Extracting spatial information from temporal odor patterns: insights from insects.

Extracting spatial information from temporal stimulus patterns is essential for sensory perception (e.g. visual motion direction detection or concurrent sound segregation), but this process remains understudied in olfaction. Animals rely on olfaction to locate resources and dangers. In open environments, where odors are dispersed by turbulent wind, detection of wind direction seems crucial for odor source localization. However, recent studies showed that insects can extract spatial information from the odor stimulus itself, independently from sensing wind direction. This remarkable ability is achieved by detecting the fine-scale temporal pattern of odor encounters, which contains information about the location and size of an odor source, and the distance between different odor sources.

Characterisation and automated quantification of induced seizure-related behaviours in Xenopus laevis tadpoles.

Epilepsy, a clinical diagnosis characterised by paroxysmal episodes known as seizures, affects 1% of people worldwide. Safe and patient-specific treatment is vital and can be achieved by the development of rapid pre-clinical models of for identified epilepsy genes. Epilepsy can result from either brain injury or gene mutations, and can also be induced chemically. Xenopus laevis tadpoles could be a useful model for confirmation of variants of unknown significance found in epilepsy patients, and for drug re-purposing screens that could eventually lead to benefits for patients. Here, we characterise and quantify seizure-related behaviours in X. laevis tadpoles arrayed in 24-well plates. To provoke acute seizure behaviours, tadpoles were chemically induced with either pentylenetetrazole (PTZ) or 4-aminopyridine (4-AP). To test the capacity to adapt this method for drug testing, we also exposed induced tadpoles to the anti-seizure drug valproate (VPA). Four induced seizure-like behaviours were described and manually quantified, and two of these (darting, circling) could be accurately detected automatically, using the video analysis software TopScan. Additionally, we recorded swimming trajectories and mean swimming velocity. Automatic detection showed that either PTZ or 4-AP induced darting behaviour and increased mean swimming velocity compared to untreated controls. Both parameters were significantly reduced in the presence of VPA. In particular, darting behaviour was a shown to be a sensitive measure of epileptic seizure activity. While we could not automatically detect the full range of seizure behaviours, this method shows promise for future studies since X. laevis is a well-characterised and genetically tractable model organism.

Reduced olfactory acuity in recently flightless insects suggests rapid regressive evolution.

BACKGROUND: Insects have exceptionally fast smelling capabilities, and some can track the temporal structure of odour plumes at rates above 100 Hz. It has been hypothesized that this fast smelling capability is an adaptation for flying. We test this hypothesis by comparing the olfactory acuity of sympatric flighted versus flightless lineages within a wing-polymorphic stonefly species. RESULTS: Our analyses of olfactory receptor neuron responses reveal that recently-evolved flightless lineages have reduced olfactory acuity. By comparing flighted versus flightless ecotypes with similar genetic backgrounds, we eliminate other confounding factors that might have affected the evolution of their olfactory reception mechanisms. Our detection of different patterns of reduced olfactory response strength and speed in independently wing-reduced lineages suggests parallel evolution of reduced olfactory acuity. CONCLUSIONS: These reductions in olfactory acuity echo the rapid reduction of wings themselves, and represent an olfactory parallel to the convergent phenotypic shifts seen under selective gradients in other sensory systems (e.g. parallel loss of vision in cave fauna). Our study provides evidence for the hypothesis that flight poses a selective pressure on the speed and strength of olfactory receptor neuron responses and emphasizes the energetic costs of rapid olfaction.

Sugar alcohols have the potential as bee-safe baits for the common wasp.

BACKGROUND: Pest insects are often baited with poisoned feeding stimulants, the most common of which are sugars. However, sugars are attractive for most animal species, which makes it difficult to target only a specific pest insect species. Here, we assessed different sugar alcohols for their potential as more species-selective feeding stimulants for pest insects. RESULTS: We tested the attractiveness of the sugar alcohols sorbitol, xylitol and erythritol with a capillary feeder assay in wasps (as potential pest insects, because introduced wasps are a pest in many regions) and bees (as non-target insects). For the common wasp (Vespula vulgaris), sorbitol and xylitol acted as nutritive feeding stimulants, and erythritol acted as a non-nutritive feeding stimulant. For the buff-tailed bumble bee (Bombus terrestris), sorbitol acted as a feeding stimulant, while for the honey bee (Apis mellifera), none of the sugar alcohols acted as feeding stimulant. CONCLUSION: The species-specific preferences for sugar alcohols suggest their potential as species-selective insect baits. The wasp-specific preference for xylitol suggests its potential as a bee-safe alternative to sugar-containing bait for controlling the common wasp. © 2022 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Alpha oscillations govern interhemispheric spike timing coordination in the honey bee brain.

