Odor-mediated Behavior Research Articles

Damage to Olfactory Organs of Adult Zebrafish Induced by Diesel Particulate Matter.

Particulate matter (PM) is an environmental hazard that is associated with various human health risks. The olfactory system is directly exposed to PM; therefore, the influence of PM exposure on olfactory function must be investigated. In this study, we propose a zebrafish olfactory model to evaluate the effects of exposure to diesel particulate matter (DPM), which was labeled Korean diesel particulate matter (KDP20). KDP20 comprises heavy metals and polycyclic aromatic hydrocarbons (PAHs). KDP20 exposed olfactory organs exhibited reduced cilia and damaged epithelium. Olfactory dysfunction was confirmed using an odor-mediated behavior test. Furthermore, the olfactory damage was analyzed using Alcian blue and anti-calretinin staining. KDP20 exposed olfactory organs exhibited histological damages, such as increased goblet cells, decreased cell density, and calretinin level. Quantitative real-time polymerase chain reaction (qRT-PCR) revealed that PAHs exposure related genes (AHR2 and CYP1A) were upregulated. Reactive oxidation stress (ROS) (CAT) and inflammation (IL-1B) related genes were upregulated. Furthermore, olfactory sensory neuron (OSN) related genes (OMP and S100) were downregulated. In conclusion, KDP20 exposure induced dysfunction of the olfactory system. Additionally, the zebrafish olfactory system exhibited a regenerative capacity with recovery conditions. Thus, this model may be used in future investigating PM-related diseases.

Behavior Matters-Future Need for Insect Studies on Odor-Mediated Host Plant Recognition with the Aim of Making Use of Allelochemicals for Plant Protection.

Allelochemicals, chemical cues that, among other things, mediate insect-plant interactions, such as host plant recognition, have attracted notable interest as tools for ecological control of pest insects. Advances have recently been made in methods for sampling and analyzing volatile compounds and technology for tracking insects in their natural habitat. However, progress in odor-mediated behavioral bioassays of insects has been relatively slow. This perspective highlights this odor-mediated insect behavior, particularly in a natural setting and considering the whole behavioral sequence involved in the host location, which is the key to understanding the mechanisms underlying host plant recognition. There is thus a need to focus on elaborate behavioral bioassays in future studies, particularly if the goal is to use allelochemicals in pest control. Future directions for research are discussed.

Mosquito Host Seeking in 3D Using a Versatile Climate-Controlled Wind Tunnel System.

Future anthropogenic climate change is predicted to impact sensory-driven behaviors. Building on recent improvements in computational power and tracking technology, we have developed a versatile climate-controlled wind tunnel system, in which to study the effect of climate parameters, including temperature, precipitation, and elevated greenhouse gas levels, on odor-mediated behaviors in insects. To establish a baseline for future studies, we here analyzed the host-seeking behavior of the major malaria vector mosquito, Anopheles gambiae sensu strico, to human odor and carbon dioxide (CO2), under tightly controlled climatic conditions, and isolated from potential background contamination by the presence of an experimenter. When presented with a combination of human foot odor and CO2 (case study I), mosquitoes engaged in faster crosswind flight, spent more time in the filamentous odor plume and targeted the odor source more successfully. In contrast, female An. gambiae s. s. presented with different concentrations of CO2 alone, did not display host-seeking behavior (case study II). These observations support previous findings on the role of human host-associated cues in host seeking and confirm the role of CO2 as a synergist, but not a host-seeking cue on its own. Future studies are aimed at investigating the effect of climate change on odor-mediated behavior in mosquitoes and other insects. Moreover, the system will be used to investigate detection and processing of olfactory information in various behavioral contexts, by providing a fine-scale analysis of flight behavior.

Effects of Nicotine and Tobacco-Related Products on the Feeding Behavior of the German Cockroach (Blattodea: Blattellidae).

