Insect Brain Research Articles

Identification of sex-biased and neurodevelopment genes via brain transcriptome in Ostrinia furnacalis.

Insect brains play important roles in the regulation of sex-biased behaviors such as mating and oviposition. The neural structure and function of brain differences between males and females have been identified, in which the antenna lobes (AL) showed the most discrepancy, however, the whole repertoire of the genes expressed in the brains and the molecular mechanism of neural signaling and structural development are still unclear. In this study, high-throughput transcriptome analysis of male and female brains was carried on in the Asia corn borer, Ostrinia furnacalis, and a total of 39.23 Gb data and 34,092 unigenes were obtained. Among them, 276 genes displayed sex-biased expression by DEG analysis, of which 125 genes were highly expressed in the males and 151 genes were highly expressed in the females. Besides, by homology analysis against genes that have been confirmed to be related to brain neurodevelopment, a total of 24 candidate genes were identified in O. furnacalis. In addition, to further screen the core genes that may be important for sex-biased nerve signaling and neurodevelopment, protein-protein interaction networks were constructed for the sex-biased genes and neurodevelopment genes. We identified 10 (Mhc, Mlc1, Mlc2, Prm, Mf, wupA, TpnC25D, fln, l(2)efl, and Act5C), 11 (PPO2, GNBP3, Spn77Ba, Ppn, yellow-d2, PGRP-LB, PGRP-SD, PGRP-SC2, Hml, Cg25C, and vkg) and 8 (dac, wg, hh, ci, run, Lim1, Rbp9, and Bx) core hub genes that may be related to brain neural development from male-biased, female-biased, and neurodevelopment gene groups. Our results provide a reference for further analysis of the dimorphism of male and female brain structures in agricultural pests.

Feature maps: How the insect visual system organizes information

A new study explores how a population of neurons in the insect brain responds to different features of visual scenes and discovers an unusual topographic map that organizes the information they encode.

Evolution: The arthropod brain — a saga in three parts

The arthropod brain comprises three segments and innervates three appendages. How it evolved has been a matter of intense debate. New Burgess Shale fossils preserving nervous tissue bring us closer to solving this mystery but evaluating the many hypotheses they could fit is not easy.

Descending control of nociception in insects?

Modulation of nociception allows animals to optimize chances of survival by adapting their behaviour in different contexts. In mammals, this is executed by neurons from the brain and is referred to as the descending control of nociception. Whether insects have such control, or the neural circuits allowing it, has rarely been explored. Based on behavioural, neuroscientific and molecular evidence, we argue that insects probably have descending controls for nociception. Behavioural work shows that insects can modulate nocifensive behaviour. Such modulation is at least in part controlled by the central nervous system since the information mediating such prioritization is processed by the brain. Central nervous system control of nociception is further supported by neuroanatomical and neurobiological evidence showing that the insect brain can facilitate or suppress nocifensive behaviour, and by molecular studies revealing pathways involved in the inhibition of nocifensive behaviour both peripherally and centrally. Insects lack the endogenous opioid peptides and their receptors that contribute to mammalian descending nociception controls, so we discuss likely alternative molecular mechanisms for the insect descending nociception controls. We discuss what the existence of descending control of nociception in insects may reveal about pain perception in insects and finally consider the ethical implications of these novel findings.

Myoinhibitory peptides in the central complex of the locust Schistocerca gregaria and colocalization with locustatachykinin‐related peptides

The central complex in the brain of insects provides a neural network for sensorimotor processing that is essential for spatial navigation and locomotion and plays a role in sleep control. Studies on the neurochemical architecture of the central complex have been performed especially in the fruit fly Drosophila melangoaster and the desert locust, Schistocerca gregaria. In several insect species, myoinhibitory peptides (MIPs) are involved in circadian control and sleep-wake regulation. To identify neurons that might underlie these functions, we investigated the distribution of MIPs in the central complex of the locust. In silico transcript analysis suggests the presence of eight different MIPs in the desert locust. Through immunolabeling, we identified five systems of central-complex neurons that express MIP-like peptides. Two systems constitute columnar neurons of the protocerebral bridge and the lower division of the central body, while the other three systems are columnar neurons (two systems) and tangential neurons (one system) of the upper division of the central body. The innervation pattern and cell count of two systems of columnar neurons revealed the existence of 18 instead of 16 columns of the protocerebral bridge. Immunostaining of preparations containing intracellularly stained single cells allowed us to further specify subtypes of labeled columnar neurons. Double-label experiments showed that three systems of MIP-immunostained columnar neurons are also locustatachykinin-immunoreactive. No colocalization was found with serotonin immunostaining. The data provide novel insights into the architecture of the locust central complex and suggest that MIPs play a prominent role within the central-complex network.

