A diabetes-like biochemical and behavioural phenotype of Drosophila induced by predator stress.

Predation can have both lethal and non-lethal effects on prey. The non-lethal effects of predation can instil changes in prey life history, behaviour, morphology and physiology, causing adaptive evolution. The chronic stress caused by sustained predation on prey is comparable to chronic stress conditions in humans. Conditions like anxiety, depression, and post-traumatic stress syndrome have also been implicated in the development of metabolic disorders such as obesity and diabetes. In this study, we found that predator stress induced during larval development in fruit flies Drosophila melanogaster impairs carbohydrate metabolism by systemic inhibition of Akt protein kinase, which is a central regulator of glucose uptake. However, Drosophila grown with predators survived better under direct spider predation in the adult phase. Administration of metformin and 5-hydroxytryptophan (5-HTP), a precursor of the neurotransmitter serotonin, reversed these effects. Our results demonstrate a direct link between predator stress and metabolic impairment, suggesting that a diabetes-like biochemical phenotype may be adaptive in terms of survival and reproductive success. We provide a novel animal model to explore the mechanisms responsible for the onset of these metabolic disorders, which are highly prevalent in human populations.

Neural Code for Ambient Heat Detection in Elaterid Beetles.

Environmental thermal conditions play a major role at all levels of biological organization; however, there is little information on noxious high temperature sensation crucial in behavioral thermoregulation and survival of small ectothermic animals such as insects. So far, a capability to unambiguously encode heat has been demonstrated only for the sensory triad of the spike bursting thermo- and two bimodal hygro-thermoreceptor neurons located in the antennal dome-shaped sensilla (DSS) in a carabid beetle. We used extracellular single sensillum recording in the range of 20-45°C to demonstrate that a similar sensory triad in the elaterid Agriotes obscurus also produces high temperature-induced bursty spike trains. Several parameters of the bursts are temperature dependent, allowing the neurons in a certain order to encode different, but partly overlapping ranges of heat up to lethal levels in a graded manner. ISI in a burst is the most useful parameter out of six. Our findings consider spike bursting as a general, fundamental quality of the classical sensory triad of antennal thermo- and hygro-thermoreceptor neurons widespread in many insect groups, being a flexible and reliable mode of coding unfavorably high temperatures. The possible involvement of spike bursting in behavioral thermoregulation of the beetles is discussed. By contrast, the mean firing rate of the neurons in regular and bursty spike trains combined does not carry useful thermal information at the high end of noxious heat.

Bursty spike trains of antennal thermo- and bimodal hygro-thermoreceptor neurons encode noxious heat in elaterid beetles.

The main purpose of this study was to explain the internal fine structure of potential antennal thermo- and hygroreceptive sensilla, their innervation specifics, and responses of the sensory neurons to thermal and humidity stimuli in an elaterid beetle using focused ion beam scanning electron microscopy and electrophysiology, respectively. Several essential, high temperature induced turning points in the locomotion were determined using automated video tracking. Our results showed that the sensilla under study, morphologically, are identical to the dome-shaped sensilla (DSS) of carabids. A cold-hot neuron and two bimodal hygro-thermoreceptor neurons, the moist-hot and dry-hot neuron, innervate them. Above 25-30 °C, all the three neurons, at different threshold temperatures, switch from regular spiking to temperature dependent spike bursting. The percentage of bursty DSS neurons on the antenna increases with temperature increase suggesting that this parameter of the neurons may encode noxious heat in a graded manner. Thus, we show that besides carabid beetles, elaterids are another large group of insects with this ability. The threshold temperature of the beetles for onset of elevated locomotor activity (OELA) was lower by 11.9 °C compared to that of critical thermal maximum (39.4 °C). Total paralysis occurred at 41.8 °C. The threshold temperatures for spike bursting of the sensory neurons in DSS and OELA of the beetles coincide suggesting that probably the spike bursts are responsible for encoding noxious heat when confronted. In behavioural thermoregulation, spike bursting DSS neurons serve as a fast and firm three-fold early warning system for the beetles to avoid overheating and death.