Animals use olfaction to detect developmentally significant volatile organic compounds (VOCs) in their local environment. As part of a wider study aiming to demonstrate that the olfactory responses of animals to VOCs can be modified through the creation of a drug-addicted status and association with a selected VOC, we investigated nicotine and tobacco smoke particulate (TSP) extract as possible addictive compounds for male German cockroaches, Blattella germanica (Linnaeus). In feeding experiments using an artificial food stimulus, food treated with TSP extract was preferred over untreated food. Surprisingly, nicotine, which was expected to be the most important addictive tobacco component, did not induce noticeable effects on cockroach behavior. Both TSP extract and nicotine were shown to be phagostimulants. Olfactometry assays that measured odor-mediated insect behavior demonstrated that male B. germanica did not choose TSP-extract-treated food even when attempts were made specifically to train them via this modality. These results support a hypothesis that B. germanica needs to consume TSP-containing food to show a clear preference for this stimulus and that gustatory mechanisms are involved due to compounds present in the TSP extract.

Dengue infection modulates locomotion and host seeking in Aedes aegypti.

Pathogens may manipulate their human and mosquito hosts to enhance disease transmission. Dengue, caused by four viral serotypes, is the fastest-growing transmissible disease globally resulting in 50–100 million infections annually. Transmission of the disease relies on the interaction between humans and the vector Aedes aegypti and is largely dependent on the odor-mediated host seeking of female mosquitoes. In this study, we use activity monitors to demonstrate that dengue virus-1 affects the locomotion and odor-mediated behavior of Ae. aegypti, reflecting the progression of infection within the mosquito. Mosquitoes 4–6 days post-infection increase locomotion, but do not alter their odor-driven host-seeking response. In contrast, females 14–16 days post-infection are less active, yet more sensitive to human odors as assessed by behavioral and electrophysiological assays. Such an increase in physiological and behavioral sensitivity is reflected by the antennal-specific increase in abundance of neural signaling transcripts in 14 days post-infection females, as determined by transcriptome analysis. This suggests that the sensitivity of the mosquito peripheral olfactory system is altered by the dengue virus by enhancing the overall neural responsiveness of the antenna, rather than the selective regulation of chemosensory-related genes. Our study reveals that dengue virus-1 enhances vector-related behaviors in the early stages post-infection that aid in avoiding predation and increasing spatial exploration. On the other hand, at the later stages of infection, the virus enhances the host-seeking capacity of the vector, thereby increasing the risk of virus transmission. A potential mechanism is discussed.

Patterns of cell death in the embryonic antenna of the grasshopper Schistocerca gregaria.

We have investigated the pattern of apoptosis in the antennal epithelium during embryonic development of the grasshopper Schistocerca gregaria. The molecular labels lachesin and annulin reveal that the antennal epithelium becomes subdivided into segment-like meristal annuli within which sensory cell clusters later differentiate. To determine whether apoptosis is involved in the development of such sensory cell clusters, we examined the expression pattern of the cell death labels acridine orange and TUNEL in the epithelium. We found stereotypic, age-dependent, wave-like patterns of cell death in the antenna. Early in embryogenesis, apoptosis is restricted to the most basal meristal annuli but subsequently spreads to encompass almost the entire antenna. Cell death then declines in more basal annuli and is only found in the tip region later in embryogenesis. Apoptosis is restricted throughout to the midregion of a given annulus and away from its border with neighboring annuli, arguing against a causal role in annular formation. Double-labeling for cell death and sensory cell differentiation reveals apoptosis occurring within bands of differentiating sensory cell clusters, matching the meristal organization of the apical antenna. Examination of the individual epithelial lineages which generate sensory cells reveals that apoptosis begins peripherally within a lineage and with age expands to encompass the differentiated sensory cell at the base. We conclude that complete lineages can undergo apoptosis and that the youngest cells in these lineages appear to die first, with the sensory neuron dying last. Lineage-based death in combination with cell death patterns in different regions of the antenna may contribute to odor-mediated behaviors in the grasshopper.

Feeding-induced changes in allatostatin-A and short neuropeptide F in the antennal lobes affect odor-mediated host seeking in the yellow fever mosquito, Aedes aegypti.

