Martin Kollmann

Discovery of a Novel Insect Neuropeptide Signaling System Closely Related to the Insect Adipokinetic Hormone and Corazonin Hormonal Systems

Neuropeptides and their G protein-coupled receptors (GPCRs) play a central role in the physiology of insects. One large family of insect neuropeptides are the adipokinetic hormones (AKHs), which mobilize lipids and carbohydrates from the insect fat body. Other peptides are the corazonins that are structurally related to the AKHs but represent a different neuropeptide signaling system. We have previously cloned an orphan GPCR from the malaria mosquito Anopheles gambiae that was structurally intermediate between the A. gambiae AKH and corazonin GPCRs. Using functional expression of the receptor in cells in cell culture, we have now identified the ligand for this orphan receptor as being pQVTFSRDWNAamide, a neuropeptide that is structurally intermediate between AKH and corazonin and that we therefore named ACP (AKH/corazonin-related peptide). ACP does not activate the A. gambiae AKH and corazonin receptors and, vice versa, AKH and corazonin do not activate the ACP receptor, showing that the ACP/receptor couple is an independent and so far unknown peptidergic signaling system. Because ACP is structurally intermediate between AKH and corazonin and the ACP receptor between the AKH and corazonin receptors, this is a prominent example of receptor/ligand co-evolution, probably originating from receptor and ligand gene duplications followed by mutations and evolutionary selection, thereby yielding three independent hormonal systems. The ACP signaling system occurs in the mosquitoes A. gambiae, Aedes aegypti, and Culex pipiens (Diptera), the silkworm Bombyx mori (Lepidoptera), the red flour beetle Tribolium castaneum (Coleoptera), the parasitic wasp Nasonia vitripennis (Hymenoptera), and the bug Rhodnius prolixus (Hemiptera). However, the ACP system is not present in 12 Drosophila species (Diptera), the honeybee Apis mellifera (Hymenoptera), the pea aphid Acyrthosiphon pisum (Hemiptera), the body louse Pediculus humanus (Phthiraptera), and the crustacean Daphnia pulex, indicating that it has been lost several times during arthropod evolution. In particular, this frequent loss of hormonal systems is unique for arthropods compared with vertebrates.

Confocal Laser Scanning Microscopy Method for Quantitative Characterization of Silica Monolith Morphology

We present a fast, nondestructive, and quantitative approach to characterize the morphology of capillary silica-based monolithic columns by reconstruction from confocal laser scanning microscopy images. The method comprises column pretreatment, image acquisition, image processing, and statistical analysis of the image data. The received morphological data are chord length distributions for the bulk macropore space and skeleton of the silica monolith. The morphological information is shown to be comparable to that derived from transmission electron microscopy, but far easier to access. The approach is generally applicable to silica-based capillary columns, monolithic or particulate. It allows the rapid acquisition of hundreds of longitudinal and cross-sectional images in a single session, resolving a multitude of morphological details in the column.

Cockchafer Larvae Smell Host Root Scents in Soil

In many insect species olfaction is a key sensory modality. However, examination of the chemical ecology of insects has focussed up to now on insects living above ground. Evidence for behavioral responses to chemical cues in the soil other than CO2 is scarce and the role played by olfaction in the process of finding host roots below ground is not yet understood. The question of whether soil-dwelling beetle larvae can smell their host plant roots has been under debate, but proof is as yet lacking that olfactory perception of volatile compounds released by damaged host plants, as is known for insects living above ground, occurs. Here we show that soil-dwelling larvae of Melolontha hippocastani are well equipped for olfactory perception and respond electrophysiologically and behaviorally to volatiles released by damaged host-plant roots. An olfactory apparatus consisting of pore plates at the antennae and about 70 glomeruli as primary olfactory processing units indicates a highly developed olfactory system. Damage induced host plant volatiles released by oak roots such as eucalyptol and anisol are detected by larval antennae down to 5 ppbv in soil air and elicit directed movement of the larvae in natural soil towards the odor source. Our results demonstrate that plant-root volatiles are likely to be perceived by the larval olfactory system and to guide soil-dwelling white grubs through the dark below ground to their host plants. Thus, to find below-ground host plants cockchafer larvae employ mechanisms that are similar to those employed by the adult beetles flying above ground, despite strikingly different physicochemical conditions in the soil.

