On the role of olfaction in sexual and interspecies recognition in crickets (Acheta and Gryllus)

Crickets appear to rely less upon olfactory communication than do their non-acoustical relatives, but males and females of house (Acheta) and field (Gryllus) crickets can determine sex by odour, and males can distinguish odours of conspecific and heterospecific females. In the presence of female odours, female-deprived males groom more vigorously and are more aggressive.     

Roles of NO Signaling in Long-Term Memory Formation in Visual Learning in an Insect

Many insects exhibit excellent capability of visual learning, but the molecular and neural mechanisms are poorly understood. This is in contrast to accumulation of information on molecular and neural mechanisms of olfactory learning in insects. In olfactory learning in insects, it has been shown that cyclic AMP (cAMP) signaling critically participates in the formation of protein synthesis-dependent long-term memory (LTM) and, in some insects, nitric oxide (NO)-cyclic GMP (cGMP) signaling also plays roles in LTM formation. In this study, we examined the possible contribution of NO-cGMP signaling and cAMP signaling to LTM formation in visual pattern learning in crickets. Crickets that had been subjected to 8-trial conditioning to associate a visual pattern with water reward exhibited memory retention 1 day after conditioning, whereas those subjected to 4-trial conditioning exhibited 30-min memory retention but not 1-day retention. Injection of cycloheximide, a protein synthesis inhibitor, into the hemolymph prior to 8-trial conditioning blocked formation of 1-day memory, whereas it had no effect on 30-min memory formation, indicating that 1-day memory can be characterized as protein synthesis-dependent long-term memory (LTM). Injection of an inhibitor of the enzyme producing an NO or cAMP prior to 8-trial visual conditioning blocked LTM formation, whereas it had no effect on 30-min memory formation. Moreover, injection of an NO donor, cGMP analogue or cAMP analogue prior to 4-trial conditioning induced LTM. Induction of LTM by an NO donor was blocked by DDA, an inhibitor of adenylyl cyclase, an enzyme producing cAMP, but LTM induction by a cAMP analogue was not impaired by L-NAME, an inhibitor of NO synthase. The results indicate that cAMP signaling is downstream of NO signaling for visual LTM formation. We conclude that visual learning and olfactory learning share common biochemical cascades for LTM formation.     

Time course of protein synthesis-dependent phase of olfactory memory in the cricket Gryllus bimaculatus

The cricket Gryllus bimaculatus forms a stable olfactory memory that lasts for practically a lifetime. As a first step to elucidate the cellular mechanisms of olfactory learning and memory retention in crickets, we studied the dependency of memory retention on the de novo brain protein synthesis by injecting the protein synthesis inhibitor cycloheximide (CHX) into the head capsule. Injection of CHX inhibited (3)H-leucine incorporation into brain proteins by > 90% for 3 hr. Crickets were trained to associate peppermint odor with water (reward) and vanilla odor with saline solution (non-reward) and were injected with CHX before or at different times after training. Their odor preferences were tested at 2 hr, 1 day and 4 days after training. Memory retention at 2 hr after training was unaffected by CHX injection. However, the level of retention at 1 day and 4 days after training was lowered when CHX was injected 1 hour before training or at 1 hr or 6 hr after training. To study the time course of the development of CHX-sensitive memory phase, crickets that had been injected with CHX at 1 hr after training were tested at different times from 2 to 12 hr after training. The level of retention was unaffected up to 4 hr after training but significantly lowered at 5 hr after training, and the CHX-sensitive memory phase developed gradually during the next several hours. CHX dissociates two phases of olfactory memory in crickets: earlier protein synthesis-independent phase (< 4 hr) and later (> 5 hr) protein synthesis-dependent phase.     

Neonatal representation of odour objects: distinct memories of the whole and its parts

Extraction of relevant information from highly complex environments is a prerequisite to survival. Within odour mixtures, such information is contained in the odours of specific elements or in the mixture configuration perceived as a whole unique odour. For instance, an AB mixture of the element A (ethyl isobutyrate) and the element B (ethyl maltol) generates a configural AB percept in humans and apparently in another species, the rabbit. Here, we examined whether the memory of such a configuration is distinct from the memory of the individual odorants. Taking advantage of the newborn rabbit''s ability to learn odour mixtures, we combined behavioural and pharmacological tools to specifically eliminate elemental memory of A and B after conditioning to the AB mixture and evaluate consequences on configural memory of AB. The amnesic treatment suppressed responsiveness to A and B but not to AB. Two other experiments confirmed the specific perception and particular memory of the AB mixture. These data demonstrate the existence of configurations in certain odour mixtures and their representation as unique objects: after learning, animals form a configural memory of these mixtures, which coexists with, but is relatively dissociated from, memory of their elements. This capability emerges very early in life.     