Aedes aegypti is a model species in which the endogenous regulation of odor-mediated host seeking behavior has received some attention. Sugar feeding and host seeking in female A. aegypti are transiently inhibited following a blood meal. This inhibition is partially mediated by short neuropeptide F (sNPF). The paired antennal lobes (ALs), as the first processing centers for olfactory information, has been shown to play a significant role in the neuropeptidergic regulation of odor-mediated behaviors in insects. The expression of sNPF, along with other peptides in the ALs of A. aegypti, indicate parallel neuromodulatory systems that may affect olfactory processing. To identify neuropeptides involved in regulating the odor-mediated host seeking behavior in A. aegypti, we use a semi-quantitative neuropeptidomic analysis of single ALs to analyze changes in the levels of five individual neuropeptides in response to different feeding regimes. Our results show that the level of sNPF-2, allatostatin-A-5 (AstA-5) and neuropeptide-like precursor-1-5 (NPLP-1-5), but not of tachykinin-related-peptides and SIFamide (SIFa), in the AL of female mosquitoes, changes 24 h and 48 h post-blood meal, and are dependent on prior access to sugar. To assess the role of these neuropeptides in modulating host seeking behavior, when systemically injected individually, sNPF-2 and AstA-5 significantly reduced host seeking behavior. However, only the injection of the binary mixture of the two neuropeptides lead to a host seeking inhibition similar to that observed in blood fed females. We conclude that modulation of the odor mediated host seeking behavior of A. aegypti is likely regulated by a dual neuropeptidergic pathway acting in concert in the ALs.

Odor, Not Performance, Dictates Bemisia tabaci's Selection between Healthy and Virus Infected Plants.

Although, insect herbivores are generally thought to select hosts that favor the fitness of their progeny, this “mother-knows-best” hypothesis may be challenged by the presence of a plant virus. Our previous study showed that the whitefly, Bemisia tabaci, the obligate vector for transmitting Tomato yellow leaf curl virus (TYLCV), preferred to settle and oviposit on TYLCV-infected rather than healthy host plant, Datura stramonium. The performances of B. tabaci larvae and adults were indeed improved on virus-infected D. stramonium, which is consistent with “mother-knows-best” hypothesis. In this study, B. tabaci Q displayed the same preference to settle and oviposit on Tomato spotted wilt virus (TSWV)-infected host plants, D. stramonium and Capsicum annuum, respectively. As a non-vector of TSWV, however, insect performance was impaired since adult body size, longevity, survival, and fecundity were reduced in TSWV infected D. stramonium. This appears to be an odor-mediated behavior, as plant volatile profiles are modified by viral infection. Infected plants have reduced quantities of o-xylene and α-pinene, and increased levels of phenol and 2-ethyl-1-hexanol in their headspace. Subsequent behavior experiments showed that o-xylene and α-pinene are repellant, while phenol and 2-ethyl-1-hexanol are attractive. This indicates that the preference of B. tabaci for virus-infected plants is modulated by the dynamic changes in the volatile profiles rather than the subsequent performances on virus-infected plants.

Octopamine regulates antennal sensory neurons via daytime-dependent changes in cAMP and IP3 levels in the hawkmoth Manduca sexta.

The biogenic amine octopamine (OA) mediates reward signals in olfactory learning and memory as well as circadian rhythms of sleep and activity. In the crepuscular hawkmoth Manduca sexta, OA changed pheromone detection thresholds daytime-dependently, suggesting that OA confers circadian control of olfactory transduction. Thus, with enzyme-linked immunosorbent assays we searched hawkmoth antennae for daytime-dependent changes in the concentration of OA and its respective second messengers. Antennal stimulation with OA raised cAMP- and IP3 levels. Furthermore, antennae expressed daytime-dependent changes in the concentration of OA, with maxima at Zeitgebertime (ZT) 20 when moths were active and also maximal concentrations of cAMP occurred. Maximal IP3 levels at ZT 18 and 23 correlated with maximal flight activity of male moths, while minimal IP3 levels at dusk correlated with peaks of feeding activity. Half maximal effective concentration (EC50) for activation of the OA-receptor decreased during the moth’s activity phase suggesting daytime-dependent changes in OA receptor sensitivity. With an antiserum against tyramine, the precursor of OA, two centrifugal neurons were detected projecting out into the sensory cell layer of the antenna, possibly mediating more rapid stimulus-dependent OA actions. Indeed, in fast kinetic assays OA receptor stimulation increased cAMP concentrations within 50 msec. Thus, we hypothesize that fast, stimulus-dependent centrifugal control of OA-release in the antenna occurs. Additional slow systemic OA actions might be based upon circadian release of OA into the hemolymph mediating circadian rhythms of antennal second messenger levels. The resulting rhythms of odor sensitivity are suggested to underlie circadian rhythms in odor-mediated behavior.

