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The antennal lobe (AL) is the first olfactory center of the insect brain and is constituted of different functional units, the glomeruli. In the AL, odors are coded as spatiotemporal patterns of glomerular activity. In honeybees, olfactory learning during early adulthood modifies neural activity in the AL on a long-term scale and also enhances later memory retention. By means of behavioral experiments, we first verified that olfactory learning between the fifth and eighth day of adulthood induces better retention performances at a late adult stage than the same experience acquired before or after this period. We checked that the specificity of memory for the odorants used was improved. We then studied whether such early olfactory learning also induces long-term structural changes in the AL consistent with the formation of long-term olfactory memories. We also measured the volume of 15 identified glomeruli in the ALs of 17-day-old honeybees that either experienced an odor associated with sucrose solution between the fifth and eighth day of adulthood or were left untreated. We found that early olfactory experience induces glomerulus-selective increases in volume that were specific to the learned odor. By comparing our volumetric measures with calcium-imaging recordings from a previous study, performed in 17-day-old bees subjected to the same treatment and experimental conditions, we found that glomeruli that showed structural changes after early learning were those that exhibited a significant increase in neural activity. Our results make evident a correlation between structural and functional changes in the AL following early olfactory learning. [ABSTRACT FROM AUTHOR]

In the antennal lobe (AL), the first olfactory centre of the insect brain, odorants are represented as spatiotemporal patterns of glomerular activity. Whether and how such patterns are modified in the long term after precocious olfactory experiences (i.e. in the first days of adulthood) remains unknown. To address this question, we used in vivo optical imaging of calcium activity in the antennal lobe of 17-day-old honeybees which either experienced an odorant associated with sucrose solution 5–8 days after emergence or were left untreated. In both cases, we imaged neural responses to the learned odor and to three novel odors varying in functional group and carbon-chain length. Two different odor concentrations were used. We also measured behavioral responses of 17-day-old honeybees, treated and untreated, to these stimuli. We show that precocious olfactory experience increased general odor-induced activity and the number of activated glomeruli in the adult AL, but also affected qualitative odor representations, which appeared shifted in the neural space of treated animals relative to control animals. Such effects were not limited to the experienced odor, but were generalized to other perceptually similar odors. A similar trend was found in behavioral experiments, in which increased responses to the learned odor extended to perceptually similar odors in treated bees. Our results show that early olfactory experiences have long-lasting effects, reflected in behavioral responses to odorants and concomitant neural activity in the adult olfactory system. [ABSTRACT FROM AUTHOR]

Neonicotinoids represent over a quarter of the global pesticide market. Research on their environmental impact has revealed their adverse effect on the cognitive functions of pollinators, in particular of bees. Cognitive impairments, mostly revealed by behavioural studies, are the phenotypic expression of an alteration in the underlying neural circuits, a matter deserving greater attention. Here, we reviewed studies on the impact of field-relevant doses of neonicotinoids on the neurophysiology and neurodevelopment of bees. In particular, we focus on their olfactory system as much knowledge has been gained on the different brain areas that participate in odour processing. Recent studies have revealed the detrimental effects of neonicotinoids at multiple levels of the olfactory system, including modulation of odorant-induced activity in olfactory sensory neurons, diminished neural responses in the antennal lobe (the first olfactory processing centre) and abnormal development of the neural connectivity within the mushroom bodies (central neuropils involved in multisensory integration, learning and memory storage, among others). Given the importance of olfactory perception for multiple aspects of bee biology, the reported disruption of the olfactory circuit, which can occur even upon exposure to sublethal doses of neonicotinoids, has severe consequences at both individual and colony levels. Moreover, the effects reported for a multimodal structure such as the mushroom bodies indicate that neonicotinoids' impact translates to other sensory domains. Assessing the impact of field-relevant doses of pesticides on bee neurophysiology is crucial for understanding how neonicotinoids influence their behaviour in ecological contexts and for defining effective and sustainable agricultural practices., (© 2024 The Author(s). European Journal of Neuroscience published by Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Understanding the neural principles governing taste perception in species that bear economic importance or serve as research models for other sensory modalities constitutes a strategic goal. Such is the case of the honey bee (Apis mellifera), which is environmentally and socioeconomically important, given its crucial role as pollinator agent in agricultural landscapes and which has served as a traditional model for visual and olfactory neurosciences and for research on communication, navigation, and learning and memory. Here we review the current knowledge on honey bee gustatory receptors to provide an integrative view of peripheral taste detection in this insect, highlighting specificities and commonalities with other insect species. We describe behavioral and electrophysiological responses to several tastant categories and relate these responses, whenever possible, to known molecular receptor mechanisms. Overall, we adopted an evolutionary and comparative perspective to understand the neural principles of honey bee taste and define key questions that should be answered in future gustatory research centered on this insect., (© 2021 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