The Role of Central Complex Neurons in Prey Detection and Tracking in the Freely Moving Praying Mantis (Tenodera sinensis).

Complex tasks like hunting moving prey in an unpredictable environment require high levels of motor and sensory integration. An animal needs to detect and track suitable prey objects, measure their distance and orientation relative to its own position, and finally produce the correct motor output to approach and capture the prey. In the insect brain, the central complex (CX) is one target area where integration is likely to take place. In this study, we performed extracellular multi-unit recordings on the CX of freely hunting praying mantises (Tenodera sinensis). Initially, we recorded the neural activity of freely moving mantises as they hunted live prey. The recordings showed activity in cells that either reflected the mantis's own movements or the actions of a prey individual, which the mantises focused on. In the latter case, the activity increased as the prey moved and decreased when it stopped. Interestingly, cells ignored the movement of the other prey than the one to which the mantis attended. To obtain quantitative data, we generated simulated prey targets presented on an LCD screen positioned below the clear floor of the arena. The simulated target oscillated back and forth at various angles and distances. We identified populations of cells whose activity patterns were strongly linked to the appearance, movement, and relative position of the virtual prey. We refer to these as sensory responses. We also found cells whose activity preceded orientation movement toward the prey. We call these motor responses. Some cells showed both sensory and motor properties. Stimulation through tetrodes in some of the preparations could also generate similar movements. These results suggest the crucial importance of the CX to prey-capture behavior in predatory insects like the praying mantis and, hence, further emphasize its role in behaviorally and ecologically relevant contexts.

In Search for the Retrievable Memory Trace in an Insect Brain

The search strategy for the memory trace and its semantics is exemplified for the case of olfactory learning in the honeybee brain. The logic of associative learning is used to guide the experimental approach into the brain by identifying the anatomical and functional convergence sites of the conditioned stimulus and unconditioned stimulus pathways. Two of the several convergence sites are examined in detail, the antennal lobe as the first-order sensory coding area, and the input region of the mushroom body as a higher order integration center. The memory trace is identified as the pattern of associative changes on the level of synapses. The synapses are recruited, drop out, and change the transmission properties for both specifically associated stimulus and the non-associated stimulus. Several rules extracted from behavioral studies are found to be mirrored in the patterns of synaptic change. The strengths and the weaknesses of the honeybee as a model for the search for the memory trace are addressed in a comparison with Drosophila. The question is discussed whether the memory trace exists as a hidden pattern of change if it is not retrieved and whether an external reading of the content of the memory trace may ever be possible. Doubts are raised on the basis that the retrieval circuits are part of the memory trace. The concept of a memory trace existing beyond retrieval is defended by referring to two well-documented processes also in the honeybee, memory consolidation during sleep, and transfer of memory across brain areas.

A three-dimensional atlas of the honeybee central complex, associated neuropils and peptidergic layers of the central body.

The central complex (CX) in the brain of insects is a highly conserved group of midline-spanning neuropils consisting of the upper and lower division of the central body, the protocerebral bridge, and the paired noduli. These neuropils are the substrate for a number of behaviors, most prominently goal-oriented locomotion. Honeybees have been a model organism for sky-compass orientation for more than 70 years, but there is still very limited knowledge about the structure and function of their CX. To advance and facilitate research on this brain area, we created a high-resolution three-dimensional atlas of the honeybee's CX and associated neuropils, including the posterior optic tubercles, the bulbs, and the anterior optic tubercles. To this end, we developed a modified version of the iterative shape averaging technique, which allowed us to achieve high volumetric accuracy of the neuropil models. For a finer definition of spatial locations within the central body, we defined layers based on immunostaining against the neuropeptides locustatachykinin, FMRFamide, gastrin/cholecystokinin, and allatostatin and included them into the atlas by elastic registration. Our honeybee CX atlas provides a platform for future neuroanatomical work.

