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The large conductance Ca2+-activated potassium channel (pSlo) of the cockroach Periplaneta americana: structure, localization in neurons and electrophysiology.

Voltage-activated, Ca2+-sensitive K+ channels (BK or maxi K,Ca channels) play a major role in the control of neuronal excitability. We have cloned pSlo, the BK channel alpha subunit of the cockroach Periplaneta americana. The amino acid sequence of pSlo shows 88% identity to dSlo from Drosophila. There are five alternatively spliced positions in pSlo showing differential expression in various tissues. A pSlo-specific antibody prominently stained the octopaminergic dorsal unpaired median (DUM) neurons and peptidergic midline neurons in Periplaneta abdominal ganglia. HEK293 cells expressing pSlo exhibit K+ channels of 170 pS conductance. They have a tendency for brief closures, exhibit subconductance states and show slight inward rectification. Activation kinetics and voltage dependence are controlled by cytoplasmic [Ca2+]. In contrast to dSlo, pSlo channels are sensitive to charybdotoxin and iberiotoxin. Mutagenesis at two positions (E254 and Q285) changed blocking efficacy of charybdotoxin. In contrast to pSlo expressed in HEK293 cells, native IbTx-sensitive K,Ca currents in DUM and in peptidergic neurons, exhibited rapid, partial inactivation. The fast component of the K,Ca current partly accounts for the repolarization and the early after-hyperpolarization of the action potential. By means of Ca2+-induced repolarization, BK channels may reduce the risk of Ca2+ overload in cockroach neurons. Interestingly, the neurons expressing pSlo were also found to express taurine, a messenger that is likely to limit overexcitation by an autocrine mechanism in mammalian central neurons.

Alternative Splicing↗

Sex pheromone for the brownbanded cockroach is an unusual dialkyl-substituted alpha-pyrone.

Female brownbanded cockroaches, Supella longipalpa, emit a sex pheromone that attracts males from a distance. This pheromone was isolated and identified as 5-(2,4-dimethylheptanyl)-3-methyl-2H-pyran-2-one (which we refer to as supellapyrone), and its structure was confirmed by synthesis. A racemic blend of the synthetic compound elicited behavioral and electrophysiological responses comparable to the natural pheromone across a range of doses. This compound is not only a very different type of cockroach pheromone but also makes up an additional class of natural products--namely, 3,5-dialkyl-substituted alpha-pyrones.

Journal Article↗

Cytochrome P450 MA expression in insecticide-resistant German cockroaches (Dictyoptera: Blattellidae).

Cytochrome P450 monooxygenases are detoxification enzymes commonly involved in insecticide resistance by insects. Recently, an overexpressed form of this enzyme, P450 MA, was purified from an insecticide-resistant strain of German cockroach, Blattella germanica (L.), and polyclonal antisera (anti-P450 MA) was produced. To test hypotheses that the overexpressed condition of P450 MA has evolved in > 1 geographic location and that P450 MA might be involved in insecticide resistance to specific insecticides, investigations were conducted using 4 insecticide-resistant and 1 susceptible German cockroach strains. In western blots that used anti-P450 MA antiserum as a probe, substantial differences in expression of P450 MA were observed. Strains showing the highest P450 MA expression had both the highest tolerance to the organophosphate insecticide chlorpyrifos and cytochrome P450-mediated demethylation activity. Results support the hypothesis that cytochrome P450 MA is potentially overexpressed in insecticide-resistant populations on a global scale.

Animals↗

Contamination affects the performance of insecticidal baits against German cockroaches (Dictyoptera: Blattellidae).

The effects of contamination of insecticidal bait formulations, by using mint oil and silica aerogel, were evaluated in a series of laboratory experiments against the German cockroach, Blattella germanica (L.). Bait consumption at 3 d for uncontaminated baits ranged from 0.01 g for Avert dust to 0.399 g for Combat dry bait with hydramethylnon presented in a station. LT50 values for uncontaminated gel bait deposits ranged from 0.4 d for PreEmpt containing imidacloprid to 4.1 d for Maxforce containing hydramethylnon. As a group, significantly more gel bait was consumed than solid formulations even when both formulations had similar concentrations of the same active ingredient. As a result, gel baits were significantly more toxic than solid formulations. Application of mint oil directly to bait deposits significantly decreased bait consumption and increased overall LT50 values. When bait formulation types were examined individually, there was no difference in consumption or toxicity between contaminated and uncontaminated gel formulations. Contaminated solid baits, however, had significantly greater LT50 values and less consumption than uncontaminated solid baits. Gel formulations probably absorbed the contaminants and removed them from the surface of the bait deposits resulting in normal bait consumption and toxicity. Gel and solid bait deposits, inside plastic exposure stations or alone with no station and placed onto mint oil-contaminated substrates, had significantly lower bait consumption and greater LT50 values than baits placed on uncontaminated substrates. Contamination of a baited area is more likely than contamination of just the bait deposit and therefore a more realistic test of the effects of contamination on bait performance. The importance of contamination to the performance of cockroach baits is discussed.

