Wolfgang Blenau

Distribution of serotonin (5-HT) and its receptors in the insect brain with focus on the mushroom bodies: lessons from Drosophila melanogaster and Apis mellifera.

The biogenic amine serotonin (5-hydroxytryptamine, 5-HT) plays a key role in regulating and modulating various physiological and behavioral processes in both protostomes and deuterostomes. The specific functions of serotonin are mediated by its binding to and subsequent activation of membrane receptors. The vast majority of these receptors belong to the superfamily of G-protein-coupled receptors. We report here the in vivo expression pattern of a recently characterized 5-HT(1) receptor of the honeybee Apis mellifera (Am5-HT(1A)) in the mushroom bodies. In addition, we summarize current knowledge on the distribution of serotonin and serotonin receptor subtypes in the brain and specifically in the mushroom bodies of the fruit fly Drosophila melanogaster and the honeybee. Functional studies in these two species have shown that serotonergic signaling participates in various behaviors including aggression, sleep, circadian rhythms, responses to visual stimuli, and associative learning. The molecular, pharmacological, and functional properties of identified 5-HT receptor subtypes from A. mellifera and D. melanogaster will also be summarized in this review.

Am5‐HT7: molecular and pharmacological characterization of the first serotonin receptor of the honeybee (Apis mellifera)

The biogenic amine serotonin (5‐HT) plays a key role in the regulation and modulation of many physiological and behavioural processes in both vertebrates and invertebrates. These functions are mediated through the binding of serotonin to its receptors, of which 13 subtypes have been characterized in vertebrates. We have isolated a cDNA from the honeybee Apis mellifera (Am5‐ht7) sharing high similarity to members of the 5‐HT7 receptor family. Expression of the Am5‐HT7 receptor in HEK293 cells results in an increase in basal cAMP levels, suggesting that Am5‐HT7 is expressed as a constitutively active receptor. Serotonin application to Am5‐ht7‐transfected cells elevates cyclic adenosine 3′,5′‐monophosphate (cAMP) levels in a dose‐dependent manner (EC50 = 1.1–1.8 nm). The Am5‐HT7 receptor is also activated by 5‐carboxamidotryptamine, whereas methiothepin acts as an inverse agonist. Receptor expression has been investigated by RT‐PCR, in situ hybridization, and western blotting experiments. Receptor mRNA is expressed in the perikarya of various brain neuropils, including intrinsic mushroom body neurons, and in peripheral organs. This study marks the first comprehensive characterization of a serotonin receptor in the honeybee and should facilitate further analysis of the role(s) of the receptor in mediating the various central and peripheral effects of 5‐HT.

Unique features of a global human ectoparasite identified through sequencing of the bed bug genome

The bed bug, Cimex lectularius, has re-established itself as a ubiquitous human ectoparasite throughout much of the world during the past two decades. This global resurgence is likely linked to increased international travel and commerce in addition to widespread insecticide resistance. Analyses of the C. lectularius sequenced genome (650 Mb) and 14,220 predicted protein-coding genes provide a comprehensive representation of genes that are linked to traumatic insemination, a reduced chemosensory repertoire of genes related to obligate hematophagy, host–symbiont interactions, and several mechanisms of insecticide resistance. In addition, we document the presence of multiple putative lateral gene transfer events. Genome sequencing and annotation establish a solid foundation for future research on mechanisms of insecticide resistance, human–bed bug and symbiont–bed bug associations, and unique features of bed bug biology that contribute to the unprecedented success of C. lectularius as a human ectoparasite.

The cloning, phylogenetic relationship and distribution pattern of two new putative GPCR-type octopamine receptors in the desert locust (Schistocerca gregaria).