Non-elemental learning constitutes a cognitive challenge because events to be learned are usually ambiguous in terms of reinforcement outcome, contrary to elemental learning, which relies on unambiguous associations. Negative patterning (NP) constitutes a paradigmatic case of non-elemental learning, as subjects have to learn that single elements are reinforced while their simultaneous presentation is not reinforced (A+, B+ vs. AB-). Solving NP requires treating AB as being different from the linear sum of its components in order to overcome the ambiguity of stimulus reinforcement (i.e. A and B are as often reinforced as not reinforced). The honeybee is currently the only insect mastering NP as shown by studies restricted mainly to the olfactory domain. Here, we tested the bees' capacity to solve a NP discrimination in the visual domain and used to this end a virtual reality (VR) environment in which a tethered bee walking stationary on a spherical treadmill faces visual stimuli projected on a semicircular screen. We show that bees learn a composite grating made of alternated green and blue bars in an elemental way, and generalize their response to both a blue and a green grating. Yet, after NP training, one-quarter of the bees inhibited elemental processing and responded significantly more to the single-coloured gratings than to the composite grating. Alternative strategies were used by the other bees, which achieved partial NP learning. These results offer attractive perspectives to study different forms of visual learning in a controlled VR environment, and dissect their underlying mechanisms., (© 2019 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.)

We studied the involvement of the α8 subunit of nicotinic acetylcholine receptors (nAChRs) in olfactory learning and memory in Apis mellifera. We have previously shown, by injecting different nicotinic antagonists into the bee brain, that pharmacologically different subtypes of nAChRs are important for honeybee memory -α-bungarotoxin-sensitive receptors are necessary for memory consolidation and mecamylamine-sensitive receptors are involved in retrieval processes. Here, we took advantage of the honeybee genome sequencing and the development of a small interfering RNA (siRNA) tool to focus on the role of the α8 subunit, which has been shown to be expressed in brain areas important for olfactory learning, such as the antennal lobes and mushroom bodies. We first demonstrated the efficacy of the siRNA tool by showing a decrease of the α8 protein level at 6 h after brain injection of α8 siRNA. We then tested the general role of this subunit in olfactory conditioning, using brain systemic or localized siRNA injections in the antennal lobes or the calyces and vertical lobes of the mushroom bodies. These injections were performed at either 6 h before the learning acquisition or 6 h before the memory test. The most prominent result was that 6-h pre-test injection of siRNA in the mushroom body vertical lobes impaired memory retrieval at 24 and 48 h post-training. This indicated the importance of cholinergic extrinsic neurons and nAChRs containing the α8 subunit for this process., (© 2012 Federation of European Neuroscience Societies and Blackwell Publishing Ltd.)

The formation of a stable olfactory memory requires activity within several brain regions. The honeybee provides a valuable model to map complex olfactory learning tasks onto certain brain areas. To this end, we used injections of the local anaesthetic procaine to reversibly block spike activity in a specific brain region, the mushroom body (MB). We first investigated the physiological effects of procaine on cultured MB neurons from adult honeybee brains. Using the whole-cell configuration of the patch-clamp technique, we show that procaine blocks voltage-gated Na+ and K+ currents in a dose-dependent manner between 0.1 and 10 mm. The effects are reversible within a few minutes of wash. Lidocaine acts similarly, but is less effective at the tested concentrations. We then studied the role of the MBs during reversal learning by blocking the neural activity within these structures by injecting procaine. During reversal learning bees learn to revert their responses to two odorants, one rewarded (A+) and one unrewarded (B-), if their contingencies are changed (A- vs B+). Injecting procaine into the MBs impaired reversal learning. Procaine treatment during acquisition left the later retention of the initial learning (A+ vs B-) intact. Similarly, a differential conditioning task involving novel odorants (C+ vs D-) was intact under procaine treatment. Our experiments show that local injections of procaine can be used to map learning tasks onto specific regions of the insect brain. We conclude that intact MB activity is required for the acquisition of reversal learning, but not for simple differential learning tasks.