Behavioural effects of the neonicotinoid insecticide thiamethoxam on the predatory insect Platynus assimilis.

Little information is available regarding sublethal effects of neonicotinoids on insect predators, many of which perform important roles in ecosystem functioning and biocontrol. In this study, dose-dependent sublethal effects of a dietary administered neonicotinoid insecticide thiamethoxam on two basic behaviours, locomotion and feeding, were quantified in the carabid Platynus assimilis (Coleoptera, Carabidae) using automated video-tracking and weighing of consumed food, respectively. Acute toxicity tests showed that, when orally administered, the LD50 of thiamethoxam for P. assimilis beetles was 114.5 ng/g. Thiamethoxam at 108.1 ng/g caused a short-term locomotor hyperactivity within several hours of treatment. Next day after exposure to the insecticide, all the beetles were in a state of locomotor hypoactivity independent of the administered dose ranging from 1.1 to 108.1 ng/g. Reduction in clean food consumption rate (CFCR) is another altered behavioural endpoint of poisoned insect predators as first demonstrated in this study. On the first day of thiamethoxam administration, a remarkable reduction in feeding only occurred in beetles treated at 108.1 ng/g but on the next day, this negative effect appeared even at doses ten to a hundred-fold lower. Recovery from locomotion abnormalities and reduced feeding took several days. Both locomotor activity and CFCR are sensitive and valuable ecotoxicological biomarkers of carabids which should be taken into account in Integrated Pest Management programs where optimal combination of reduced insecticide use and biological control by predatory insects is crucial to achieve best results.

Encoding noxious heat by spike bursts of antennal bimodal hygroreceptor (dry) neurons in the carabid Pterostichus oblongopunctatus.

Despite thermosensation being crucial in effective thermoregulation behaviour, it is poorly studied in insects. Very little is known about encoding of noxious high temperatures by peripheral thermoreceptor neurons. In carabids, thermo- and hygrosensitive neurons innervate antennal dome-shaped sensilla (DSS). In this study, we demonstrate that several essential fine structural features of dendritic outer segments of the sensory neurons in the DSS and the classical model of insect thermo- and hygrosensitive sensilla differ fundamentally. Here, we show that spike bursts produced by the bimodal dry neurons in the antennal DSS may contribute to the sensation of noxious heat in P. oblongopunctatus. Our electrophysiological experiments showed that, at temperatures above 25 °C, these neurons switch from humidity-dependent regular spiking to temperature-dependent spike bursting. Five out of seven measured parameters of the bursty spike trains, the percentage of bursty dry neurons, the CV of ISIs in a spike train, the percentage of bursty spikes, the number of spikes in a burst and the ISIs in a burst, are unambiguously dependent on temperature and thus may precisely encode both noxious high steady temperatures up to 45 °C as well as rapid step-changes in it. The cold neuron starts to produce temperature-dependent spike bursts at temperatures above 30-35 °C. Thus, the two neurons encode different but largely overlapping ranges in noxious heat. The extent of dendritic branching and lamellation of the neurons largely varies in different DSS, which might be the structural basis for their variation in threshold temperatures for spike bursting.

The opening-closing rhythms of the subelytral cavity associated with gas exchange patterns in diapausing Colorado potato beetle, Leptinotarsa decemlineata.

The opening-closing rhythms of the subelytral cavity and associated gas exchange patterns were monitored in diapausing Leptinotarsa decemlineata beetles. Measurements were made by means of a flow-through CO2 analyser and a coulometric respirometer. Under the elytra of these beetles there is a more or less tightly enclosed space, the subelytral cavity (SEC). When the cavity was tightly closed, air pressure inside was sub-atmospheric, as a result of oxygen uptake into the tracheae by the beetle. In about half of the beetles, regular opening-closing rhythms of the SEC were observed visually and also recorded; these beetles displayed a discontinuous gas exchange pattern. The SEC opened at the start of the CO2 burst and was immediately closed. On opening, a rapid passive suction inflow of atmospheric air into the SEC occurred, recorded coulometrically as a sharp upward peak. As the CO2 burst lasted beyond the closure of the SEC, we suggest that most of the CO2 was expelled through the mesothoracic spiracles. In the remaining beetles, the SEC was continually semi-open, and cyclic gas exchange was exhibited. The locking mechanisms and structures between the elytra and between the elytra and the body were examined under a stereomicroscope and by means of microphotography. We conclude that at least some of the L. decemlineata diapausing beetles were able to close their subelytral cavity tightly, and that the cavity then served as a water-saving device.