Perceptual and neural olfactory similarity in honeybees

The question of whether or not neural activity patterns recorded in the olfactory centres of the brain correspond to olfactory perceptual measures remains unanswered. To address this question, we studied olfaction in honeybees Apis mellifera using the olfactory conditioning of the proboscis extension response. We conditioned bees to odours and tested generalisation responses to different odours. Sixteen odours were used, which varied both in their functional group (primary and secondary alcohols, aldehydes and ketones) and in their carbon-chain length (from six to nine carbons).The results obtained by presentation of a total of 16 × 16 odour pairs show that (i) all odorants presented could be learned, although acquisition was lower for short-chain ketones; (ii) generalisation varied depending both on the functional group and the carbon-chain length of odours trained; higher generalisation was found between long-chain than between short-chain molecules and between groups such as primary and secondary alcohols; (iii) for some odour pairs, cross-generalisation between odorants was asymmetric; (iv) a putative olfactory space could be defined for the honeybee with functional group and carbon-chain length as inner dimensions; (v) perceptual distances in such a space correlate well with physiological distances determined from optophysiological recordings of antennal lobe activity. We conclude that functional group and carbon-chain length are inner dimensions of the honeybee olfactory space and that neural activity in the antennal lobe reflects the perceptual quality of odours.     

Olfactory conditioning of the proboscis extension in bumble bees

The foraging behaviour of bumble bees is well documented for nectar and/or pollen gathering, but little is known about the learning processes underlying such behaviour. We report olfactory conditioning in worker bumble bees Bombus terrestris L. (Hymenoptera: Apidae) obtained under laboratory conditions on restrained individuals. The protocol was adapted from the proboscis extension conditioning previously described in the honey bee Apis mellifera L. Bumble bees were found to be able to learn a pure odorant when it was presented in paired association with a sugar reward, but not when odour and reward were presented in an explicitly unpaired procedure. This suggests an associative basis for this olfactory learning. Bumble bees showed similar conditioning abilities when stimulated with two different floral odours. An effect of the sugar reward concentration on the learning performances was found.     

棉铃虫成虫对两种关键花香气味的联系性学习

为了明确棉铃虫对苯乙醛和乙酸苯甲酯两种关键花香气味的联系性学习行为,在室内分别测定了不同学习训练的棉铃虫雌雄成虫对两种关键花香气味的选择偏好性反应和雄成虫的触角电位反应。偏好性测定结果表明,无花香气味接触经历的棉铃虫对苯乙醛和乙酸苯甲酯的选择频次均无显著差异,且单纯的花香气味接触经历也不能诱导棉铃虫气味偏好性的显著改变,唯有花香气味接触经历伴随蔗糖溶液进行强化训练时,才能诱导气味偏好性的显著性改变。从对两种花香气味的联系性学习看,棉铃虫更容易将苯乙醛与蔗糖溶液进行联系性学习,而不容易将乙酸苯甲酯与蔗糖溶液进行联系性学习。从性别差异看,雌虫比雄虫具有更强的将花香气味与蔗糖溶液进行联系性学习的能力。触角电位测定结果表明,不同学习训练的棉铃虫雄成虫对两种花香气味和绿叶气味顺-3-己烯-1-醇的EAG反应值之间不存在显著性差异。总之,花香气味伴随食物资源的联系性学习经历,能够明显提高棉铃虫成虫对花香气味的选择偏好,可以此为基础进一步优化花香引诱剂的配方设计和应用技术。     

Demystifying wine expertise: olfactory threshold,perceptual skill and semantic memory in expert and novice wine judges