Olfactory Strategies in the Defensive Behaviour of Insects.

Simple SummaryInsects have several methods to protect themselves and their resources from danger. One of them is to use their sense of smell. In this review, we describe how insects use smell to detect threats and perform behaviours of ‘flight or fight’ such as avoidance, escape or attack, in order to protect themselves. We also discuss how group-living insects share the information of danger through semiochemicals called alarm pheromones, to act as a collective. In the second section of this paper, we review how these odours are processed in insect brains. We discuss how the two kinds of neural architectures observed in olfactory areas, labelled-lines and across-fiber patterns, support the processing of alarm pheromones. Finally, we give an outlook on potential future studies that will help us understand this field better. Most animals must defend themselves in order to survive. Defensive behaviour includes detecting predators or intruders, avoiding them by staying low-key or escaping or deterring them away by means of aggressive behaviour, i.e., attacking them. Responses vary across insect species, ranging from individual responses to coordinated group attacks in group-living species. Among different modalities of sensory perception, insects predominantly use the sense of smell to detect predators, intruders, and other threats. Furthermore, social insects, such as honeybees and ants, communicate about danger by means of alarm pheromones. In this review, we focus on how olfaction is put to use by insects in defensive behaviour. We review the knowledge of how chemical signals such as the alarm pheromone are processed in the insect brain. We further discuss future studies for understanding defensive behaviour and the role of olfaction.

Dual nicotinic acetylcholine receptor subunit gene knockouts reveal limits to functional redundancy

The nicotinic acetylcholine receptor (nAChR) subunit gene family consists of ten members in Drosophila melanogaster. The mature nAChR is a pentamer assembled from these subunits. Despite recent advances in the in vitro expression of some receptor subunit combinations (nAChR subtypes), the in vivo combinations and stoichiometry of these subtypes remains poorly defined. In addition, there are many potential nAChR signalling roles for different subtypes in insect behaviour, development and physiology. Prior work has shown that nAChR subunit mutants can display altered sleep and mating behaviour, disrupted hormone signalling and reduced locomotion, climbing ability and longevity. Teasing out the specific receptor subunits that are involved in these different functions is potentially made more difficult given that the structural similarity between members of gene families often means that there is a degree of functional redundancy. In order to circumvent this, we created a dual knockout strain for the Dα1 and Dβ2 nAChR subunit genes and examined four traits including insecticide resistance. These subunits had been previously implicated in the response to a neonicotinoid insecticide, imidacloprid. The use of the dual knockout revealed that Dα1 and Dβ2 subunits are involved in signalling that leads to the inflation of wings following adult emergence from the pupal case. The Dβ1 subunit had previously been implicated as a contributor to this function. The lack of a phenotype or low penetrance of the phenotype in the Dα1 and Dβ2 single mutants compared to the dual knockout suggests that these subunits are, to some extent, functionally redundant. We also observed stronger reductions in climbing ability and longevity in the dual knockout. Our findings demonstrate that a dual knockout approach to examining members of the nAChR subunit gene family may increase the power of genetic approaches linking individual subunits and combinations thereof to particular biological functions. This approach will be valuable as the nAChRs are so widely expressed in the insect brain that they are likely to have many functions that hereto remain undetected.

An atlas of the developing Tribolium castaneum brain reveals conservation in anatomy and divergence in timing to Drosophila melanogaster.

