Honey bees infer source location from the dances of returning foragers.

Honeybees (Apis mellifera carnica) communicate the direction and distance to a food source by means of a waggle dance. We ask whether bees recruited by the dance use it only as a flying instruction, with the technical form of a polar vector, or also translate it into a location vector that enables them to set courses directed toward the food source from arbitrary locations within their familiar territory. The flights of recruits captured on exiting the hive and released at distant sites were tracked by radar. The recruits performed first a straight flight in approximately the compass direction indicated by the dance. However, this "vector" portion of their flights and the ensuing tortuous "search" portion were strongly and differentially affected by the release site. Searches were biased toward the true location of the food and away from the location specified by translating the origin for the danced polar vector to the release site. We conclude that by following the dance recruits get two messages, a polar flying instruction (bearing and range from the hive) and a location vector that enables them to approach the source from anywhere in their familiar territory. The dance communication is much richer than thought so far.

Insights from the past: the work of Hans von Alten on the evolution of brain structure, ecological adaptation, and cognition in hymenopteran species.

In his treatise on arthropod brains, Hans von Alten (1910) focuses on a specific functional group of insects-the flying Hymenoptera-which exhibit a spectrum of lifestyles ranging from solitary to social. His work presents a distinctive comparative neuro-anatomical approach rooted in an eco-evolutionary and eco-behavioral background. We regard his publication as an exceptionally valuable source of information and seek to inspire the research community dedicated to the study of the insect brain to explore its insights further, even after more than 110 years. We have translated and annotated his work, expecting it to engage researchers not just with its remarkable drawings but also with its substantive content and exemplary research strategy. The present text is designed to complement von Alten's publication, situating it within the temporal context of nineteenth-century and early twentieth-century studies, and to draw connections to contemporary perspectives, especially concerning a central brain structure: the mushroom body.

Unpacking the navigation toolbox: insights from comparative cognition.

The study of navigation is informed by ethological data from many species, laboratory investigation at behavioural and neurobiological levels, and computational modelling. However, the data are often species-specific, making it challenging to develop general models of how biology supports behaviour. Wiener et al. outlined a framework for organizing the results across taxa, called the 'navigation toolbox' (Wiener et al. In Animal thinking: contemporary issues in comparative cognition (eds R Menzel, J Fischer), pp. 51-76). This framework proposes that spatial cognition is a hierarchical process in which sensory inputs at the lowest level are successively combined into ever-more complex representations, culminating in a metric or quasi-metric internal model of the world (cognitive map). Some animals, notably humans, also use symbolic representations to produce an external representation, such as a verbal description, signpost or map that allows communication of spatial information or instructions between individuals. Recently, new discoveries have extended our understanding of how spatial representations are constructed, highlighting that the hierarchical relationships are bidirectional, with higher levels feeding back to influence lower levels. In the light of these new developments, we revisit the navigation toolbox, elaborate it and incorporate new findings. The toolbox provides a common framework within which the results from different taxa can be described and compared, yielding a more detailed, mechanistic and generalized understanding of navigation.

Navigation and dance communication in honeybees: a cognitive perspective.

Flying insects like the honeybee experience the world as a metric layout embedded in a compass, the time-compensated sun compass. The focus of the review lies on the properties of the landscape memory as accessible by data from radar tracking and analyses of waggle dance following. The memory formed during exploration and foraging is thought to be composed of multiple elements, the aerial pictures that associate the multitude of sensory inputs with compass directions. Arguments are presented that support retrieval and use of landscape memory not only during navigation but also during waggle dance communication. I argue that bees expect landscape features that they have learned and that are retrieved during dance communication. An intuitive model of the bee's navigation memory is presented that assumes the picture memories form a network of geographically defined locations, nodes. The intrinsic components of the nodes, particularly their generalization process leads to binding structures, the edges. In my view, the cognitive faculties of landscape memory uncovered by these experiments are best captured by the term cognitive map.

Innovation in solitary bees is driven by exploration, shyness and activity levels.