Responses of the antennal bimodal hygroreceptor neurons to innocuous and noxious high temperatures in the carabid beetle, Pterostichus oblongopunctatus.

Electrophysiological responses of thermo- and hygroreceptor neurons from antennal dome-shaped sensilla of the carabid beetle Pterostichus oblongopunctatus to different levels of steady temperature ranging from 20 to 35°C and rapid step-changes in it were measured and analysed at both constant relative and absolute ambient air humidity conditions. It appeared that both hygroreceptor neurons respond to temperature which means that they are bimodal. For the first time in arthropods, the ability of antennal dry and moist neurons to produce high temperature induced spike bursts is documented. Burstiness of the spike trains is temperature dependent and increases with temperature increase. Threshold temperatures at which the two neurons switch from regular spiking to spike bursting are lower compared to that of the cold neuron, differ and approximately coincide with the upper limit of preferred temperatures of the species. We emphasise that, in contrast to various sensory systems studied, the hygroreceptor neurons of P. oblongopunctatus have stable and continuous burst trains, no temporal information is encoded in the timing of the bursts. We hypothesise that temperature dependent spike bursts produced by the antennal thermo- and hygroreceptor neurons may be responsible for detection of noxious high temperatures important in behavioural thermoregulation of carabid beetles.

Low doses of the common alpha-cypermethrin insecticide affect behavioural thermoregulation of the non-targeted beneficial carabid beetle Platynus assimilis (Coleoptera: Carabidae).

Sub-lethal effects of pesticides on behavioural endpoints are poorly investigated in non-targeted beneficial carabids. Conspicuous changes in locomotor activity of carabids exposed to sub-lethal doses of neurotoxic insecticides suggest that many other behaviours of these insects might be severely injured as well. We hypothesize that behavioural thermoregulation of carabids may be affected by low doses of neurotoxic pyrethroid insecticide alpha-cypermethrin which may have direct deleterious consequences for the fitness and populations of the beetles in the field. Automated video tracking of the carabid beetle Platynus assimilis Paykull (Coleoptera: Carabidae) on an experimental thermal mosaic arena using EthoVision XT Version 9 software (Noldus Information Technology, Wageningen, The Netherlands) showed that brief exposure to alpha-cypermethrin at sub-lethal concentrations (0.1-10mgL(-1)) drastically reduces the ability of the beetles for behavioural thermoregulation. At noxious high temperature, a considerable number of the beetles died due to thermo-shock. Other intoxicated beetles that survived exposure to high temperature displayed behavioural abnormalities. During heating of the arena from 25 to 45°C, insecticide treated beetles showed a significant fall in tendency to hide in a cool shelter (20°C) and prolonged exposure to noxious high temperatures, accompanied by changes in locomotor activity. Next day after insecticide treatment the beetles recovered from behavioural abnormalities to a large extent but they still were considerably longer exposed to noxious high temperatures compared to the negative control beetles. Our results demonstrated that behavioural thermoregulation is a sensitive and important etho-toxicological biomarker in ground-dwelling carabids. Prolonged exposure to unfavourably high temperatures has an array of negative effects decreasing fitness and survival of these insects at elevated thermal conditions with deep temperature gradients, typical of agricultural habitats. These results may have importance in IPM programs promoting reduced insecticide use.

Sub-lethal effects of the neurotoxic pyrethroid insecticide Fastac 50EC on the general motor and locomotor activities of the non-targeted beneficial carabid beetle Platynus assimilis (Coleoptera: Carabidae).