Insect brains are formed by conserved sets of neural lineages whose fibers form cohesive bundles with characteristic projection patterns. Within the brain neuropil, these bundles establish a system of fascicles constituting the macrocircuitry of the brain. The overall architecture of the neuropils and the macrocircuitry appear to be conserved. However, variation is observed, for example, in size, shape, and timing of development. Unfortunately, the developmental and genetic basis of this variation is poorly understood, although the rise of new genetically tractable model organisms such as the red flour beetle Tribolium castaneum allows the possibility to gain mechanistic insights. To facilitate such work, we present an atlas of the developing brain of T. castaneum, covering the first larval instar, the prepupal stage, and the adult, by combining wholemount immunohistochemical labeling of fiber bundles (acetylated tubulin) and neuropils (synapsin) with digital 3D reconstruction using the TrakEM2 software package. Upon comparing this anatomical dataset with the published work in Drosophila melanogaster, we confirm an overall high degree of conservation. Fiber tracts and neuropil fascicles, which can be visualized by global neuronal antibodies like antiacetylated tubulin in all invertebrate brains, create a rich anatomical framework to which individual neurons or other regions of interest can be referred to. The framework of a largely conserved pattern allowed us to describe differences between the two species with respect to parameters such as timing of neuron proliferation and maturation. These features likely reflect adaptive changes in developmental timing that govern the change from larval to adult brain.

A Protocol for Immunohistochemistry and RNA In-situ Distribution within Early Drosophila Embryo.

Calcium induced calcium release signaling (CICR) plays a critical role in many biological processes. Every cellular activity from cell proliferation and apoptosis, development and ageing, to neuronal synaptic plasticity and regeneration have been associated with Ryanodine receptors (RyRs). Despite the importance of calcium signaling, the exact mechanism of its function in early development is unclear. As an organism with a short gestational period, the embryos of Drosophila melanogaster are prime study subjects for investigating the distribution and localization of CICR associated proteins and their regulators during development. However, because of their lipid-rich embryos and chitin-rich chorion, their utility is limited by the difficulty of mounting embryos on glass surfaces. In this work, we introduce a practical protocol that significantly enhances the attachment of Drosophila embryo onto slides and detail methods for successful histochemistry, immunohistochemistry, and in-situ hybridization. The chrome alum gelatin slide-coating method and embryo pre-embedding method dramatically increases the yield in studying Drosophila embryo protein and RNA expression. To demonstrate this approach, we studied DmFKBP12/Calstabin, a well-known regulator of RyR during early embryonic development of Drosophila melanogaster. We identified DmFKBP12 in as early as the syncytial blastoderm stage and report the dynamic expression pattern of DmFKBP12 during development: initially as an evenly distributed protein in the syncytial blastoderm, then preliminarily localizing to the basement layer of the cortex during cellular blastoderm, before distributing in the primitive neuronal and digestion architecture during the three-gem layer phase in early gastrulation. This distribution may explain the critical role RyR plays in the vital organ systems that originate in from these layers: the suboesophageal and supraesophageal ganglion, ventral nervous system, and musculoskeletal system.

Categorizing Visual Information in Subpopulations of Honeybee Mushroom Body Output Neurons.

Multisensory integration plays a central role in perception, as all behaviors usually require the input of different sensory signals. For instance, for a foraging honeybee the association of a food source includes the combination of olfactory and visual cues to be categorized as a flower. Moreover, homing after successful foraging using celestial cues and the panoramic scenery may be dominated by visual cues. Hence, dependent on the context, one modality might be leading and influence the processing of other modalities. To unravel the complex neural mechanisms behind this process we studied honeybee mushroom body output neurons (MBON). MBONs represent the first processing level after olfactory-visual convergence in the honeybee brain. This was physiologically confirmed in our previous study by characterizing a subpopulation of multisensory MBONs. These neurons categorize incoming sensory inputs into olfactory, visual, and olfactory-visual information. However, in addition to multisensory units a prominent population of MBONs was sensitive to visual cues only. Therefore, we asked which visual features might be represented at this high-order integration level. Using extracellular, multi-unit recordings in combination with visual and olfactory stimulation, we separated MBONs with multisensory responses from purely visually driven MBONs. Further analysis revealed, for the first time, that visually driven MBONs of both groups encode detailed aspects within this individual modality, such as light intensity and light identity. Moreover, we show that these features are separated by different MBON subpopulations, for example by extracting information about brightness and wavelength. Most interestingly, the latter MBON population was tuned to separate UV-light from other light stimuli, which were only poorly differentiated from each other. A third MBON subpopulation was neither tuned to brightness nor to wavelength and encoded the general presence of light. Taken together, our results support the view that the mushroom body, a high-order sensory integration, learning and memory center in the insect brain, categorizes sensory information by separating different behaviorally relevant aspects of the multisensory scenery and that these categories are channeled into distinct MBON subpopulations.