Behavioural innovation and problem solving are widely considered to be important mechanisms by which animals respond to novel environmental challenges, including those induced by human activities. Despite their functional and ecological relevance, much of our current understanding of these processes comes from studies in vertebrates. Understanding of these processes in invertebrates has lagged behind partly because they are not perceived to have the cognitive machinery required. This perception is, however, challenged by recent evidence demonstrating sophisticated cognitive capabilities in insects despite their small brains. Here, we studied innovation, defined as the capacity to solve a new task, of a solitary bee (Osmia cornuta) in the laboratory by exposing naive individuals to an obstacle removal task. We also studied the underlying cognitive and non-cognitive mechanisms through a battery of experimental tests designed to measure associative learning, exploration, shyness and activity levels. We found that solitary bees can innovate, with 11 of 29 individuals (38%) being able to solve a new task consisting of lifting a lid to reach a reward. However, the propensity to innovate was uncorrelated with the measured learning capacity, but increased with exploration, boldness and activity. These results provide solid evidence that non-social insects can solve new tasks, and highlight the importance of interpreting innovation in the light of non-cognitive processes.

The neonicotinoid clothianidin impairs memory processing in honey bees.

Neonicotinoids act as agonists on the nicotinic Acetylcholine receptor (nAChR) in insect brains, an essential molecular component of central brain structures involved in learning and memory formation. Sublethal doses might, therefore, impair neural processes necessary for adaptive experience dependent behaviour and thus reduce the fitness of pollinating insects on the individual and community level. First, the question was addressed whether clothianidin has an aversive taste for honey bees and concluded with both a laboratory and a semi-field experiment that bees are unable to distinguish between control and contaminated sucrose solutions. In the laboratory, proboscis extension response conditioning was performed with forager bees exposed to different concentrations of clothianidin (0.1, 0.3 and 0.8 ng/bee) before learning, after learning during memory consolidation, and just before memory retention. These tests at different timings allowed uncovering an impairment of the consolidation and retrieval of memory due to the exposure to clothianidin. It was concluded that an acute exposure to clothianidin has an adverse effect on memory processing in honey bees.

A neural network model for familiarity and context learning during honeybee foraging flights.

How complex is the memory structure that honeybees use to navigate? Recently, an insect-inspired parsimonious spiking neural network model was proposed that enabled simulated ground-moving agents to follow learned routes. We adapted this model to flying insects and evaluate the route following performance in three different worlds with gradually decreasing object density. In addition, we propose an extension to the model to enable the model to associate sensory input with a behavioral context, such as foraging or homing. The spiking neural network model makes use of a sparse stimulus representation in the mushroom body and reward-based synaptic plasticity at its output synapses. In our experiments, simulated bees were able to navigate correctly even when panoramic cues were missing. The context extension we propose enabled agents to successfully discriminate partly overlapping routes. The structure of the visual environment, however, crucially determines the success rate. We find that the model fails more often in visually rich environments due to the overlap of features represented by the Kenyon cell layer. Reducing the landmark density improves the agents route following performance. In very sparse environments, we find that extended landmarks, such as roads or field edges, may help the agent stay on its route, but often act as strong distractors yielding poor route following performance. We conclude that the presented model is valid for simple route following tasks and may represent one component of insect navigation. Additional components might still be necessary for guidance and action selection while navigating along different memorized routes in complex natural environments.

The honeybee as a model for understanding the basis of cognition.

Honeybees contradict the notion that insect behaviour tends to be relatively inflexible and stereotypical. Indeed, they live in colonies and exhibit complex social, navigational and communication behaviours, as well as a relatively rich cognitive repertoire. Because these relatively complex behaviours are controlled by a brain consisting of only 1 million or so neurons, honeybees offer an opportunity to study the relationship between behaviour and cognition in neural networks that are limited in size and complexity. Most recently, the honeybee has been used to model learning and memory formation, highlighting its utility for neuroscience research, in particular for understanding the basis of cognition.

Effects of sublethal doses of thiacloprid and its formulation Calypso® on the learning and memory performance of honey bees.

Learning and memory play a central role in the behavior and communication of foraging bees. We have previously shown that chronic uptake of the neonicotinoid thiacloprid affects the behavior of honey bees in the field. Foraging behavior, homing success, navigation performance and social communication were impaired. Thiacloprid collected at a feeding site at low doses accumulates in foragers over time. Here, we applied a laboratory standard procedure (the proboscis-extension response conditioning) in order to assess which processes, acquisition, memory consolidation and/or memory retrieval were compromised after bees were fed either with thiacloprid or the formulation of thiacloprid named Calypso® at different sublethal doses. Extinction and generalization tests allowed us to investigate whether bees respond to a learned stimulus, and how selectively. We showed that thiacloprid, as active substance and as formulation, poses a substantial risk to honey bees by disrupting learning and memory functions. These data support and specify the data collected in the field.