Animals↗

Behavioral and physiological resistance of the German cockroach to gel baits (Blattodea: Blattellidae).

A gel bait-resistant German cockroach, Blattella germanica (L.), strain Cincy was collected in Cincinnati, OH. This strain exhibited a high level of behavioral resistance to Avert (0.05% abamectin) and Maxforce FC (0.01% fipronil) gel baits. Topical application assays indicated moderate levels of physiological resistance of the Cincy strain to abamectin and fipronil. Resistance ratios (based on LD50 values from topical applications) to abamectin and fipronil were 2.5 and 8.7, respectively. The Cincy strain of had a significantly lower LD50 value to abamectin than a nonaverse field strain (Dorie) and similar LD50 values to fipronil as the Dorie strain. The aversion behavior (avoidance of gel baits) was therefore caused by food ingredients in the gel baits. The Cincy strain showed avoidance of agar containing fructose, glucose, maltose, and sucrose, which are phagostimulants to the laboratory strain. Modifications of the inert ingredients in the Maxforce FC gel bait significantly improved the efficacy against the Cincy strain. The Cincy strain produced significantly smaller oothecae and lower numbers of eggs in each egg capsule than the nonaverse Jwax and Dorie strains of cockroaches, suggesting fitness costs are associated with resistance.

Animals↗

Expression, production and excretion of Bla g 1, a major human allergen, in relation to food intake in the German cockroach, Blattella germanica.

The German cockroach, Blattella germanica (Linnaeus) (Dictyoptera: Blattellidae), produces several potent human allergens, one of which, Blattella germanica allergen 1 (Bla g 1), is produced in the midgut and excreted in faeces. We tested with descriptive and experimental approaches the hypothesis that Bla g 1 production is related to food intake in adult males and females of the German cockroach. Bla g 1 mRNA expression in the female midgut (assayed by real time quantitative polymerase chain reaction), her Bla g 1 content (assayed by enzyme-linked immunosorbent assay), and the female's faeces production and its Bla g 1 content tracked a cyclic pattern in relation to the gonadotrophic cycle. All four measures rose as food intake increased, declined before oviposition in relation to diminishing food intake, and remained low while the female carried an egg case for 20 days. After her first clutch of embryos hatched, the female resumed feeding, and faeces and Bla g 1 production increased concomitantly. Both Bla g 1 mRNA expression and Bla g 1 protein levels remained low in experimentally starved females. However, when starved females were allowed to feed, Bla g 1 production elevated and the gonadotrophic cycle resumed. Bla g 1 mRNA expression also increased six-fold in response to feeding compared to starved females. By contrast, there were no apparent cycles in the pattern of Bla g 1 production in males, reflecting their low and non-cyclic food intake. Our results therefore demonstrate that Bla g 1 production in B. germanica is modulated in relation to food intake.

Allergens↗

Dieldrin: interaction with nerve components of cockroaches.

We have evidence that the nerve components of the dieldrin-resistant German cockroach have less binding capacity for dieldrin than those of the susceptible cockroach; the highest interstrain difference was in the crudenucleus fraction. The dieldrin-nerve complexes are not extracted by many organic solvents.

Animals↗

Carboxymethyl cellulose decomposition by intestinal bacteria of cockroaches.

The amounts of carboxymethyl cellulase present in the midguts and hindguts of two species of cockroaches were determined. Numbers of colonies from the guts which digested carboxymethyl cellulose, visualized by flooding plates with cetyltrimethylammonium bromide after incubation, were counted. The numbers indicated the presence of a resident, multiplying population of predominantly anaerobic carboxymethyl cellulose-digesting bacteria in both species of cockroaches. Anaerobic and facultatively anaerobic species of bacteria were isolated, and their carboxymethylcellulase activities were assayed.