BACKGROUND: Sub-lethal effects of pesticides on behavioural endpoints are poorly studied in carabids (Coleoptera: Carabidae) though changes in behaviour caused by chemical stress may affect populations of these non-targeted beneficial insects. General motor activity and locomotion are inherent in many behavioural patterns, and changes in these activities that result from xenobiotic influence mirror an integrated response of the insect to pesticides. Influence of pyrethroid insecticides over a wide range of sub-lethal doses on the motor activities of carabids still remains unclear. RESULTS: Video tracking of Platynus assimilis showed that brief exposure to alpha-cypermethrin at sub-lethal concentrations ranged from 0.01 to 100 mg L(-1) caused initial short-term (< 2 h) locomotor hyperactivity followed by a long-term (>24 h) locomotor hypo-activity. In addition, significant short- and long-term concentration and time-dependent changes occurred in general motor activity patterns and rates. CONCLUSION: Conspicuous changes in motor activity of Platynus assimilis beetles treated at alpha-cypermethrin concentrations up to 75,000-fold lower than maximum field recommended concentration (MFRC) suggest that many, basic fitness-related behaviours might be severely injured as well. These changes may negatively affect carabid populations in agro-ecosystems. Long-term hypo-activity could directly contribute to decreased trap captures of carabids frequently observed after insecticide application in the field.

Electrophysiological identification of hygroreceptor neurons from the antennal dome-shaped sensilla in the ground beetle Pterostichus oblongopunctatus.

This study gives the first electrophysiological evidence of hygroreceptors in carabids. Extracellular recordings from the antennal dome-shaped sensilla of the carabid beetle Pterostichus oblolongopunctatus (Coleoptera, Carabidae) clearly show the presence of moist and dry neuron antagonistically responding to humidity changes. The cold neuron of the same sensillum did not respond to changes in humidity. For the first time, we demonstrate that the binary system of two antagonistic hygroreceptor neurons discriminates differences between steady-state humidity levels more sensitively than either neuron separately. Another advantage of the binary system is that it guarantees immediate and strong phasic-tonic response to rapid humidity changes in either direction. In the hygrosensing system of carabids, this would allow detection of subtle step-changes in humidity with greater sensitivity than differences in steady-state values of humidity. Thus, construction of the hygrosensing system with opposing receptor neurons may allow insects to detect environmental humidity differences critical for their habitat and microhabitat selection, and survival with great precision.

Spike bursts generated by the thermosensitive (cold) neuron from the antennal campaniform sensilla of the ground beetle Platynus assimilis.

Responses of the antennal thermosensitive neuron of the ground beetle Platynus assimilis to warming from 20 to 50 degrees C were measured and analysed. During warming, neurons switched from regular spiking to bursting. ISI analysis showed that the number of spikes in the burst and spike frequency within the burst were temperature dependent and may precisely encode unfavourably or dangerously high temperatures in a graded manner. In contrast, regular spikes of the neuron encode moderate temperatures at 20-30 degrees C. The threshold temperature of spike bursting varied in different neurons from 25 to 47 degrees C. As a result, the number of bursting neurons increased with temperature increase. Therefore, in addition to the burst characteristics, the total number of bursting neurons may also contain useful information on external temperature. A relationship between the spike bursts and locomotor activity of the beetles was found which may have importance in behavioural thermoregulation of the species. At 44.4+/-0.6 degrees C, first indications of partial paralysis (of the hind legs) were observed. We emphasize, that in contrast to various sensory systems studied, the thermoreceptor neuron of P. assimilis has a stable and continuous burst train, no temporal information is encoded in the timing of the bursts.

Electrophysiological responses from neurons of antennal taste sensilla in the polyphagous predatory ground beetle Pterostichus oblongopunctatus (Fabricius 1787) to plant sugars and amino acids.