Way-finding in displaced clock-shifted bees proves bees use a cognitive map.

Mammals navigate by means of a metric cognitive map. Insects, most notably bees and ants, are also impressive navigators. The question whether they, too, have a metric cognitive map is important to cognitive science and neuroscience. Experimentally captured and displaced bees often depart from the release site in the compass direction they were bent on before their capture, even though this no longer heads them toward their goal. When they discover their error, however, the bees set off more or less directly toward their goal. This ability to orient toward a goal from an arbitrary point in the familiar environment is evidence that they have an integrated metric map of the experienced environment. We report a test of an alternative hypothesis, which is that all the bees have in memory is a collection of snapshots that enable them to recognize different landmarks and, associated with each such snapshot, a sun-compass-referenced home vector derived from dead reckoning done before and after previous visits to the landmark. We show that a large shift in the sun-compass rapidly induced by general anesthesia does not alter the accuracy or speed of the homeward-oriented flight made after the bees discover the error in their initial postrelease flight. This result rules out the sun-referenced home-vector hypothesis, further strengthening the now extensive evidence for a metric cognitive map in bees.

Environment-specific modulation of odorant representations in the honeybee brain.

Ca2+ imaging techniques were applied to investigate the neuronal behavior of projection neurons in the honeybee antennal lobe (AL) to examine the effects of long-lasting adaptation on odorant coding. Responses to eight test odorants were measured before, during, and after an odor adaptation phase. Bees were exposed to the adapting odor for 30 min. Test odorant responses were only recorded from a sub-population of accessible glomeruli on the AL surface. Projection neurons, the output neurons of the antennal lobes, are projecting through the lateral, mediolateral, and medial AL tract to higher centers of the olfactory pathway. Due to our staining techniques, we primarily focused our study on projection neurons going through the lateral and medial tract. Test odorants comprised compounds with different functional groups (alcohol, aldehyde, ketone, and ester) representing floral and/or pheromone odorants. Strength and discriminability between combinatorial activity patterns induced by the test odorants were quantified. In two independent experiments, we investigated one group of animals adapted to a colony odor and another adapted to a synthetic odor. Within the experimental groups, we found test odorant responses either decreased or increased in AL projection neurons. Additionally, the discriminability between test odorant patterns became less distinct in the colony odor experiment and more distinct during adaptation in the synthetic mixture experiment. These results are interpreted as odor dependent adaptation effects, increasing or decreasing response strength and discriminability by altered neural coding mechanisms in the AL neuropile.

Neural correlates of side-specific odour memory in mushroom body output neurons.

Humans and other mammals as well as honeybees learn a unilateral association between an olfactory stimulus presented to one side and a reward. In all of them, the learned association can be behaviourally retrieved via contralateral stimulation, suggesting inter-hemispheric communication. However, the underlying neuronal circuits are largely unknown and neural correlates of across-brain-side plasticity have yet not been demonstrated. We report neural plasticity that reflects lateral integration after side-specific odour reward conditioning. Mushroom body output neurons that did not respond initially to contralateral olfactory stimulation developed a unique and stable representation of the rewarded compound stimulus (side and odour) predicting its value during memory retention. The encoding of the reward-associated compound stimulus is delayed by about 40 ms compared with unrewarded neural activity, indicating an increased computation time for the read-out after lateral integration.

Honey Bees' Behavior Is Impaired by Chronic Exposure to the Neonicotinoid Thiacloprid in the Field.

The decline of pollinators worldwide is of growing concern and has been related to the use of plant-protecting chemicals. Most studies have focused on three neonicotinoid insecticides (clothianidin, imidacloprid, and thiamethoxam) currently subject to a moratorium in the EU. Here, we focus on thiacloprid, a widely used cyano-substituted neonicotinoid thought to be less toxic to honey bees and of which use has increased in the last years. Honey bees (Apis mellifera carnica) were exposed chronically to thiacloprid in the field for several weeks at a sublethal concentration. Foraging behavior, homing success, navigation performance, and social communication were impaired, and thiacloprid residue levels increased both in the foragers and the nest mates over time. The effects observed in the field were not due to a repellent taste of the substance. For the first time, we present the necessary data for the risk evaluation of thiacloprid taken up chronically by honey bees in field conditions.