We investigated recognition and identification of wine-relevant odours as a function of domain-specific expertise. Eleven wine experts and 11 wine novices participated in tasks measuring olfactory threshold, odour recognition, odour identification, and consistency of odour naming. Twenty-four wine-relevant odorants were sampled orthonasally by each participant in the semantic (identification; consistency of naming) and episodic (recognition) memory tasks. Results showed superior olfactory recognition by expert wine judges, despite their olfactory sensitivity and bias measures being similar to those of novices. Contrary to predictions based on reports of an association between odour memory and semantic processing, wine experts did not perform better than novices on the verbal memory tasks. Further, ability to recognize odours and ability to name odours were not positively correlated, although the novices' data showed a trend in this direction. The results imply that the source of superior odour recognition in wine experts was not enhanced semantic memory and linguistic capabilities for wine-relevant odours. One interpretation of the data is that wine experts were less susceptible than wine novices to verbal overshadowing. When forced to identify the odorants, experts' superior perceptual skills protected them from verbal interference, whereas novices' generated verbal representations of the odours were emphasized at the expense of the odorant itself. This has implications for training in wine-evaluation skills.     

Complete nutrient content of four species of commercially available feeder insects fed enhanced diets during growth

Commercially raised feeder insects used to feed captive insectivores are a good source of many nutrients but are deficient in several key nutrients. Current methods used to supplement insects include dusting and gut‐loading. Here, we report on the nutrient composition of four species of commercially raised feeder insects fed a special diet to enhance their nutrient content. Crickets, mealworms, superworms, and waxworms were analyzed for moisture, crude protein, fat, ash, acid detergent fiber, total dietary fiber, minerals, amino acids, fatty acids, vitamins, taurine, carotenoids, inositol, and cholesterol. All four species contained enhanced levels of vitamin E and omega 3 fatty acids when compared to previously published data for these species. Crickets, superworms, and mealworms contained β‐carotene although using standard conversion factors only crickets and superworms would likely contain sufficient vitamin A activity for most species of insectivores. Waxworms did not contain any detectable β‐carotene but did contain zeaxanthin which they likely converted from dietary β‐carotene. All four species contained significant amounts of both inositol and cholesterol. Like previous reports all insects were a poor source of calcium and only superworms contained vitamin D above the limit of detection. When compared to the nutrient requirements as established by the NRC for growing rats or poultry, these species were good sources of most other nutrients although the high fat and low moisture content of both waxworms and superworms means when corrected for energy density these two species were deficient in more nutrients than crickets or mealworms. These data show the value of modifying the diet of commercially available insects as they are growing to enhance their nutrient content. They also suggest that for most insectivores properly supplemented lower fat insects such as crickets, or smaller mealworms should form the bulk of the diet. Zoo Biol. 34:554–564, 2015. © 2015 The Authors. Zoo Biology published by Wiley Periodicals, Inc.     

The Sound and the Fury—Bees Hiss when Expecting Danger

Honey bees are important model systems for the investigation of learning and memory and for a better understanding of the neuronal basics of brain function. Honey bees also possess a rich repertoire of tones and sounds, from queen piping and quacking to worker hissing and buzzing. In this study, we tested whether the worker bees’ sounds can be used as a measure of learning. We therefore conditioned honey bees aversively to odours in a walking arena and recorded both their sound production and their movement. Bees were presented with two odours, one of which was paired with an electric shock. Initially, the bees did not produce any sound upon odour presentation, but responded to the electric shock with a strong hissing response. After learning, many bees hissed at the presentation of the learned odour, while fewer bees hissed upon presentation of another odour. We also found that hissing and movement away from the conditioned odour are independent behaviours that can co-occur but do not necessarily do so. Our data suggest that hissing can be used as a readout for learning after olfactory conditioning, but that there are large individual differences between bees concerning their hissing reaction. The basis for this variability and the possible ecological relevance of the bees’ hissing remain to be investigated.     

The attractiveness of different odour sources from the fruit–host complex on Leptopilina boulardi, a larval parasitoid of frugivorous Drosophila spp.

In parasitoid insects, successful offspring development depends on the female’s ability to find a suitable host. Specific recognition is often based on responses to olfactory cues, but their source and nature have rarely been determined. –This paper deals with the recognition of odours involved in host location by Leptopilina boulardi[Barbotin, Carton & Kelner-Pillault] (Hymenoptera: Eucoilidae), a larval parasitoid of Drosophila species that develops in mature fruits. The nature and origin of volatile stimuli recognized among odours of the host–fruit complex, and the effect of learning on this recognition, were investigated. Oriented responses to these odours were observed in a four-armed olfactometer and were analysed with the observer software (Noldus Information Technology). Fruit odours alone (banana and pear) were not spontaneously attractive to naive parasitoids, whereas naturally-infested bananas were highly attractive. The attraction was related to the odour that adult Drosophila left on the substrate but not to Drosophila oviposition activity or larval development. A synergism between some fruit odours (banana and pear) and the odour left by adult Drosophila on damp filter paper was observed. However, when testing a non-fruit substrate (mushroom), no synergism was observed. Thus, female L. boulardi may innately recognize host–food substrate odours associated with odours from the adult stage of their host. In addition, an oviposition experience on an infested banana allows L. boulardi females to memorise the fruit odour itself through associative learning. The adaptive significance of this process is discussed.     