Dopamine Modulation of Drosophila Ellipsoid Body Neurons, a Nod to the Mammalian Basal Ganglia.

The central complex (CX) is a neural structure located on the midline of the insect brain that has been widely studied in the last few years. Its role in navigation and goal-oriented behaviors resembles those played by the basal ganglia in mammals. However, the neural mechanisms and the neurotransmitters involved in these processes remain unclear. Here, we exploited an in vivo bioluminescence Ca2+ imaging technique to record the activity in targeted neurons of the ellipsoid body (EB). We used different drugs to evoke excitatory Ca2+-responses, depending on the putative neurotransmitter released by their presynaptic inputs, while concomitant dopamine administration was employed to modulate those excitations. By using a genetic approach to knockdown the dopamine 1-like receptors, we showed that different dopamine modulatory effects are likely due to specific receptors expressed by the targeted population of neurons. Altogether, these results provide new data concerning how dopamine modulates and shapes the response of the ellipsoid body neurons. Moreover, they provide important insights regarding the similitude with mammals as far as the role played by dopamine in increasing and stabilizing the response of goal-related information.

The neural basis of interspecies interactions in insects.

As insects move through the world, they continuously engage in behavioral interactions with other species. These interactions take on a spectrum of forms, from inconsequential encounters to predation, defense, and specialized symbiotic partnerships. All such interactions rely on sensorimotor pathways that carry out efficient categorization of different organisms and enact behaviors that cross species boundaries. Despite the universality of interspecies interactions, how insect brains perceive and process salient features of other species remains unexplored. Here, we present an overview of major questions concerning the neurobiology and evolution of behavioral interactions between species, providing a framework for future research on this critical role of the insect nervous system.

Multimodal Information Processing and Associative Learning in the Insect Brain.

Simple SummaryInsect behaviors are a great indicator of evolution and provide useful information about the complexity of organisms. The realistic sensory scene of an environment is complex and replete with multisensory inputs, making the study of sensory integration that leads to behavior highly relevant. We summarize the recent findings on multimodal sensory integration and the behaviors that originate from them in our review.The study of sensory systems in insects has a long-spanning history of almost an entire century. Olfaction, vision, and gustation are thoroughly researched in several robust insect models and new discoveries are made every day on the more elusive thermo- and mechano-sensory systems. Few specialized senses such as hygro- and magneto-reception are also identified in some insects. In light of recent advancements in the scientific investigation of insect behavior, it is not only important to study sensory modalities individually, but also as a combination of multimodal inputs. This is of particular significance, as a combinatorial approach to study sensory behaviors mimics the real-time environment of an insect with a wide spectrum of information available to it. As a fascinating field that is recently gaining new insight, multimodal integration in insects serves as a fundamental basis to understand complex insect behaviors including, but not limited to navigation, foraging, learning, and memory. In this review, we have summarized various studies that investigated sensory integration across modalities, with emphasis on three insect models (honeybees, ants and flies), their behaviors, and the corresponding neuronal underpinnings.

Identification of neuropeptide receptors from the brain of the bean pod borer, Maruca vitrata

• The brain transcriptomes of Maruca vitrata were analyzed. • Twenty neuropeptide receptors exhibited significant expression level in the brain. • The brain expressed various neuropeptide receptors belonging to GPCR family. • Neuropeptide receptors in the brain can be putative control targets. Neuropeptides in the insect brain have been involved in various physiological activities such as circadian rhythm, learning and memory, olfactory processing, feeding behavior, etc. Neuropeptides bind to their receptors which transduce their signals, resulting in behavioral and physiological changes. To identify neuropeptides and examine their diurnal expression from the brain of Maruca vitrata , the brain transcriptomes collected from total RNAs with 4 h interval were analyzed. Total trimmed reads from six brain transcriptomes were 0.69 billion and 40,487 contigs exhibiting significant FPKM value were identified. Twenty contigs showing significant expression level were identified as neuropeptide receptors including tachykinin-like peptide receptor, diuretic hormone receptor, SIFamide receptor, sex peptide receptor, somatostatin receptor, RYamide receptor, etc. These receptors mainly belonged to G protein-coupled receptor family. Relative expressions of neuropeptide receptors were evaluated based on FPKM values. Expressions of various neuropeptide receptors in the brain can provide information on putative control targets for insect pests.