Mushroom body extrinsic neurons in the honeybee (Apis mellifera) brain integrate context and cue values upon attentional stimulus selection.

Multimodal GABA-immunoreactive feedback neurons in the honeybee brain connecting the output region of the mushroom body with its input are expected to tune the input to the mushroom body in an experience-dependent way. These neurons are known to change their rate responses to learned olfactory stimuli. In this work we ask whether these neurons also transmit learned attentional effects during multisensory integration. We find that a visual context and an olfactory cue change the rate responses of these neurons after learning according to the associated values of both context and cue. The learned visual context promotes attentional response selection by enhancing olfactory stimulus valuation at both the behavioral and the neural level. During a rewarded visual context, bees reacted faster and more reliably to a rewarded odor. We interpreted this as the result of the observed enhanced neural discharge toward the odor. An unrewarded context reduced already low rate responses to the unrewarded odor. In addition to stimulus valuation, these feedback neurons generate a neural error signal after an incorrect behavioral response. This might act as a learning signal in feedback neurons. All of these effects were exclusively found in trials in which the animal prepares for a motor response that happens during attentional stimulus selection. We discuss possible implications of the results for the feedback connections of the mushroom body.

The memory structure of navigation in honeybees.

The analytical approach to navigation studies aims to identify elementary sensory motor processes that guide an animal to a remote site. This approach will be used here to characterize components of navigation in a flying insect, the honeybee. However, navigation studies need to go beyond an analysis of behavioral routines to come up with a synthesis. We will defend the concept of an active memory structure guiding navigation in bees that is best described as a mental or cognitive map. In our opinion, spatial/temporal relations of landmarks are stored in a mental map in such a way that behavioral routines such as expectation and planning, as indicated by shortcutting, are possible. We view the mental map of animals including the honeybee as an "action memory of spatial relations" rather than as a sensory representation as we humans experience it by introspection. Two components characterize the mental map, the relational representation of landmarks and the meaning of locations to the animal. As yet, there is little data to suggest that bees assign meaning to the experienced locations. To explore this possibility, further studies will be needed, whereby honeybees provide a unique model to address this question.

The neonicotinoid clothianidin interferes with navigation of the solitary bee Osmia cornuta in a laboratory test.

Pollinating insects provide a vital ecosystem service to crops and wild plants. Exposure to low doses of neonicotinoid insecticides has sub-lethal effects on social pollinators such as bumblebees and honeybees, disturbing their navigation and interfering with their development. Solitary Hymenoptera are also very important ecosystem service providers, but the sub-lethal effects of neonicotinoids have not yet been studied well in those animals. We analyzed the ability of walking Osmia to remember a feeding place in a small environment and found that Osmia remembers the feeding place well after 4 days of training. Uptake of field-realistic amounts of the neonicotinoid clothianidin (0.76 ng per bee) altered the animals' sensory responses to the visual environment and interfered with the retrieval of navigational memory. We conclude that the neonicotinoid clothianidin compromises visual guidance and the use of navigational memory in the solitary bee Osmia cornuta.

Effects of sub-lethal doses of glyphosate on honeybee navigation.

Glyphosate (GLY) is a herbicide that is widely used in agriculture for weed control. Although reports about the impact of GLY in snails, crustaceans and amphibians exist, few studies have investigated its sub-lethal effects in non-target organisms such as the honeybee Apis mellifera, the main pollen vector in commercial crops. Here, we tested whether exposure to three sub-lethal concentrations of GLY (2.5, 5 and 10 mg/L corresponding to 0.125, 0.250 and 0.500 µg/animal) affects the homeward flight path of honeybees in an open field. We performed an experiment in which forager honeybees were trained to an artificial feeder, and then captured, fed with sugar solution containing GLY traces and released from a novel site (the release site, RS) either once or twice. Their homeward trajectories were tracked using harmonic radar technology. We found that honeybees that had been fed with solution containing 10 mg/L GLY spent more time performing homeward flights than control bees or bees treated with lower GLY concentrations. They also performed more indirect homing flights. Moreover, the proportion of direct homeward flights performed after a second release at the RS increased in control bees but not in treated bees. These results suggest that, in honeybees, exposure to GLY doses commonly found in agricultural settings impairs the cognitive capacities needed to retrieve and integrate spatial information for a successful return to the hive. Therefore, honeybee navigation is affected by ingesting traces of the most widely used herbicide worldwide, with potential long-term negative consequences for colony foraging success.