Divergent responses of two cereal aphids to previous infestation of their host plant

The role of pollen odour in resource location by the pollen beetle, Meligethes aeneus (Fabricius) (Coleoptera: Nitidulidae), a pollen-feeding insect regarded as a pest of oilseed rape, Brassica napus L., (Brassicaceae) crops, was investigated in a linear track olfactometer. Both male and female beetles were attracted to the odour of whole oilseed rape flowers, indicating that these insects can locate their host plants using floral odours as cues. The attractive odour of flowers was found to emanate from all floral parts tested: the petals/sepals, the anthers, and from pollen itself. Therefore, at least part of the attractive odour of oilseed rape flowers emanates from pollen. Beetles were more attracted to floral samples containing anthers than those without anthers when these odours were directly compared in a choice-test, and this indicates that there were detectable differences between them. Anthers and pollen may therefore release distinctive odours that are quantitatively and/or qualitatively different from the odour of the rest of the flower. These experiments support the hypothesis that pollen-seeking insects use pollen odour cues to locate this food source.     

Drosophila rely on learning while foraging under semi‐natural conditions

Learning is predicted to affect manifold ecological and evolutionary processes, but the extent to which animals rely on learning in nature remains poorly known, especially for short‐lived non‐social invertebrates. This is in particular the case for Drosophila, a favourite laboratory system to study molecular mechanisms of learning. Here we tested whether Drosophila melanogaster use learned information to choose food while free‐flying in a large greenhouse emulating the natural environment. In a series of experiments flies were first given an opportunity to learn which of two food odours was associated with good versus unpalatable taste; subsequently, their preference for the two odours was assessed with olfactory traps set up in the greenhouse. Flies that had experienced palatable apple‐flavoured food and unpalatable orange‐flavoured food were more likely to be attracted to the odour of apple than flies with the opposite experience. This was true both when the flies first learned in the laboratory and were then released and recaptured in the greenhouse, and when the learning occurred under free‐flying conditions in the greenhouse. Furthermore, flies retained the memory of their experience while exploring the greenhouse overnight in the absence of focal odours, pointing to the involvement of consolidated memory. These results support the notion that even small, short lived insects which are not central‐place foragers make use of learned cues in their natural environments.     

Reward-motivated learning: mesolimbic activation precedes memory formation

We examined anticipatory mechanisms of reward-motivated memory formation using event-related FMRI. In a monetary incentive encoding task, cues signaled high- or low-value reward for memorizing an upcoming scene. When tested 24 hr postscan, subjects were significantly more likely to remember scenes that followed cues for high-value rather than low-value reward. A monetary incentive delay task independently localized regions responsive to reward anticipation. In the encoding task, high-reward cues preceding remembered but not forgotten scenes activated the ventral tegmental area, nucleus accumbens, and hippocampus. Across subjects, greater activation in these regions predicted superior memory performance. Within subject, increased correlation between the hippocampus and ventral tegmental area was associated with enhanced long-term memory for the subsequent scene. These findings demonstrate that brain activation preceding stimulus encoding can predict declarative memory formation. The findings are consistent with the hypothesis that reward motivation promotes memory formation via dopamine release in the hippocampus prior to learning.     

A combined perceptual, physico-chemical, and imaging approach to 'odour-distances' suggests a categorizing function of the Drosophila antennal lobe

How do physico-chemical stimulus features, perception, and physiology relate? Given the multi-layered and parallel architecture of brains, the question specifically is where physiological activity patterns correspond to stimulus features and/or perception. Perceived distances between six odour pairs are defined behaviourally from four independent odour recognition tasks. We find that, in register with the physico-chemical distances of these odours, perceived distances for 3-octanol and n-amylacetate are consistently smallest in all four tasks, while the other five odour pairs are about equally distinct. Optical imaging in the antennal lobe, using a calcium sensor transgenically expressed in only first-order sensory or only second-order olfactory projection neurons, reveals that 3-octanol and n-amylacetate are distinctly represented in sensory neurons, but appear merged in projection neurons. These results may suggest that within-antennal lobe processing funnels sensory signals into behaviourally meaningful categories, in register with the physico-chemical relatedness of the odours.     