In 1929 Hans Berger discovered the alpha oscillations: prominent, ongoing oscillations around 10 Hz in the electroencephalogram of the human brain. These alpha oscillations are among the most widely studied brain signals, related to cognitive phenomena such as attention, memory and consciousness. However, the mechanisms by which alpha oscillations affect human cognition await demonstration. Here, we suggest the honey bee brain as an experimentally more accessible model system for investigating the functional role of alpha oscillations. We found a prominent spontaneous oscillation around 18 Hz that is reduced in amplitude upon olfactory stimulation. Similar to alpha oscillations in primates, the phase of this oscillation biased both timing of neuronal spikes and amplitude of high-frequency gamma activity (40-450 Hz). These results suggest a common role of alpha oscillations across phyla and provide an unprecedented new venue for causal studies on the relationship between neuronal spikes, brain oscillations and cognition.

Social foraging extends associative odor-food memory expression in an automated learning assay for Drosophila melanogaster.

Animals socially interact during foraging and share information about the quality and location of food sources. The mechanisms of social information transfer during foraging have been mostly studied at the behavioral level, and its underlying neural mechanisms are largely unknown. Fruit flies have become a model for studying the neural bases of social information transfer, because they provide a large genetic toolbox to monitor and manipulate neuronal activity, and they show a rich repertoire of social behaviors. Fruit flies aggregate, they use social information for choosing a suitable mating partner and oviposition site, and they show better aversive learning when in groups. However, the effects of social interactions on associative odor-food learning have not yet been investigated. Here, we present an automated learning and memory assay for walking flies that allows the study of the effect of group size on social interactions and on the formation and expression of associative odor-food memories. We found that both inter-fly attraction and the duration of odor-food memory expression increase with group size. This study opens up opportunities to investigate how social interactions during foraging are relayed in the neural circuitry of learning and memory expression.

Segregation of Unknown Odors From Mixtures Based on Stimulus Onset Asynchrony in Honey Bees.

Animals use olfaction to search for distant objects. Unlike vision, where objects are spaced out, olfactory information mixes when it reaches olfactory organs. Therefore, efficient olfactory search requires segregating odors that are mixed with background odors. Animals can segregate known odors by detecting short differences in the arrival of mixed odorants (stimulus onset asynchrony). However, it is unclear whether animals can also use stimulus onset asynchrony to segregate odorants that they had no previous experience with and which have no innate or learned relevance (unknown odorants). Using behavioral experiments in honey bees, we here show that stimulus onset asynchrony also improves segregation of those unknown odorants. The stimulus onset asynchrony necessary to segregate unknown odorants is in the range of seconds, which is two orders of magnitude larger than the previously reported stimulus asynchrony sufficient for segregating known odorants. We propose that for unknown odorants, segregating odorant A from a mixture with B requires sensory adaptation to B.

Olfactory Object Recognition Based on Fine-Scale Stimulus Timing in Drosophila.

Odorants of behaviorally relevant objects (e.g., food sources) intermingle with those from other sources. Therefore to determine whether an odor source is good or bad-without actually visiting it-animals first need to segregate the odorants from different sources. To do so, animals could use temporal stimulus cues, because odorants from one source exhibit correlated fluctuations, whereas odorants from different sources are less correlated. However, the behaviorally relevant timescales of temporal stimulus cues for odor source segregation remain unclear. Using behavioral experiments with free-flying flies, we show that (1) odorant onset asynchrony increases flies' attraction to a mixture of two odorants with opposing innate or learned valence and (2) attraction does not increase when the attractive odorant arrives first. These data suggest that flies can use stimulus onset asynchrony for odor source segregation and imply temporally precise neural mechanisms for encoding odors and for segregating them into distinct objects.

Odorant mixtures elicit less variable and faster responses than pure odorants.

In natural environments, odors are typically mixtures of several different chemical compounds. However, the implications of mixtures for odor processing have not been fully investigated. We have extended a standard olfactory receptor model to mixtures and found through its mathematical analysis that odorant-evoked activity patterns are more stable across concentrations and first-spike latencies of receptor neurons are shorter for mixtures than for pure odorants. Shorter first-spike latencies arise from the nonlinear dependence of binding rate on odorant concentration, commonly described by the Hill coefficient, while the more stable activity patterns result from the competition between different ligands for receptor sites. These results are consistent with observations from numerical simulations and physiological recordings in the olfactory system of insects. Our results suggest that mixtures allow faster and more reliable olfactory coding, which could be one of the reasons why animals often use mixtures in chemical signaling.

High Precision of Spike Timing across Olfactory Receptor Neurons Allows Rapid Odor Coding in Drosophila.