Effects of sublethal doses of glyphosate on honeybee navigation.

Glyphosate (GLY) is a herbicide that is widely used in agriculture for weed control. Although reports about the impact of GLY in snails, crustaceans and amphibians exist, few studies have investigated its sublethal effects in non-target organisms such as the honeybee Apis mellifera, the main pollen vector in commercial crops. Here, we tested whether exposure to three sublethal concentrations of GLY (2.5, 5 and 10 mg l(-1): corresponding to 0.125, 0.250 and 0.500 µg per animal) affects the homeward flight path of honeybees in an open field. We performed an experiment in which forager honeybees were trained to an artificial feeder, and then captured, fed with sugar solution containing traces of GLY and released from a novel site either once or twice. Their homeward trajectories were tracked using harmonic radar technology. We found that honeybees that had been fed with solution containing 10 mg l(-1) GLY spent more time performing homeward flights than control bees or bees treated with lower concentrations. They also performed more indirect homing flights. Moreover, the proportion of direct homeward flights performed after a second release from the same site increased in control bees but not in treated bees. These results suggest that, in honeybees, exposure to levels of GLY commonly found in agricultural settings impairs the cognitive capacities needed to retrieve and integrate spatial information for a successful return to the hive. Therefore, honeybee navigation is affected by ingesting traces of the most widely used herbicide worldwide, with potential long-term negative consequences for colony foraging success.

General anesthesia alters time perception by phase shifting the circadian clock.

Following general anesthesia, people are often confused about the time of day and experience sleep disruption and fatigue. It has been hypothesized that these symptoms may be caused by general anesthesia affecting the circadian clock. The circadian clock is fundamental to our well-being because it regulates almost all aspects of our daily biochemistry, physiology, and behavior. Here, we investigated the effects of the most common general anesthetic, isoflurane, on time perception and the circadian clock using the honeybee (Apis mellifera) as a model. A 6-h daytime anesthetic systematically altered the time-compensated sun compass orientation of the bees, with a mean anticlockwise shift in vanishing bearing of 87° in the Southern Hemisphere and a clockwise shift in flight direction of 58° in the Northern Hemisphere. Using the same 6-h anesthetic treatment, time-trained bees showed a delay in the start of foraging of 3.3 h, and whole-hive locomotor-activity rhythms were delayed by an average of 4.3 h. We show that these effects are all attributable to a phase delay in the core molecular clockwork. mRNA oscillations of the central clock genes cryptochrome-m and period were delayed by 4.9 and 4.3 h, respectively. However, this effect is dependent on the time of day of administration, as is common for clock effects, and nighttime anesthesia did not shift the clock. Taken together, our results suggest that general anesthesia during the day causes a persistent and marked shift of the clock effectively inducing "jet lag" and causing impaired time perception. Managing this effect in humans is likely to help expedite postoperative recovery.

Mushroom body extrinsic neurons in the honeybee brain encode cues and contexts differently.

Free-flying honeybees (Apis mellifera carnica) are known to learn the context to solve discrimination tasks. Here we apply classical conditioning of the proboscis extension response in restrained bees in combination with single-unit extracellular recordings from mushroom body (MB) extrinsic neurons elucidating the neural correlates of context-dependent olfactory discrimination. The contexts were light, colors, and temperatures, either alone or in combination. We found that bees learn context rules quickly and use them for better discrimination. They also solved a transwitching and a cue/context reversal task. Neurons extrinsic to the α lobe of the MB reduced the responses to the rewarded odor, whereas they increased their responses to the context. These results indicate that MB extrinsic neurons encode cues and contexts differently. Data are discussed with reference to MB function.