The biogenic amine octopamine functions as a neuromodulator, neurotransmitter and neurohormone in insect nervous systems. It plays a prominent role in modulating multiple physiological and behavioural processes in invertebrates. Octopamine exerts its effects by binding to specific receptor proteins that belong to the superfamily of G protein-coupled receptors. We found two partial sequences of putative octopamine receptors in the desert locust Schistocerca gregaria (SgOctalphaR and SgOctbetaR) and investigated their transcript levels in males and females of both phases and during the transition between long-term solitarious and gregarious locusts. The transcript levels of SgOctalphaR are the highest in the central nervous system, whereas those of SgOctbetaR are the highest in the flight muscles, followed by the central nervous system. Both SgOctalphaR and SgOctbetaR show higher transcript levels in long-term gregarious locusts as compared to solitarious ones. The rise of SgOctbetaR transcript levels already appears during the first 4h of gregarisation, during which also the behavioural changes take place.

Molecular and Pharmacological Characterization of Serotonin 5-HT2α and 5-HT7 Receptors in the Salivary Glands of the Blowfly Calliphora vicina

Secretion in blowfly (Calliphora vicina) salivary glands is stimulated by the biogenic amine serotonin (5-hydroxytryptamine, 5-HT), which activates both inositol 1,4,5-trisphosphate (InsP3)/Ca2+ and cyclic adenosine 3′,5′-monophosphate (cAMP) signalling pathways in the secretory cells. In order to characterize the signal-inducing 5-HT receptors, we cloned two cDNAs (Cv5-ht2α, Cv5-ht7) that share high similarity with mammalian 5-HT2 and 5-HT7 receptor genes, respectively. RT-PCR demonstrated that both receptors are expressed in the salivary glands and brain. Stimulation of Cv5-ht2α-transfected mammalian cells with 5-HT elevates cytosolic [Ca2+] in a dose-dependent manner (EC50 = 24 nM). In Cv5-ht7-transfected cells, 5-HT produces a dose-dependent increase in [cAMP]i (EC50 = 4 nM). We studied the pharmacological profile for both receptors. Substances that appear to act as specific ligands of either Cv5-HT2α or Cv5-HT7 in the heterologous expression system were also tested in intact blowfly salivary gland preparations. We observed that 5-methoxytryptamine (100 nM) activates only the Cv5-HT2α receptor, 5-carboxamidotryptamine (300 nM) activates only the Cv5-HT7 receptor, and clozapine (1 µM) antagonizes the effects of 5-HT via Cv5-HT7 in blowfly salivary glands, providing means for the selective activation of each of the two 5-HT receptor subtypes. This study represents the first comprehensive molecular and pharmacological characterization of two 5-HT receptors in the blowfly and permits the analysis of the physiological role of these receptors, even when co-expressed in cells, and of the modes of interaction between the Ca2+- and cAMP-signalling cascades.

Plant essential oils and formamidines as insecticides/acaricides: what are the molecular targets?

The parasitic mite Varroa destructor is the main cause of the severe reduction in beekeeping during the last few decades. Therefore, efforts have been made to develop chemical treatments against the parasite. In the past, synthetic products were preferentially used to combat Varroa mites. Nowadays, mainly plant essential oils and organic acids are applied because they are safer and impose less unfavorable effects on the environment. Essential oils contain mixtures of mostly volatile and odorous terpenoid constituents. The molecular targets of these substances are tyramine and/or octopamine receptors that control and modulate vital functions ranging from metabolism to behavior. Disturbing the native function of these receptors in the mite results in deleterious effects in this parasite. This overview considers not only tyramine and octopamine receptors but also other potential targets of essential oils including ionotropic GABAA receptors, TRP type ion channels, and acetylcholinesterase.

Function and Distribution of 5-HT2 Receptors in the Honeybee (Apis mellifera)