In recent years, it has become evident that olfaction is a fast sense, and millisecond short differences in stimulus onsets are used by animals to analyze their olfactory environment. In contrast, olfactory receptor neurons are thought to be relatively slow and temporally imprecise. These observations have led to a conundrum: how, then, can an animal resolve fast stimulus dynamics and smell with high temporal acuity? Using parallel recordings from olfactory receptor neurons in Drosophila, we found hitherto unknown fast and temporally precise odorant-evoked spike responses, with first spike latencies (relative to odorant arrival) down to 3 ms and with a SD below 1 ms. These data provide new upper bounds for the speed of olfactory processing and suggest that the insect olfactory system could use the precise spike timing for olfactory coding and computation, which can explain insects' rapid processing of temporal stimuli when encountering turbulent odor plumes.

Corrigendum: Calcium in Kenyon Cell Somata as a Substrate for an Olfactory Sensory Memory in Drosophila.

[This corrects the article on p. 128 in vol. 12, PMID: 29867361.].

Calcium in Kenyon Cell Somata as a Substrate for an Olfactory Sensory Memory in Drosophila.

Animals can form associations between temporally separated stimuli. To do so, the nervous system has to retain a neural representation of the first stimulus until the second stimulus appears. The neural substrate of such sensory stimulus memories is unknown. Here, we search for a sensory odor memory in the insect olfactory system and characterize odorant-evoked Ca2+ activity at three consecutive layers of the olfactory system in Drosophila: in olfactory receptor neurons (ORNs) and projection neurons (PNs) in the antennal lobe, and in Kenyon cells (KCs) in the mushroom body. We show that the post-stimulus responses in ORN axons, PN dendrites, PN somata, and KC dendrites are odor-specific, but they are not predictive of the chemical identity of past olfactory stimuli. However, the post-stimulus responses in KC somata carry information about the identity of previous olfactory stimuli. These findings show that the Ca2+ dynamics in KC somata could encode a sensory memory of odorant identity and thus might serve as a basis for associations between temporally separated stimuli.

The Role of the Sucrose-Responsive IR60b Neuron for Drosophila melanogaster: A Hypothesis.

In a recent paper, Joseph and colleagues (Joseph et al. 2017) have characterized an IR60b receptor-expressing neuron in Drosophila. They showed that it responds to sucrose and serves to limit sucrose consumption, and proposed that it may thereby act to prevent overfeeding. Here, we propose an alternative hypothesis for the functional role of sucrose feeding control, and for how this limitation of sucrose uptake is accomplished. Adult fruit flies feed by excreting saliva onto the food, and imbibing the predigested liquefied food, or by filling the crop, where the food is predigested. Enzymes in the saliva hydrolyze starch and disaccharides into absorbable monosaccharides. Premature ingestion into the midgut would not give the enzymes in the saliva enough time to predigest the food. Thus, IR60b neurons might serve as a sensor to monitor the digestive state of external food or crop content: when disaccharides (sucrose) concentration is high, ingestion to the gut is inhibited, keeping a low concentration of starch and disaccharides in the midgut.

Task-Related Phasing of Circadian Rhythms in Antennal Responsiveness to Odorants and Pheromones in Honeybees.

The insect antennae receive olfactory information from the environment. In some insects, it has been shown that antennal responsiveness is dynamically regulated by circadian clocks. However, it is unknown how general this phenomenon is and what functions it serves. Circadian regulation in honeybee workers is particularly interesting in this regard because they show natural task-related chronobiological plasticity. Forager bees show strong circadian rhythms in behavior and brain gene expression, whereas nurse bees tend brood around-the-clock and have attenuated circadian rhythms in activity and whole-brain gene expression. Here, we tested the hypothesis that there is task-related plasticity in circadian rhythms of antennal responsiveness to odorants in worker honeybees. We used electroantennogram (EAG) to measure the antennal responsiveness of nurses and foragers to general odorants and pheromones around the day. The capacity to track 10-Hz odorant pulses varied with time of day for both task groups but with different phases. The antennal pulse-tracking capacity was higher during the subjective day for the day-active foragers, whereas it was better during the night for around-the-clock active nurses. The task-related phases of pulse-tracking rhythms were similar for all the tested stimuli. We also found evidence for circadian rhythms in the EAG response magnitude of foragers but not of nurses. To the best of our knowledge, these results provide the first evidence for circadian regulation of antennal olfactory responsiveness and odorant pulse-tracking capacity in bees or any other hymenopteran insect. Importantly, our study shows for the first time that the circadian phase of olfactory responsiveness may be socially regulated.

Trace Conditioning in Drosophila Induces Associative Plasticity in Mushroom Body Kenyon Cells and Dopaminergic Neurons.

