In recent years, bumblebees have become a prominent insect model organism for a variety of biological disciplines, particularly to investigate learning behaviors as well as visual performance. Understanding these behaviors and their underlying neurobiological principles requires a clear understanding of brain anatomy. Furthermore, to be able to compare neuronal branching patterns across individuals, a common framework is required, which has led to the development of 3D standard brain atlases in most of the neurobiological insect model species. Yet, no bumblebee 3D standard brain atlas has been generated. Here we present a brain atlas for the buff-tailed bumblebee Bombus terrestris using micro-computed tomography (micro-CT) scans as a source for the raw data sets, rather than traditional confocal microscopy, to produce the first ever micro-CT-based insect brain atlas. We illustrate the advantages of the micro-CT technique, namely, identical native resolution in the three cardinal planes and 3D structure being better preserved. Our Bombus terrestris brain atlas consists of 30 neuropils reconstructed from ten individual worker bees, with micro-CT allowing us to segment neuropils completely intact, including the lamina, which is a tissue structure often damaged when dissecting for immunolabeling. Our brain atlas can serve as a platform to facilitate future neuroscience studies in bumblebees and illustrates the advantages of micro-CT for specific applications in insect neuroanatomy.

To-be-memorized information in working-memory could be protected against distracting influences by processes of functional inhibition or prioritization. Modulations of oscillations in the alpha to beta range in task-relevant sensory regions have been suggested to play an important role for both mechanisms. We adapted a Sternberg task variant to the auditory modality, with a strong or a weak distracting sound presented at a predictable time during the retention period. Using a time-generalized decoding approach, relatively decreased strength of memorized information was found prior to strong distractors, paralleled by decreased pre-distractor alpha/beta power in the left superior temporal gyrus (lSTG). Over the entire group, reduced beta power in lSTG was associated with relatively increased strength of memorized information. The extent of alpha power modulations within participants was negatively correlated with strength of memorized information. Overall, our results are compatible with a prioritization account, but point to nuanced differences between alpha and beta oscillations.

Division of labor among workers is a key feature of social insects and frequently characterized by an age‐related transition between tasks, which is accompanied by considerable structural changes in higher brain centers. Bumble bees (Bombus terrestris), in contrast, exhibit a size‐related rather than an age‐related task allocation, and thus workers may already start foraging at two days of age. We ask how this early behavioral maturation and distinct size variation are represented at the neuronal level and focused our analysis on the mushroom bodies (MBs), brain centers associated with sensory integration, learning and memory. To test for structural neuronal changes related to age, light exposure, and body size, whole‐mount brains of age‐marked workers were dissected for synapsin immunolabeling. MB calyx volumes, densities, and absolute numbers of olfactory and visual projection neuron (PN) boutons were determined by confocal laser scanning microscopy and three‐dimensional image analyses. Dark‐reared bumble bee workers showed an early age‐related volume increase in olfactory and visual calyx subcompartments together with a decrease in PN‐bouton density during the first three days of adult life. A 12:12  h light‐dark cycle did not affect structural organization of the MB calyces compared to dark‐reared individuals. MB calyx volumes and bouton numbers positively correlated with body size, whereas bouton density was lower in larger workers. We conclude that, in comparison to the closely related honey bees, neuronal maturation in bumble bees is completed at a much earlier stage, suggesting a strong correlation between neuronal maturation time and lifestyle in both species.

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Maintaining information in working memory normally happens in dynamic environments, with a multitude of distracting events. This is particularly evident in the auditory system, for example, when trying to memorize a telephone number during ongoing background noise. How relevant to-be-memorized information is protected against the adverse influence of a temporally predictable distractor was the main goal of the present study. For this purpose we adapted a Sternberg task variant established in the visual modality, with either a strong or a weak distracting sound presented at a fixed time during the retention period. Our behavioral analysis confirmed a small, albeit significant deterioration of memory performance in the strong distractor condition. We used a time-generalized decoding approach applied to magnetoencephalography (MEG) data to investigate the extent of memory probe-related information prior to the anticipated distractor onset and found a relative decrease for the strong distractor condition. This effect was paralleled by a predistractor alpha power decrease in the left superior temporal gyrus (STG), a cortical region putatively holding memory content relevant information. Based on gating frameworks of alpha oscillations, these results could be interpreted as a failed inhibition of an anticipated strong (more salient) distractor. However, in a critical analysis we found that reduced alpha power in the left STG was associated with relatively increased memory probe-related information. Our results therefore support the view of alpha power reductions in relevant sensory (here auditory) cortical areas to be a mechanism by which to-be-remembered information is prioritized during working memory retention periods.Auditory cortical alpha prioritizes to-be-memorized information

The diffraction limit of light microscopy poses a problem that is frequently faced in structural analyses of social insect brains. With the introduction of expansion microscopy (ExM), a tool became available to overcome this limitation by isotropic physical expansion of preserved specimens. Our analyses focus on synaptic microcircuits (microglomeruli, MG) in the mushroom body (MB) of social insects, high-order brain centers for sensory integration, learning, and memory. MG undergo significant structural reorganizations with age, sensory experience, and during long-term memory formation. However, the changes in subcellular architecture involved in this plasticity have only partially been accessed yet. Using the western honeybee Apis mellifera as an experimental model, we established ExM for the first time in a social insect species and applied it to investigate plasticity in synaptic microcircuits within MG of the MB calyces. Using combinations of antibody staining and neuronal tracing, we demonstrate that this technique enables quantitative and qualitative analyses of structural neuronal plasticity at high resolution in a social insect brain.

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