Natural olfactory stimuli occur as mixtures of many single odors. We studied whether the representation of a mixture in the brain retains single-odor information and how much mixture-specific information it includes. To understand mixture representation in the honeybee brain, we used in vivo calcium imaging at the level of the antennal lobe, and systematically measured odor-evoked activity in 24 identified glomeruli in response to four single odorants and all their possible binary, ternary and quaternary mixtures. Qualitatively, mixture-induced activity patterns always contained glomeruli belonging to the pattern of at least one of the components, suggesting a high conservation of component information in olfactory mixtures. Quantitatively, glomerular activity saturated quickly and increasing the number of components resulted in an increase of cases in which the response of a glomerulus to the mixture was lower than that to the strongest component ('suppression'). This shows global inhibition in the antennal lobe, probably acting as overall gain control. Single components were not equally salient (in terms of number of active glomeruli) and mixture activity patterns were always more similar to the more salient components, in a way that could be predicted linearly. Thus, although a gain control system in the honeybee antennal lobe prevents saturation of the olfactory system, mixture representation follows essentially elemental rules.

We combined behavioural and electrophysiological experiments to study whether bitter taste is perceived at the antennal level in honeybees, Apis mellifera. Our behavioural studies showed that neither quinine nor salicin delivered at one antenna at different concentrations induced a retraction of the proboscis once it was extended in response to 1 M sucrose solution delivered to the opposite antenna. Bees that extended massively their proboscis to 1 M sucrose responded only partially when stimulated with a mixture of 1 M sucrose and 100 mM quinine. The mixture of 1 m sucrose and 100 mM salicin had no such suppressive effect. No behavioural suppression was found for mixtures of salt solution and either bitter substance. Electrophysiological recordings of taste sensillae at the antennal tip revealed sensillae that responded specifically either to sucrose or salt solutions, but none responded to the bitter substances quinine and salicin at the different concentrations tested. The electrophysiological responses of sensillae to 15 mM sucrose solution were inhibited by a mixture of 15 mM sucrose and 0.1 mM quinine, but not by a mixture of 15 mM sucrose and 0.1 mM salicin. The responses of sensillae to 50 mM NaCl were reduced by a mixture of 50 mm NaCl and 1 mM quinine but not by a mixture of 50 mM NaCl and 1 mM salicin. We concluded that no receptor cells for the bitter substances tested, exist at the level of the antennal tip of the honeybee and that antennal bitter taste is not represented as a separate perceptual quality.

The mushroom bodies (MBs) are central structures in the insect brain that have been associated with olfactory learning and memory. Here we used hydroxyurea (HU) to treat honeybee larvae and induce partial MB ablations at the adult stage. We studied olfactory learning in honeybees with unilateral loss of the median calyces of their MBs and compared their ability to solve different forms of olfactory discrimination. When odorants were delivered in a side-specific manner, ablated bees could not solve either discrimination of the unambiguous problem (Paradigm 1: A+, B- on one antenna, C+, D- on the other; A+B-/C+D-) whereas they could solve at least one of both discriminations of the ambiguous problem (Paradigm 2: A+B-/A-B+), namely that proposed to their intact brain side. Non-ablated bees could learn side-specific discriminations on both brain sides. When odorants were delivered simultaneously to both antennae (Paradigm 3: A+B-C+D-), HU-ablated bees learned slower than HU-normal bees. Thus, in all three paradigms, the unilateral loss of a median calyx affected olfactory learning. We propose that the MBs are required for solving elemental olfactory tasks whose complexity is increased by the number of stimuli involved and that MB ablations could have an effect on the inhibition of information exchange between brain hemispheres.