The responses of antennal contact chemoreceptors, in the polyphagous predatory ground beetle Pterostichus oblongopunctatus, to twelve 1-1,000 mmol l(-1) plant sugars and seven 10-100 mmol l(-1) amino acids were tested. The disaccharides with an alpha-1.4-glycoside linkage, sucrose and maltose, were the two most stimulatory sugars for the sugar-sensitive neuron innervating these contact chemosensilla. The firing rates they evoked were concentration dependent and reached up to 70 impulses/s at 1,000 mmol l(-1). The stimulatory effect of glucose on this neuron was approximately two times lower. This can be partly explained by the fact that glucose exists in at least two anomeric forms, alpha and beta. These two forms interconvert over a timescale of hours in aqueous solution, to a final stable ratio of alpha:beta 36:64, in a process called mutarotation. So the physiologically active alpha-anomere forms only 36% of the glucose solution which was reflected in its relatively low dose/response curve. Due to the partial herbivory of P. oblongopunctatus these plant sugars are probably involved in its search for food, for example, for conifer seeds. Several carbohydrates, in addition to glucose, such as cellobiose, arabinose, xylose, mannose, rhamnose and galactose are known as components of cellulose and hemicelluloses. They are released by brown-rot fungi during enzymatic wood decay. None of them stimulated the antennal sugar-sensitive neuron. They are therefore not implicated in the search for hibernation sites, which include rotting wood, by this beetle. The weak stimulating effect (below 3 impulses/s) of some 100 mmol l(-1) amino acids (methionine, serine, alanine, glutamine) to the 4th chemosensory neuron of these sensilla was characterized as non-specific, or modulating the responses of non-target chemosensory neurons.

Electrophysiological identification of the sugar cell in antennal taste sensilla of the predatory ground beetle Pterostichus aethiops.

By single sensillum tip recording technique, in addition to the salt and pH cells found in antennal taste sensilla of some ground beetles earlier, the third chemosensory cell of four innervating these large sensilla was electrophysiologically identified as a sugar cell in the ground beetle Pterostichus aethiops. This cell generated action potentials of considerably smaller amplitude than those of the salt and pH cells, and phasic-tonically responded to sucrose and glucose over the range of 1-1000 mM tested. Responses were concentration dependent, with sucrose generating more spikes than glucose. During the first second of the response, maximum rates of firing of the sugar cell reached up to 19 and 37 imp/s when stimulated with 1000 mM glucose and sucrose, respectively. Three to four seconds later, the responses decreased close to zero. Both sugars are important in plant carbohydrate metabolism. These ground dwelling insects may come into contact with live and decayed plant material everywhere in their habitat including their preferred overwintering sites in brown-rot decayed wood. In conclusion, we hypothesize that high content of soluble sugars in their overwintering sites and refugia is unfavourable for these ground beetles, most probably to avoid contact with dangerous fungi.

A new method for electrophysiological identification of antennal pH receptor cells in ground beetles: the example of Pterostichus aethiops (Panzer, 1796) (Coleoptera, Carabidae).

The responses of antennal taste sensilla of the ground beetle Pterostichus aethiops to 100mM Na(+)-salts and their mixtures with 1 and 10mM NaOH were compared. An increase in pH by 0.3-0.6 units in 100mM Na(+)-salt solutions, caused by the content of 1mM NaOH, was too small, except for alkaline Na(2)HPO(4), to influence the firing rate of the cation cell and pH cell significantly. However, different sensitivity of the two cells to increased pH was clearly demonstrated when the concentration of NaOH in 100mM stimulating salt solutions was increased to 10mM. Increasing pH by 1.2-2 units caused the 1st s firing rate to increase by 140-1050% and 0-26% in the pH cell and cation cell, respectively. Compared to the buffer series method used for identification of the pH receptors in ground beetles earlier, considerably stronger responses of the pH cell to a similar increase in pH were observed when the NaOH method was used for testing. At the same time, undesirable changes in salt ions concentration that occur when stimulating solutions differing by 1-2 pH units are prepared were much smaller using the latter method. Behavioural and ecological relevance of the results is discussed.

Responses of antennal campaniform sensilla to rapid temperature changes in ground beetles of the tribe platynini with different habitat preferences and daily activity rhythms.