Statistical Atlases and Automatic Labeling Strategies to Accelerate the Analysis of Social Insect Brain Evolution

Current methods used to quantify brain size and compartmental scaling relationships in studies of social insect brain evolution involve manual annotations of images from histological samples, confocal microscopy or other sources. This process is susceptible to human bias and error and requires time-consuming effort by expert annotators. Standardized brain atlases, constructed through 3D registration and automatic segmentation, surmount these issues while increasing throughput to robustly sample diverse morphological and behavioral phenotypes. Here we design and evaluate three strategies to construct statistical brain atlases, or templates, using ants as a model taxon. The first technique creates a template by registering multiple brains of the same species. Brain regions are manually annotated on the template, and the labels are transformed back to each individual brain to obtain an automatic annotation, or to any other brain aligned with the template. The second strategy also creates a template from multiple brain images but obtains labels as a consensus from multiple manual annotations of individual brains comprising the template. The third technique is based on a template comprising brains from multiple species and the consensus of their labels. We used volume similarity as a metric to evaluate the automatic segmentation produced by each method against the inter- and intra-individual variability of human expert annotators. We found that automatic and manual methods are equivalent in volume accuracy, making the template technique an extraordinary tool to accelerate data collection and reduce human bias in the study of the evolutionary neurobiology of ants and other insects.

Performance of polarization-sensitive neurons of the locust central complex at different degrees of polarization

The polarization pattern of the sky is exploited by many insects for spatial orientation and navigation. It derives from Rayleigh scattering in the atmosphere and depends directly on the position of the sun. In the insect brain, the central complex (CX) houses neurons tuned to the angle of polarization (AoP), that together constitute an internal compass for celestial navigation. Polarized light is not only characterized by the AoP, but also by the degree of polarization (DoP), which can be highly variable, depending on sky conditions. Under a clear sky, the DoP of polarized sky light may reach up to 0.75 but is usually much lower especially when light is scattered by clouds or haze. To investigate how the polarization-processing network of the CX copes with low DoPs, we recorded intracellularly from neurons of the locust CX at different stages of processing, while stimulating with light of different DoPs. Significant responses to polarized light occurred down to DoPs of 0.05 indicating reliable coding of the AoP even at unfavorable sky conditions. Moreover, we found that the activity of neurons at the CX input stage may be strongly influenced by nearly unpolarized light, while the activity of downstream neurons appears less affected.

Seasonal variations of serotonin in the visual system of an ant revealed by immunofluorescence and a machine learning approach.

Hibernation, as an adaptation to seasonal environmental changes in temperate or boreal regions, has profound effects on mammalian brains. Social insects of temperate regions hibernate as well, but despite abundant knowledge on structural and functional plasticity in insect brains, the question of how seasonal activity variations affect insect central nervous systems has not yet been thoroughly addressed. Here, we studied potential variations of serotonin-immunoreactivity in visual information processing centres in the brain of the long-lived ant species Lasius niger. Quantitative immunofluorescence analysis revealed stronger serotonergic signals in the lamina and medulla of the optic lobes of wild or active laboratory workers than in hibernating animals. Instead of statistical inference by testing, differentiability of seasonal serotonin-immunoreactivity was confirmed by a machine learning analysis using convolutional artificial neuronal networks (ANNs) with the digital immunofluorescence images as input information. Machine learning models revealed additional differences in the third visual processing centre, the lobula. We further investigated these results by gradient-weighted class activation mapping. We conclude that seasonal activity variations are represented in the ant brain, and that machine learning by ANNs can contribute to the discovery of such variations.