Brain organization in Collembola (springtails)

Arthropoda is comprised of four major taxa: Hexapoda, Crustacea, Myriapoda and Chelicerata. Although this classification is widely accepted, there is still some debate about the internal relationships of these groups. In particular, the phylogenetic position of Collembola remains enigmatic. Some molecular studies place Collembola into a close relationship to Protura and Diplura within the monophyletic Hexapoda, but this placement is not universally accepted, as Collembola is also regarded as either the sister group to Branchiopoda (a crustacean taxon) or to Pancrustacea (crustaceans + hexapods). To contribute to the current debate on the phylogenetic position of Collembola, we examined the brains in three collembolan species: Folsomia candida, Protaphorura armata and Tetrodontophora bielanensis, using antennal backfills, series of semi-thin sections, and immunostaining technique with several antisera, in conjunction with confocal laser scanning microscopy and three-dimensional reconstructions. We identified several neuroanatomical structures in the collembolan brain, including a fan-shaped central body showing a columnar organization, a protocerebral bridge, one pair of antennal lobes with 20-30 spheroidal glomeruli each, and a structure, which we interpret as a simply organized mushroom body. The results of our neuroanatomical study are consistent with the phylogenetic position of Collembola within the Hexapoda and do not contradict the hypothesis of a close relationship of Collembola, Protura and Diplura.

Morphological and Transcriptomic Analysis of a Beetle Chemosensory System Reveals a Gnathal Olfactory Center

BackgroundThe red flour beetle Tribolium castaneum is an emerging insect model organism representing the largest insect order, Coleoptera, which encompasses several serious agricultural and forest pests. Despite the ecological and economic importance of beetles, most insect olfaction studies have so far focused on dipteran, lepidopteran, or hymenopteran systems.ResultsHere, we present the first detailed morphological description of a coleopteran olfactory pathway in combination with genome-wide expression analysis of the relevant gene families involved in chemoreception. Our study revealed that besides the antennae, also the mouthparts are highly involved in olfaction and that their respective contribution is processed separately. In this beetle, olfactory sensory neurons from the mouthparts project to the lobus glomerulatus, a structure so far only characterized in hemimetabolous insects, as well as to a so far non-described unpaired glomerularly organized olfactory neuropil in the gnathal ganglion, which we term the gnathal olfactory center. The high number of functional odorant receptor genes expressed in the mouthparts also supports the importance of the maxillary and labial palps in olfaction of this beetle. Moreover, gustatory perception seems equally distributed between antenna and mouthparts, since the number of expressed gustatory receptors is similar for both organs.ConclusionsOur analysis of the T. castaneum chemosensory system confirms that olfactory and gustatory perception are not organotopically separated to the antennae and mouthparts, respectively. The identification of additional olfactory processing centers, the lobus glomerulatus and the gnathal olfactory center, is in contrast to the current picture that in holometabolous insects all olfactory inputs allegedly converge in the antennal lobe. These findings indicate that Holometabola have evolved a wider variety of solutions to chemoreception than previously assumed.

Neuropeptides in insect mushroom bodies

Owing to their experimental amenability, insect nervous systems continue to be in the foreground of investigations into information processing in - ostensibly - simple neuronal networks. Among the cerebral neuropil regions that hold a particular fascination for neurobiologists are the paired mushroom bodies, which, despite their function in other behavioral contexts, are most renowned for their role in learning and memory. The quest to understand the processes that underlie these capacities has been furthered by research focusing on unraveling neuroanatomical connections of the mushroom bodies and identifying key players that characterize the molecular machinery of mushroom body neurons. However, on a cellular level, communication between intrinsic and extrinsic mushroom body neurons still remains elusive. The present account aims to provide an overview on the repertoire of neuropeptides expressed in and utilized by mushroom body neurons. Existing data for a number of insect representatives is compiled and some open gaps in the record are filled by presenting additional original data.

Revisiting the anatomy of the central nervous system of a hemimetabolous model insect species: the pea aphid Acyrthosiphon pisum.