Background Serotonin plays a pivotal role in regulating and modulating physiological and behavioral processes in both vertebrates and invertebrates. In the honeybee (Apis mellifera), serotonin has been implicated in division of labor, visual processing, and learning processes. Here, we present the cloning, heterologous expression, and detailed functional and pharmacological characterization of two honeybee 5-HT2 receptors. Methods Honeybee 5-HT2 receptor cDNAs were amplified from brain cDNA. Recombinant cell lines were established constitutively expressing receptor variants. Pharmacological properties of the receptors were investigated by Ca2+ imaging experiments. Quantitative PCR was applied to explore the expression patterns of receptor mRNAs. Results The honeybee 5-HT2 receptor class consists of two subtypes, Am5-HT2α and Am5-HT2β. Each receptor gene also gives rise to alternatively spliced mRNAs that possibly code for truncated receptors. Only activation of the full-length receptors with serotonin caused an increase in the intracellular Ca2+ concentration. The effect was mimicked by the agonists 5-methoxytryptamine and 8-OH-DPAT at low micromolar concentrations. Receptor activities were blocked by established 5-HT receptor antagonists such as clozapine, methiothepin, or mianserin. High transcript numbers were detected in exocrine glands suggesting that 5-HT2 receptors participate in secretory processes in the honeybee. Conclusions This study marks the first molecular and pharmacological characterization of two 5-HT2 receptor subtypes in the same insect species. The results presented should facilitate further attempts to unravel central and peripheral effects of serotonin mediated by these receptors.

The role of serotonin in feeding and gut contractions in the honeybee.

Graphical abstract

Large-scale monitoring of effects of clothianidin-dressed oilseed rape seeds on pollinating insects in northern Germany: residues of clothianidin in pollen, nectar and honey.

This study was part of a large-scale monitoring project to assess the possible effects of Elado® (10 g clothianidin & 2 g β-cyfluthrin/kg seed)-dressed oilseed rape seeds on different pollinators in Northern Germany. Firstly, residues of clothianidin and its active metabolites thiazolylnitroguanidine and thiazolylmethylurea were measured in nectar and pollen from Elado®-dressed (test site, T) and undressed (reference site, R) oilseed rape collected by honey bees confined within tunnel tents. Clothianidin and its metabolites could not be detected or quantified in samples from R fields. Clothianidin concentrations in samples from T fields were 1.3 ± 0.9 μg/kg and 1.7 ± 0.9 μg/kg in nectar and pollen, respectively. Secondly, pollen and nectar for residue analyses were sampled from free flying honey bees, bumble bees and mason bees, placed at six study locations each in the R and T sites at the start of oilseed rape flowering. Honey samples were analysed from all honey bee colonies at the end of oilseed rape flowering. Neither clothianidin nor its metabolites were detectable or quantifiable in R site samples. Clothianidin concentrations in samples from the T site were below the limit of quantification (LOQ, 1.0 µg/kg) in most pollen and nectar samples collected by bees and 1.4 ± 0.5 µg/kg in honey taken from honey bee colonies. In summary, the study provides reliable semi-field and field data of clothianidin residues in nectar and pollen collected by different bee species in oilseed rape fields under common agricultural conditions.

Large-scale monitoring of effects of clothianidin-dressed oilseed rape seeds on pollinating insects in Northern Germany: effects on honey bees (Apis mellifera)

Possible effects of clothianidin seed-treated oilseed rape on honey bee colonies were investigated in a large-scale monitoring project in Northern Germany, where oilseed rape usually comprises 25–33 % of the arable land. For both reference and test sites, six study locations were selected and eight honey bee hives were placed at each location. At each site, three locations were directly adjacent to oilseed rape fields and three locations were situated 400 m away from the nearest oilseed rape field. Thus, 96 hives were exposed to fully flowering oilseed rape crops. Colony sizes and weights, the amount of honey harvested, and infection with parasites and diseases were monitored between April and September 2014. The percentage of oilseed rape pollen was determined in pollen and honey samples. After oilseed rape flowering, the hives were transferred to an extensive isolated area for post-exposure monitoring. Total numbers of adult bees and brood cells showed seasonal fluctuations, and there were no significant differences between the sites. The honey, which was extracted at the end of the exposure phase, contained 62.0–83.5 % oilseed rape pollen. Varroa destructor infestation was low during most of the course of the study but increased at the end of the study due to flumethrin resistance in the mite populations. In summary, honey bee colonies foraging in clothianidin seed-treated oilseed rape did not show any detrimental symptoms as compared to colonies foraging in clothianidin-free oilseed rape. Development of colony strength, brood success as well as honey yield and pathogen infection were not significantly affected by clothianidin seed-treatment during this study.