Journal Article↗

Low-threshold, slow-inactivating Na+ potentials in the cockroach giant axon.

Electrophysiological properties of the ventral giant axon in the abdominal nerve cord of the cockroach were studied by recording intracellular potentials following partial or complete block of the K+ conductance. When the K+ conductance was completely blocked by tetraethylammonium (TEA) and 3,4-diaminopyridine, the intensity of depolarizing currents necessary for eliciting the action potential was markedly decreased, and the action potential was followed by a prolonged plateau potential. During the plateau potential following the spike, the input resistance was significantly reduced. The plateau potential was not affected by changing the external Ca2+ concentration but depended on the external Na+ concentration in a manner expected from the Nernst equation and was blocked by tetrodotoxin (TTX). During the plateau potential, the Na+ conductance responsible for the spike was inactivated, whereas immediately after the plateau potential a newly evoked spike was not followed by a plateau potential, suggesting different inactivation kinetics between the spike and plateau Na+ conductances. When the K+ conductance was partially blocked by TEA alone, slow depolarizing responses were evoked at voltage levels a few millivolts more positive than the resting potential. The "threshold" for the slow potential was much lower than that for the spike potential. The slow potential produced after partial block of the K+ conductance was not affected by alterations of the external Ca2+ concentration but was blocked by TTX or in a Na+-free solution. Even in normal medium, a small TTX-sensitive depolarizing response was discernible. This response was similar in its time course and threshold to the slow potential observed after partial block of the K+ conductance. It is concluded that the cockroach giant axon has two populations of Na+ channels, which can be distinguished by differences in time course and voltage levels for activation and that the slow, low-threshold Na+-dependent potential is largely masked by delayed increases in the K+ conductance under normal conditions. It remains uncertain whether the low-threshold slow potential and the plateau potential originate from the same or different populations of Na+ channels.

4-Aminopyridine↗

Slowly inactivating outward currents in a cuticular mechanoreceptor neuron of the cockroach (Periplaneta americana).

1. Although rapid adaptation is a widespread feature of sensory receptors, its ionic basis has not been clearly established in any touch receptor, because their small sizes have severely restricted the range of experiments tat can be performed. In the cockroach tactile spine, intracellular voltage-clamp recordings are now possible. 2. The basic electrophysiological properties of the cockroach femoral tactile spine neuron were studied using discontinuous (switching) single-electrode current- and voltage-clamp recordings. A slowly inactivating voltage-sensitive K+ outward current was detected after the major inward currents were blocked with tetrodotoxin. 3. The total outward current activated in < 1 ms at voltages above 0 mV. At moderate depolarizations it did not inactivate, but at higher depolarizations an inactivation time constant of approximately 260 ms was measured. Some recordings also revealed an additional, slower inactivation time constant of approximately 2.5 s. 4. More than half of the voltage-sensitive K+ outward current could be blocked with the Ca2+ channel blockers Co2+ and Cd2+. Tetraethylammonium chloride (TEA) also reduced the amplitude of the outward current to about half of its original amplitude. The actions of both blockers were reversible and probably reflect overlapping blockades of two separate outward currents. 5. The reversal potentials of the currents that remained after block with Co2+ (-91.7 mV) or TEA (-86.8 mV) were both near the K+ equilibrium potential expected for the tactile spine neuron. The voltage dependencies of activation of the Co(2+)- and TEA-resistant currents were both well fitted by Boltzmann distributions, giving values of half maximal activation (V50) equal to -34.5 mV for the Co(2+)-resistant current and -51.3 mV for the TEA-resistant current. 6. Current-clamp recordings revealed that the TEA-sensitive K+ current was the major component of action potential repolarization but that it did not effect the frequency of action potentials evoked by steady depolarization. On the other hand, blockers of Ca(2+)-sensitive K+ currents (Cd2+, Co2+, or charybdotoxin) reduced adaptation and increased the frequency of action potentials significantly but did not effect the duration or amplitude of individual action potentials.

Animals↗

Mite and cockroach proteases activate p44/p42 MAP kinases in human lung epithelial cells.

