combination with rapid amplification of cDNA ends/polymerase chain reaction; the deduced protein exhibits high identities with previously identified octopamine/tyramine receptors in other moths. In situ hybridization within the antennae has revealed that MbraOAR/TAR is expressed at the bases of both pheromone-sensitive and non-sensitive olfactory sensilla and in cells with a neurone-like shape. In accordance with previous physiological studies that have revealed a role of biogenic amines in the electrical activity of the receptor neurones, our results suggest that biogenic amines (either octopamine or tyramine) target olfactory receptor neurones to modulate olfactory coding as early as the antennal level.     

Calcium responses to pheromones and plant odours in the antennal lobe of the male and female moth Heliothis virescens

In male moths, the primary olfactory integration centre, the antennal lobe, consists of two systems. The macroglomerular complex processes pheromone information, while the ordinary glomeruli process plant odour information. Females lack a macroglomerular complex. We measured the spatial representation of odours using in-vivo optical recording. We found that: (1) pheromone substances elicited activity exclusively in the MGC. No response was found in female antennal lobes. (2) Plant odours elicited combinatorial activity patterns in the ordinary glomeruli in both males and females. No response was found in the MGC of male moths. (3) A clean air puff often led to activity, in both males and females, suggesting that mechano-sensory information is also processed in the antennal lobe. (4) With an interstimulus interval of 5 or 10 s, strongly activated glomeruli were able to follow the temporal structure of the stimulus, while others lost their phase-locking. Some glomeruli showed "off" responses. These properties were odour dependent. This confirms and extends previous studies, showing the functional significance of the two subsystems for processing olfactory information. Pheromones are coded in a combinatorial manner within the macroglomerular complex, with each glomerulus corresponding to one information channel. Plant odours are coded in an across-glomeruli code in the ordinary glomeruli.     

Contribution of single unit analysis in insects to an understanding of olfactory function

Summary The olfactory pathway of the American cockroach is described from antennal receptor input to protocerebral output neurons. Based upon new and reviewed electrophysiological, neuroanatomical and fine structural data, connections and projections of individually identified receptor cells and central neurons are demonstrated. On the basis of investigation of input-output functions of such units modes of neural coding of naturally occurring complex odor stimuli and their constituents are proposed for the first two stages of the pathway. The role of protocerebral neurons is discussed at a functional-neuroanatomical level. A few data are added of descending neurons of moths.Abbreviations CP Copora pedunculata - MG Macroglomerulus - TOG Tractus olfactorio-globularis - LLP lateral lobe of protocerebrum     

利用单感器记录与神经元示踪结合对棉铃虫主要性信息素感器内神经元投射的鉴定

【目的】鉴定雄性棉铃虫Helicoverpa armigera成虫触角性信息素感器嗅觉受体神经元的功能、形态及中枢投射路径。【方法】利用单感器记录技术记录棉铃虫嗅觉受体神经元对性信息素的反应,同时采用荧光染料作为示踪剂染色标记嗅觉受体神经元;使用免疫组织化学方法处理相应的脑组织,标记脑内触角叶的神经纤维球结构;用激光扫描共聚焦显微镜获取图像数据,使用图形软件ZEN和Amira 4.1.1进行三维结构重建。【结果】记录到雄性棉铃虫成虫触角上长毛形感器对主要性信息素成分Z11-16∶Ald产生明显的电生理反应,并成功染色标记了该感器内的嗅觉受体神经元。染色标记显示该感器内具有两个嗅觉受体神经元,其轴突通过触角神经分别投射触角叶内的云状体神经纤维球和普通神经纤维球。【结论】单感器记录与神经元示踪两技术结合能够用于鉴定昆虫触角嗅觉受体神经元的功能、形态和投射至神经纤维球的路径。与赖氨酸钴方法比较,使用荧光染料法进行神经元示踪,操作更简便,且易于进行三维空间分析,为调查棉铃虫其他嗅觉神经元的投射路径以明确外周气味受体感受与中枢系统的联系提供了有力技术支持。     

Morphology and physiology of the serotonin-immunoreactive putative antennal lobe feedback neuron in the male silkmoth Bombyx mori

In the male silkmoth Bombyx mori, olfactory information is relayed from olfactory receptor neurons in the antennae to the antennal lobe, and then to a variety of protocerebral neuropils. Currently, very little is known about neuromodulators that may affect the dynamics of this olfactory neural network. Immunocytochemical studies have revealed the presence of a serotonin-immunoreactive (SI) neuron that, in several insect species, is thought to provide feedback to the antennal lobe. To date, no studies have revealed details of this neuron's physiology. Using intracellular recording and staining, the silkmoth SI neuron (in two individuals) was first characterized physiologically and then stained with Lucifer Yellow to reveal morphological details. Immunocytochemical methods were also used to confirm the presence of serotonin. The silkmoth SI neuron branched in many important brain neuropils such as the mushroom body, central body, lateral accessory lobe and antennal lobe. The SI neuron in both individuals fired spontaneous, long duration action potentials, and responded to mechanosensory stimuli to the antennae.     

The organization of the antennal lobe correlates not only with phylogenetic relationship, but also life history: a Basal hymenopteran as exemplar

The structure of the brain is a consequence of selective pressures and the ancestral brain structures modified by those pressures. The Hymenoptera are one of the most behaviorally complex insect orders, and the olfactory system of honeybees (one of the most derived members) has been extensively studied. To understand the context in which the olfactory system of the Hymenoptera evolved, we performed a variety of immunocytochemical and anatomical labeling techniques on the antennal lobes (ALs) of one of its most primitive members, the sawflies, to provide a comparison between the honeybee and other insect model species. The olfactory receptor neurons project from the antennae to fill the entire glomerular volume but do not form distinct tracts as in the honeybee. Labeling of projection neurons revealed 5 output tracts similar to those in moths and immunolabeling for several transmitters revealed distinct populations of local interneurons and centrifugal neurons that were also similar to moths. There were, however, no histaminergic or dopaminergic AL neurons. The similarities between sawflies and moths suggest that along with the great radiation and increased complexity of behavioral repertoire of the Hymenoptera, there were extensive modifications of AL structure.     

Responses of olfactory receptor neurones to behaviourally important odours in gregarious and solitarious desert locust, Schistocerca gregaria

Abstract.Recordings from antennal olfactory receptor neurones in young adult Schistocerca gregaria Forskål (Orthoptera: Acrididae) showed that behaviourally important odours are detected by receptor neurones present in morphologically identifiable sensillum types. Both nymph- and adult-produced aggregation pheromones activate receptor neurones housed in sensilla basiconica. The receptor neurones in this sensillum type in solitary-reared locusts display a higher sensitivity to aggregation pheromones and to some other behaviourally relevant odours than the same type of neurones in gregarious locusts. Receptor neurones present in sensilla coeloconica respond to green leaf odours, organic acids, and nymphal odours but are inhibited by mature adult-produced aggregation pheromones. Receptor neurones housed in sensilla trichodea respond to a possible sex pheromone. No phase differences were found in the response of coeloconic- or trichoid-associated receptor neurones.