Calcium Imaging in the Drosophila Olfactory System with a Genetic Indicator

Insects show sophisticated odor-mediated behaviors controlled by an olfactory system that is genetically and anatomically simpler than that of vertebrates, providing an attractive system to investigate the mechanistic link between behavior and odor perception. Advances in neuroscience have been facilitated by modern optical imaging technologies--both in instrumentation and in probe design--that permit the visualization of functional neural circuits. Imaging calcium activity in genetically defined populations of neurons provides an important tool for investigating the function of neural circuits. This article describes a two-photon imaging system for monitoring neural activity in the Drosophila antennal lobe. Odor-evoked calcium activity is followed by measuring the specific expression of the calcium-sensitive green fluorescent protein G-CaMP in Drosophila antennae-brain preparations.

Mixture and odorant processing in the olfactory systems of insects: a comparative perspective

Natural olfactory stimuli are often complex mixtures of volatiles, of which the identities and ratios of constituents are important for odor-mediated behaviors. Despite this importance, the mechanism by which the olfactory system processes this complex information remains an area of active study. In this review, we describe recent progress in how odorants and mixtures are processed in the brain of insects. We use a comparative approach toward contrasting olfactory coding and the behavioral efficacy of mixtures in different insect species, and organize these topics around four sections: (1) Examples of the behavioral efficacy of odor mixtures and the olfactory environment; (2) mixture processing in the periphery; (3) mixture coding in the antennal lobe; and (4) evolutionary implications and adaptations for olfactory processing. We also include pertinent background information about the processing of individual odorants and comparative differences in wiring and anatomy, as these topics have been richly investigated and inform the processing of mixtures in the insect olfactory system. Finally, we describe exciting studies that have begun to elucidate the role of the processing of complex olfactory information in evolution and speciation.

Antennal lobe organization in the slender pigeon louse, Columbicola columbae (Phthiraptera: Ischnocera)

This study reports on the structure of the antennal lobe of the pigeon louse, Columbicola columbae. Anterograde staining of antennal receptor neurons revealed an antennal lobe with a few diffuse compartments, an organization distinct from the typical spheroidal glomerular structure found in the olfactory bulb of vertebrates and the antennal lobe of many other insects. This anatomical arrangement of neuronal input is somewhat reminiscent of the aglomerular antennal lobe previously reported in psyllids and aphids. As in psyllids, reports on the odor-mediated behavior of C. columbae suggest that the olfactory sense is important in these animals and indicates that a glomerular organization of the antennal lobe may not be necessary to subtend odor-mediated behaviors in all insects. The diffuse or aglomerular antennal lobe organization found in these two Paraneopteran insect orders might represent an independently evolved reduction due to similar ecological constraints.

Genetic Variation in Odorant Receptors Contributes to Variation in Olfactory Behavior in a Natural Population of Drosophila melanogaster

Chemoreception is a principle modality by which organisms gain information from their environment, and extensive variation in odor-mediated behavior has been documented within and among species. To examine the mechanisms by which sensory systems mediate these responses, we ask to what extent variation in Drosophila melanogaster odorant receptor genes contributes to variation in odor-mediated behavior. Significant differences in behavioral responses to structurally similar odorants, methyl hexanoate and ethyl hexanoate, were found in a natural population. Polymorphisms in 3 genomic regions (Or22a/Or22b, Or35a, and Or47a) were identified and associated with variation in behavior to these esters. Overall similarity in association profiles for both odorants was observed, except for Or47a in which polymorphisms were associated solely with variation in responses to ethyl hexanoate. Our analyses were then extended to examine polymorphisms in 3 odorant receptors previously reported to contribute to variation in olfactory behavior for the chemically distinct odorants benzaldehyde and acetophenone. Two Or10a polymorphisms were associated with variation in response to ethyl hexanoate. Finally, differences in Or35a and Or47a expression were associated with variation in responses to ethyl hexanoate. These results demonstrate that the genetic variation at the peripheral sensory stage plays a role in mediating differences in odor-mediated behavior.

Dual antennular chemosensory pathways mediate odor-associative learning and odor discrimination in the Caribbean spiny lobster Panulirus argus.