BACKGROUND: The mechanisms underlying epithelial cell activation by indoor inhaled antigens are poorly understood. METHODS: In this study, we investigated the role of mitogen-activated protein kinases (MAPKs) in A549 epithelial cells upon exposure to antigens of house dust mite (HDMA), German cockroach (GCA), and American cockroach (ACA). RESULTS: Each of these antigens induced a significant increase in IL-8 levels compared to the medium control. Exposure of A549 cells to these antigens induced the phosphorylation of p44/42 MAPKs within 5 minutes, which reached a peak at 25 minutes later and reached baseline levels at 1 hour after exposure. PD98059, a MEK1 inhibitor, significantly decreased phosphorylation of p44/p42 MAPKs and IL-8 production. Exposure of A549 cells with antigens, which had been preincubated with different protease inhibitors, also resulted in a reduction of both MAPK phosphorylation and IL-8 production. CONCLUSION: Thus, proteolytic antigens present in HDMA, GCA and ACA activate the p44/42 MAPKs airway epithelial cells, which lead to elevated IL-8 production and initiation of the inflammatory cascade.

Journal Article↗

Temporal resolution of general odor pulses by olfactory sensory neurons in American cockroaches

Behavioral and physiological evidence indicates that insect pheromone sensory neurons are able to resolve pulses of pheromone concentration as they occur downwind from a point source, but the abilities of insect sensory neurons that are sensitive to general odors to respond to pulsatile stimuli are unknown. The temporal response characteristics of olfactory sensory neurons of female American cockroaches Periplaneta americana in response to general odors were measured using a series of short odor pulses (20&shy;400 ms). Odor pulses were delivered to olfactory sensilla in a moving airstream controlled by electromagnetic valves. The responses of sensory neurons were recorded using a tungsten electrode placed at the base of the sensillum. The temporal responses of sensory neurons followed the temporal changes in stimulus concentration, which were estimated by replacing the odorant with oil smoke and measuring the concentration of smoke passing through a light beam. Spike frequency varied with odorant concentration with surprisingly fine temporal resolution. Cockroach olfactory sensory neurons were able reliably to follow 25 ms pulses of the pure odorant 1-hexanol and 50 ms pulses of the complex odor blend coconut oil. Lower concentrations of odorants elicited responses with lower peak spike frequencies that still retained the temporal resolution of the stimulus pulses. Thus, responses of olfactory sensory neurons can reflect the fine structures of non-uniform distributions of general odorants in a turbulent odor plume as well as the average odorant concentration.

Journal Article↗

Energy absorption during running by leg muscles in a cockroach

Biologists have traditionally focused on a muscle's ability to generate power. By determining muscle length, strain and activation pattern in the cockroach Blaberus discoidalis, we discovered leg extensor muscles that operate as active dampers that only absorb energy during running. Data from running animals were compared with measurements of force and power production of isolated muscles studied over a range of stimulus conditions and muscle length changes.We studied the trochanter-femoral extensor muscles 137 and 179, homologous leg muscles of the mesothoracic and metathoracic legs, respectively. Because each of these muscles is innervated by a single excitatory motor axon, the activation pattern of the muscle could be defined precisely. Work loop studies using sinusoidal strains at 8 Hz showed these trochanter-femoral extensor muscles to be quite capable actuators, able to generate a maximum of 19-25 W kg-1 (at 25 degreesC). The optimal conditions for power output were four stimuli per cycle (interstimulus interval 11 ms), a strain of approximately 4 %, and a stimulation phase such that the onset of the stimulus burst came approximately half-way through the lengthening phase of the cycle. High-speed video analysis indicated that the actual muscle strain during running was 12 % in the mesothoracic muscles and 16 % in the metathoracic ones. Myographic recordings during running showed on average 3-4 muscle action potentials per cycle, with the timing of the action potentials such that the burst usually began shortly after the onset of shortening. Imposing upon the muscle in vitro the strain, stimulus number and stimulus phase characteristic of running generated work loops in which energy was absorbed (-25 W kg-1) rather than produced. Simulations exploring a wide parameter space revealed that the dominant parameter that determines function during running is the magnitude of strain. Strains required for the maximum power output by the trochanter-femoral extensor muscles simply do not occur during constant, average-speed running. Joint angle ranges of the coxa-trochanter-femur joint during running were 3-4 times greater than the changes necessary to produce maximum power output. None of the simulated patterns of stimulation or phase resulted in power production when strain magnitude was greater than 5 %. The trochanter-femoral extensor muscles 137/179 of a cockroach running at its preferred speed of 20 cm s-1 do not operate under conditions which maximize either power output or efficiency. In vitro measurements, however, demonstrate that these muscles absorb energy, probably to provide control of leg flexion and to aid in its reversal.