Dopaminergic neurons (DANs) signal punishment and reward during associative learning. In mammals, DANs show associative plasticity that correlates with the discrepancy between predicted and actual reinforcement (prediction error) during classical conditioning. Also in insects, such as Drosophila, DANs show associative plasticity that is, however, less understood. Here, we study associative plasticity in DANs and their synaptic partners, the Kenyon cells (KCs) in the mushroom bodies (MBs), while training Drosophila to associate an odorant with a temporally separated electric shock (trace conditioning). In most MB compartments DANs strengthened their responses to the conditioned odorant relative to untrained animals. This response plasticity preserved the initial degree of similarity between the odorant- and the shock-induced spatial response patterns, which decreased in untrained animals. Contrary to DANs, KCs (α'/ß'-type) decreased their responses to the conditioned odorant relative to untrained animals. We found no evidence for prediction error coding by DANs during conditioning. Rather, our data supports the hypothesis that DAN plasticity encodes conditioning-induced changes in the odorant's predictive power.

A High-Bandwidth Dual-Channel Olfactory Stimulator for Studying Temporal Sensitivity of Olfactory Processing.

Animals encounter fine-scale temporal patterns of odorant mixtures that contain information about the distance and number of odorant sources. To study the role of such temporal cues for odorant detection and source localization, one needs odorant delivery devices that are capable of mimicking the temporal stimulus statistics of natural odor plumes. However, current odorant delivery devices either lack temporal resolution or are limited to a single odorant channel. Here, we present an olfactory stimulator that features precise control of high-bandwidth stimulus dynamics, which allows generating arbitrary fluctuating binary odorant mixtures. We provide a comprehensive characterization of the stimulator's performance and use it to demonstrate that odor background affects the temporal resolution of insect olfactory receptor neurons, and we present a hitherto unknown odor pulse-tracking capability of up to 60 Hz in Kenyon cells, which are higher order olfactory neurons of the insect brain. This stimulator might help investigating whether and how animals use temporal stimulus cues for odor detection and source localization. Because the stimulator is easy to replicate it can facilitate generating the same odor stimulus dynamics at different experimental setups and across different labs.

Asymmetric neural coding revealed by in vivo calcium imaging in the honey bee brain.

Left-right asymmetries are common properties of nervous systems. Although lateralized sensory processing has been well studied, information is lacking about how asymmetries are represented at the level of neural coding. Using in vivo functional imaging, we identified a population-level left-right asymmetry in the honey bee's primary olfactory centre, the antennal lobe (AL). When both antennae were stimulated via a frontal odour source, the inter-odour distances between neural response patterns were higher in the right than in the left AL. Behavioural data correlated with the brain imaging results: bees with only their right antenna were better in discriminating a target odour in a cross-adaptation paradigm. We hypothesize that the differences in neural odour representations in the two brain sides serve to increase coding capacity by parallel processing.

Automated tracking and analysis of behavior in restrained insects.

BACKGROUND: Insect behavior is often monitored by human observers and measured in the form of binary responses. This procedure is time costly and does not allow a fine graded measurement of behavioral performance in individual animals. To overcome this limitation, we have developed a computer vision system which allows the automated tracking of body parts of restrained insects. NEW METHOD: Our system crops a continuous video into separate shots with a static background. It then segments out the insect's head and preprocesses the detected moving objects to exclude detection errors. A Bayesian-based algorithm is proposed to identify the trajectory of each body part. RESULTS: We demonstrate the application of this novel tracking algorithm by monitoring movements of the mouthparts and antennae of honey bees and ants, and demonstrate its suitability for analyzing the behavioral performance of individual bees using a common associative learning paradigm. COMPARISON WITH EXISTING METHODS: Our tracking system differs from existing systems in that it does not require each video to be labeled manually and is capable of tracking insects' body parts even when working with low frame-rate videos. Our system can be generalized for other insect tracking applications. CONCLUSIONS: Our system paves the ground for fully automated monitoring of the behavior of restrained insects and accounts for individual variations in graded behavior.

High-speed odor transduction and pulse tracking by insect olfactory receptor neurons.

Sensory systems encode both the static quality of a stimulus (e.g., color or shape) and its kinetics (e.g., speed and direction). The limits with which stimulus kinetics can be resolved are well understood in vision, audition, and somatosensation. However, the maximum temporal resolution of olfactory systems has not been accurately determined. Here, we probe the limits of temporal resolution in insect olfaction by delivering high frequency odor pulses and measuring sensory responses in the antennae. We show that transduction times and pulse tracking capabilities of olfactory receptor neurons are faster than previously reported. Once an odorant arrives at the boundary layer of the antenna, odor transduction can occur within less than 2 ms and fluctuating odor stimuli can be resolved at frequencies more than 100 Hz. Thus, insect olfactory receptor neurons can track stimuli of very short duration, as occur when their antennae encounter narrow filaments in an odor plume. These results provide a new upper bound to the kinetics of odor tracking in insect olfactory receptor neurons and to the latency of initial transduction events in olfaction.