Chemosensory neurons in the antennular flagella of lobsters mediate long-range responses to chemicals. These neurons are part of two parallel chemosensory pathways with different peripheral and central components. Aesthetasc sensilla on the lateral flagella are innervated by chemosensory neurons that project to the olfactory lobes. A diversity of other 'non-aesthetasc' sensilla on both lateral and medial flagella are innervated by mechano- and chemosensory neurons, and most of these non-aesthetasc neurons project to the lateral antennular neuropils. We investigated the roles of these two pathways in odor-associative learning and odor discrimination by selectively removing either aesthetasc or non-aesthetasc sensilla from the spiny lobster Panulirus argus. Lobsters lacking both aesthetasc and non-aesthetasc antennular sensilla show very reduced or no odor-mediated searching behavior. We associatively conditioned lobsters using two paradigms: aversive conditioning with generalization testing (which reveals the similarity in the lobsters' perception of odorants) and discrimination conditioning (which reveals the lobsters' ability to discriminate odorants). Sham-control intact lobsters performed these tasks well, as did lobsters lacking either aesthetascs or non-aesthetasc setae. There was a strong but statistically non-significant trend that lobsters lacking either aesthetascs or non-aesthetasc setae generalized more between complex odor mixtures than did intact lobsters. After aversive conditioning with generalization testing, aesthetasc-ablated lobsters had more difficulty discriminating among the most closely related complex mixtures than did intact or non-aesthetasc-ablated lobsters. However, after discrimination conditioning, aesthetasc-ablated lobsters were as proficient as intact animals in discriminating highly similar mixtures. These results indicate overlap and redundancy in the function of these two chemosensory pathways in odor-associative learning and odor discrimination, but these pathways also complement each other to enable better discrimination. This study presents the first evidence for a role of non-aesthetasc chemosensory neurons in complex odor-mediated behaviors such as learning and discrimination.

Ultrastructure and physiology of the hooded sensillum, a bimodal chemo-mechanosensillum of lobsters.

The antennules of decapod crustaceans are covered with thousands of chemosensilla that mediate odor discrimination and orientation behaviors. Most studies on chemoreception in decapods have focused on the prominent aesthetasc sensilla. However, previous behavioral studies on lobsters following selective sensillar ablation have revealed that input from nonaesthetasc antennular chemosensilla is sufficient for many odor-mediated behaviors. Our earlier examination of the setal types on the antennules of the Caribbean spiny lobster Panulirus argus revealed three types of nonaesthetasc chemosensilla. The most abundant and widely distributed of these is the hooded sensillum. The present study describes the detailed ultrastructure of antennular hooded sensilla and the physiological response properties of their receptor neurons. Light and scanning and transmission electron microscopy were used to examine structural characteristics, and electrophysiology was used to examine single-unit responses elicited by focal chemical and mechanical stimulation of antennular hooded sensilla. Hooded sensilla have a porous cuticle and are innervated by 9-10 chemosensory and 3 mechanosensory neurons whose dendrites project to the distal end of the sensillum. Hooded sensillar chemosensory neurons responded to waterborne chemicals, were responsive to only one of the six tested single compounds, and had different specificities. Hooded sensillar mechanosensory neurons were not spontaneously active. They had low sensitivity in that they responded to tactile but not waterborne vibrations, and they responded to sensillar deflection with phasic bursts of activity. These results support the idea that hooded sensilla are bimodal chemo-mechanosensilla and are receptors in an antennular chemosensory pathway that parallels the well-described aesthetasc chemosensory pathway.

Odor-mediated behavior of Afrotropical malaria mosquitoes.

The African mosquito species Anopheles gambiae sensu lato s.l. and Anopheles funestus rank among the world's most efficient vectors of human malaria. Their unique bionomics, particularly their anthropophilic, endophagic and endophilic characters, guarantee a strong mosquito-host interaction, favorable to malaria transmission. Olfactory cues govern the various behaviors of female mosquitoes and here we review the role of semiochemicals in the life history of African malaria vectors. Recent evidence points towards the existence of human-specific kairomones affecting host-seeking A. gambiae s.l., and efforts are under way to identify the volatiles mediating this behavior. Based on examples from other Culicidae spp., it is argued that there is good reason to assume that mating, sugar feeding, and oviposition behavior in Afrotropical malaria vectors may also be mediated by semiochemicals. It is foreseen that increased knowledge of odor-mediated behaviors will be applied in the development of novel sampling techniques and possibly alternative methods of intervention to control malaria.