The orientation of walking honeybees in odour fields with small concentration gradients

The behaviour of walking honeybees in small gradient odour fields was investigated by means of a simulation technique. The bee was kept in one place by a locomotion compensator (‘running sphere’). This compensator allowed for a precise reconstruction of the bee's actual locomotion on the sphere, and presented the bee with a stimulating odour whose concentration was controlled by feedback from the reconstructed locomotion. This rendered possible the application of well-defined odour fields and revealed that: (1) honeybees are capable of finding odour sources in the absence of optical cues and with concentration gradients too small to allow tropotactic or klinotactic orientation; (2) bees are capable of memorizing odour concentrations with a high degree of accuracy; (3) this orientation system is based on a switching over from negative to positive anemotaxis at a ‘reference’ concentration; (4) this reference is a function of the odour concentration at which a sugar reward is given. The results do not support any hypothesis for an orientation system based on the detection and comparison of successive values of odour concentration. A hypothesis on the nature of the ‘reference value’ is discussed and supported by experiments.     

Odour-genes eovariance and differential investigation of individual odours in the Mus species complex

Previous research using habituation techniques has demonstrated that greater genetic similarity between two individuals is associated with more similarity in the qualities of their individual odours ('odour-genes eovariance'). We assessed odour similarities across species in two pairs of genetically close species within the Mus species complex (M. musculus and M. domesticus; M. spicilegus and M. macedonicus). Subjects treated odours within each species pair as similar compared with an odour from the other species pair. Subjects also treated odours of M. spicilegus males from the same population as similar compared with the odour of M. spicilegus males from a different population. This confirms odour-genes eovariance across species and within populations and also supports previous findings that odour similarities are more salient than specific odour markers. When adult males were presented with odours of females from two different heterospecific species, subjects spent more time investigating the odour from his own species pair than the other species pair, indicating greater interest in the odour of the closer heterospecific and demonstrating that odour-genes eovariance is reflected in behavioural responses to odours. Implications of odour-genes eovariance as a basis for identifying degrees of genetic relatedness of unfamiliar individuals through similarities in individual odours are discussed.     

Modelling efficiency in insect olfactory information processing

The olfactory system of insects is essential for the search of food and mates, and weak signals can be detected, amplified and discriminated in a fluctuating environment. The olfactory system also allows for learning and recall of odour memories. Based on anatomical, physiological, and behavioural data from the olfactory system of insects, we have developed a cross-scale dynamical neural network model to simulate the presentation, amplification and discrimination of host plant odours and sex pheromones. In particular, we model how the spatial and temporal patterns of the odour information emerging in the glomeruli of the antennal lobe (AL) rely on the glomerular morphology, the connectivity and the complex dynamics of the AL circuits. We study how weak signals can be amplified, how different odours can be discriminated, based on stochastic (resonance) dynamics and the connectivity of the network. We further investigate the spatial and temporal coding of sex pheromone components and plant volatile compounds, in relation to the glomerular structure, arborizing patterns of the projection neurons (PNs) and timing patterns of the neuronal spiking activity.     

Social learning of floral odours inside the honeybee hive

A honeybee hive serves as an information centre in which communication among bees allows the colony to exploit the most profitable resources in a continuously changing environment. The best-studied communication behaviour in this context is the waggle dance performed by returning foragers, which encodes information about the distance and direction to the food source. It has been suggested that another information cue, floral scents transferred within the hive, is also important for recruitment to food sources, as bee recruits are more strongly attracted to odours previously brought back by foragers in both honeybees and bumble-bees. These observations suggested that honeybees learn the odour from successful foragers before leaving the hive. However, this has never been shown directly and the mechanisms and properties of the learning process remain obscure. We tested the learning and memory of recruited bees in the laboratory using the proboscis extension response (PER) paradigm, and show that recruits indeed learn the nectar odours brought back by foragers by associative learning and retrieve this memory in the PER paradigm. The associative nature of this learning reveals that information was gained during mouth-to-mouth contacts among bees (trophallaxis). Results further suggest that the information is transferred to long-term memory. Associative learning of food odours in a social context may help recruits to find a particular food source faster.