Responses of temperature sensitive (cold) cells from the antenna of ground beetles (tribe Platynini) were compared in species with different ecological preferences and daily activity rhythms. Action potential rates were characterized at various temperatures (ranges 23-39 degrees C) and during rapid changes in it (Deltat=0.5-15 degrees C). The stationary firing frequencies were nearly twice as high in eurythermic open field ground beetles Agonum muelleri and Anchomenus dorsalis (firing rates ranging from 22 to 47imp/s) than in a stenothermic forest species Platynus assimilis. In the eurythermic species, the firing rate did not significantly depend on temperature (Anchomenus dorsalis range of 23-27 degrees C and Agonum muelleri range of 23-33 degrees C) but plots of firing rate versus temperature showed rapid declines when lethally high temperatures were approached. In contrast, a nearly linear decline of the firing rate/temperature curve was observed in Platynus assimilis. Responses to rapid temperature decreases were also considerably higher in eurythermic species. Both the peak frequency of the initial burst (maximum 420-650Hz) as well as the sustained discharge in the first 4s of the response were higher than in Platynus assimilis. Long silent periods, lasting up to several seconds, that occurred at the beginning of the response to rapid warming were significantly shorter in Agonum muelleri and Anchomenus dorsalis compared to Platynus assimilis. These findings suggest that the responses of thermoreceptors to temperature changes may be correlated with specific ecological preferences.

Electrophysiological responses to salts from antennal chaetoid taste sensilla of the ground beetle Pterostichus aethiops.

Antennal gustatory sensilla of the ground beetle Pterostichus aethiops (Pz., 1797) (Coleoptera, Carabidae) respond to salts, the three sensory cells, A-, B- and C-cells, producing action potentials that are distinguished by differences in their shape, amplitude, duration and polarity of spikes. The B-cell (salt cell) was highly sensitive to both ionic composition and concentration of the tested nine salt solutions showing phasic-tonic type of reaction with a pronounced phasic component. The stimulating effect was dominated by the cations involved, and in most cases, monovalent cations were more effective stimuli than divalent cations. Salt concentration/response relations were tested with NaCl at 1, 10, 100 and 1000 mmol l(-1): mean firing rates increased from 0.8 to 44 spikes per first second of the response, respectively. The pH value of the stimulating solutions also influenced the B-cell rate of firing. By contrast, the pH level of stimulus solutions influenced the A-cells' phasic-tonic response more than the ionic composition or concentration of these solutions. Compared to a standard 100 mmol l(-1) salt (NaCl) solution (pH 6.3), alkaline solutions of the salts NaCH3COO, Na2HPO4 and Na2B4O7 (pH 7.9, 8.5 and 9.3, respectively, all 100 mmol l(-1)) induced remarkably stronger responses in the A-cell. On the other hand, the reaction to an acid solution of NaH2PO4 (pH 4.5, 100 mmol l(-1)) was minimal. A-cell responses to neutral salts like NaCl, KCl, CaCl2, MgCl2 and C5H14NOCl (pH 6.1-6.5) varied largely in strength. Very low or no responses were observed with chlorides of divalent cations, CaCl2 and MgCl2, and choline chloride (C5H14NOCl), indicating that the ionic composition of the solutions also affected A-cell responses. Neural activity of the C-cell was not influenced by the salt solutions tested.

Antennal sensilla of the ground beetle Bembidion properans Steph. (Coleoptera, Carabidae).

The arrangement of antennal sensilla was studied in female and male ground beetles Bembidion properans Steph. (Coleoptera, Carabidae) using scanning electron microscopy. The filiform antennae, 1.8-1.9 mm in length, consist of the scape, pedicel and nine flagellomeres. In both sexes, three types of sensilla chaetica, two types of sensilla trichodea, six types of sensilla basiconica, one type of sensilla coeloconica and one type of sensilla campaniformia were distinguished. The possible function of the sensilla is discussed and three types of sensilla are considered olfactory, sensilla trichodea type 2 and sensilla basiconica types 1 and 2. Olfactory sensilla form dorsal and/or ventral sensillar fields on the flagellomeres and occur sparsely or not at all outside these areas. No sexual differences in the types, number and distribution of antennal sensilla were found.