Shelter sharing and chemical courtship signals in the lobster, Homarus americanus

In a 3.7-m Y-maze flume study of social odor-mediated behavior of mature lobsters (Homarus americanus), females preferred male- but not female-occupied shelters over empty shelters: they detected them from a distance and spent much time trying to enter. Males did not show distant detection and preference for female shelters but did spend much time trying to enter female but not male shelters once nearby. Sheltered resident males showed strong aggression toward visiting males but only mild aggression to visiting females; intermolt and premolt females could enter resident male shelters, cohabit for at least 7 h, receive mating attempts, and sometimes mate. Premolt females or females with sperm plugs resisted mating attempts. Visiting females released more than four times and males more than seven times as much urine during a shelter approach compared with an equal time in isolation. Females with or without urine release entered male-occupied shelters successfully, but mating attempts rarely occurred when female urine release was blocked. Female urine block resulted in greatly increased resident male aggression toward her, reaching the same levels elicited by visiting males with or without urine block. The results show that female urine signals reduce male aggression and facilitate mating.

The Cutting Edge of Insect Olfaction

Odor discrimination by insects can be exploited by entomologists to make species-specific pheromone traps and mating disruptants or to manipulate insects via host odors. Recent neuroanatomical studies have revealed how odor blends are represented by antennal neurons and their associated target glomeruli in the first layers of the brain as the relative abundance of each individual component in the blend. Further processing by neurons deeper in the brain decodes these lines of component-specific information and integrates them to produce enhanced signals that represent the blend quality. This article describes techniques that have aided in tracing and mapping olfactory pathways, thereby improving our ability to design new strategies for shortcircuiting odor-mediated insect behavior.

Normal glomerular organization of the antennal lobes is not necessary for odor-modulated flight in female moths

A prominent hypothesis for the function of the glomerular structures in the primary olfactory neuropil of many groups of vertebrate and invertebrate animals is that they enable the processing and coding of information about the chemical compounds that compose complex odors. Previous studies have indicated that various degrees of glomerulus formation in the antennal lobes of the brain of the moth Manduca sexta can be effected by reducing the number of olfactory sensory axons that grow from the antenna into the antennal lobe during metamorphosis. To test the hypothesis that the presence of glomerular structure is necessary to process and identify odors, we substantially reduced, by surgery, the number of antennal segments in developing moths and upon metamorphosis we observed and quantified behavioral responses known to be elicited by odors. Intact and lesioned adult female moths were challenged to fly upwind to the source of an attractive host-plant odor in a wind tunnel. Some of the moths that had developed with reduced olfactory input flew upwind to the odor source. The flight behavior of these individuals was similar to the odor-mediated flight typically observed in moths that had developed normally. Histological analysis of the moths' antennal lobes revealed that the lobes of more than half of the respondents that had been lesioned during development lacked normal glomerular organization. The neuropil of these abnormally developed antennal lobes was mostly aglomerular, but with a few isolated, clearly abnormal glomerulus-like structures. This suggests either that even a few abnormal glomeruli are sufficient to mediate this specific behavior or that "canonical" glomerular organization per se is not necessary for this odor-mediated behavior.

Body Odors and Neutral-basic Peptide Mimics: A Review of Responses by Marine Organisms

A review of body odor-mediated behaviors of hermit crabs, predatory gastropods, barnacle larvae and brachyuran crabs includes a model based on the hypothesis that all the behaviors are mediated by a similar chemical detection mechanism. A key assumption is behaviors evolved due to a selective advantage conferred by detection of information in body odors. In the model, exogenous trypsin-like proteases digest structural proteins to generate neutral-basic peptides. The receptor is postulated to have resulted from a point mutation in a trypsin-like membrane protein. Transduction of the binding event is via second messenger mediated pathways. Examples of body odors functioning as cues, allomones and pheromones are presented. The model is supported by a review of structure-function studies employing synthetic neutral basic peptides and studies with other macromolecules.