Aphids show a marked phenotypic plasticity, producing asexual or sexual and winged or wingless morphs depending on environmental conditions and season. We describe here the general structure of the brain of various morphs of the pea aphid Acyrthosiphon pisum. This is the first detailed anatomical study of the central nervous system of an aphid by immunocytochemistry (synapsin, serotonin, and several neuropeptides), ethyl-gallate staining, confocal laser scanning microscopy, and three-dimensional reconstructions. The study has revealed well-developed optic lobes composed of lamina, medulla, and lobula complex. Ocelli are only present in males and winged parthenogenetic females. The central complex is well-defined, with a central body divided into two parts, a protocerebral bridge, and affiliated lateral accessory lobes. The mushroom bodies are ill-defined, lacking calyces, and only being visualized by using an antiserum against the neuropeptide orcokinin. The antennal lobes contain poorly delineated glomeruli but can be clearly visualized by performing antennal backfills. On the basis of our detailed description of the brain of winged and wingless parthenogenetic A. pisum females, an anatomical map is now available that should improve our knowledge of the way that these structures are involved in the regulation of phenotypic plasticity.

Neuropeptidome of Tribolium castaneum antennal lobes and mushroom bodies

Neuropeptides are a highly diverse group of signaling molecules that affect a broad range of biological processes in insects, including development, metabolism, behavior, and reproduction. In the central nervous system, neuropeptides are usually considered to act as neuromodulators and cotransmitters that modify the effect of “classical” transmitters at the synapse. The present study analyzes the neuropeptide repertoire of higher cerebral neuropils in the brain of the red flour beetle Tribolium castaneum. We focus on two integrative neuropils of the olfactory pathway, the antennal lobes and the mushroom bodies. Using the technique of direct peptide profiling by matrix‐assisted laser desorption/ionization time‐of‐flight (MALDI‐TOF) mass spectrometry, we demonstrate that these neuropils can be characterized by their specific neuropeptide expression profiles. Complementary immunohistological analyses of selected neuropeptides revealed neuropeptide distribution patterns within the antennal lobes and the mushroom bodies. Both approaches revealed consistent differences between the neuropils, underlining that direct peptide profiling by mass spectrometry is a fast and reliable method to identify neuropeptide content. J. Comp. Neurol. 522:337–357, 2014.

The insect central complex as model for heterochronic brain development—background, concepts, and tools

The adult insect brain is composed of neuropils present in most taxa. However, the relative size, shape, and developmental timing differ between species. This diversity of adult insect brain morphology has been extensively described while the genetic mechanisms of brain development are studied predominantly in Drosophila melanogaster. However, it has remained enigmatic what cellular and genetic mechanisms underlie the evolution of neuropil diversity or heterochronic development. In this perspective paper, we propose a novel approach to study these questions. We suggest using genome editing to mark homologous neural cells in the fly D. melanogaster, the beetle Tribolium castaneum, and the Mediterranean field cricket Gryllus bimaculatus to investigate developmental differences leading to brain diversification. One interesting aspect is the heterochrony observed in central complex development. Ancestrally, the central complex is formed during embryogenesis (as in Gryllus) but in Drosophila, it arises during late larval and metamorphic stages. In Tribolium, it forms partially during embryogenesis. Finally, we present tools for brain research in Tribolium including 3D reconstruction and immunohistochemistry data of first instar brains and the generation of transgenic brain imaging lines. Further, we characterize reporter lines labeling the mushroom bodies and reflecting the expression of the neuroblast marker gene Tc-asense, respectively.

Novel antennal lobe substructures revealed in the small hive beetle Aethina tumida

The small hive beetle, Aethina tumida, is an emerging pest of social bee colonies. A. tumida shows a specialized life style for which olfaction seems to play a crucial role. To better understand the olfactory system of the beetle, we used immunohistochemistry and 3-D reconstruction to analyze brain structures, especially the paired antennal lobes (AL), which represent the first integration centers for odor information in the insect brain. The basic neuroarchitecture of the A. tumida brain compares well to the typical beetle and insect brain. In comparison to other insects, the AL are relatively large in relationship to other brain areas, suggesting that olfaction is of major importance for the beetle. The AL of both sexes contain about 70 olfactory glomeruli with no obvious size differences of the glomeruli between sexes. Similar to all other insects including beetles, immunostaining with an antiserum against serotonin revealed a large cell that projects from one AL to the contralateral AL to densely innervate all glomeruli. Immunostaining with an antiserum against tachykinin-related peptides (TKRP) revealed hitherto unknown structures in the AL. Small TKRP-immunoreactive spherical substructures are in both sexes evenly distributed within all glomeruli. The source for these immunoreactive islets is very likely a group of about 80 local AL interneurons. We offer two hypotheses on the function of such structures.