Journal Article↗

The structural basis for water exchange between the female cockroach (Blattella germanica) and her ootheca.

Female German cockroaches usually carry their oothecae until they hatch. The success of embryogenesis may be dependent on the water-balance relationship between females and their developing oothecae. Oothecae detached from females early in embryogenesis often fail to develop, especially in low-humidity environments. Experiments reported here using tritiated water have confirmed the transport of a significant amount of water to the ootheca from the female during embryogenesis; 18% of the tritiated water injected into gravid females was recovered in their oothecae after 24 h. We describe a structural basis of water absorption by the oothecae. An area located on the proximal end of the ootheca (adjacent to the 'escutcheon-shaped vaginal imprint') contains small pores that penetrate the oothecal covering to access regions of the chorion lying beneath these pores. Experiments using microparabiotic chambers to examine transport of tritiated water and water-soluble materials across the escutcheon region of the oothecal covering, along with dark-field birefringent microscopy of the chorion, support the hypothesis that this chorionic network is capable of conducting water throughout the interior of the ootheca. Furthermore, the structural arrangement and intimate association of the female vestibulum with the oothecal pore field contained in the region of the escutcheon-shaped vaginal imprint appear to provide an efficient conduit to the chorion. The overall structural relationship might be a means for maintaining water balance between females and their oothecae during embryonic development. Evidence presented here supports observations that this species represents an important link in the transition from oviparity to ovoviviparity by internalization of cockroach oothecae.

Animals↗

Novel extraction of German cockroach (Dictyoptera: Blattellidae) fecal pellets enhances efficacy of spray formulation insecticides.

Methanol extracts of German cockroach, Blattella germanica (L.), fecal pellets have limited use as pheromone attractants in the urban environment because of their unpleasant color and odor. To eliminate these characteristics, a novel aqueous extract of German cockroach feces was formulated. Fetal material was extracted with methylene chloride and mixed with water. The aqueous phase of the extract was colorless and relatively odorless compared with methanol fecal extracts. Aqueous extract was bioassayed and compared with methanol extracts for aggregation activity. The efficacy of chlorpyrifos and boric acid formulations was tested with and without the addition of fecal extracts. In tests with chlorpyrifos, the addition of the aqueous extract produced significantly greater mortality in young nymphs than either chlorpyrifos alone or chlorpyrifos + methanol extract. There were no differences in mortality between the 2 extract + boric acid treatments during the test period, and both enhanced mortality.

Animals↗

Behavioral responses and mortality in German cockroaches (Blattodea: Blattellidae) after exposure to dishwashing liquid.

Behavioral responses and mortality of the German cockroach, Blattella germanica (L.), after exposure to solutions of common household dishwashing liquid diluted in tap water without other known insecticidal active ingredients are described. Soap solutions of 0.05, 0.1, 0.25, 0.5, 1.0, 2.0, 3.0, 4.0, and 5.0% were tested on first and second instars, fifth and sixth instars, and adults. Cockroaches were treated individually or in groups. Behavioral observations after treatment included an immediate knockdown period, an awakening and a struggle period, an unresponsive period, and either death or recovery between 30 min and 18 h after treatment. Probit analysis gave an excellent fit of unresponsive rates 18 h after treatment with the various soap concentrations, and LD50 and LD99 values were calculated to be 0.4 and 3.0% soap, respectively. Adult females were the most difficult to kill, but at soap concentrations of 1% or higher, 95% or greater unresponsiveness and death occurred. Soap concentrations of 3% or greater resulted in 100% unresponsiveness after 3 min, and eventually resulted in 100% mortality within 72 h for all adults and nymphs treated. Observations on the mode and site of action for soap are consistent with the blockage of spiracles and/or tracheae that results in asphyxiation and death. Future investigations into the efficacy of soaps against other arthropod groups are warranted and may be beneficial in certain situations.

Animals↗

Ovicidal activity of noviflumuron when fed to adult German cockroaches (Dictyoptera: Blattellidae).

Ovicidal activity of the benzoylphenylurea noviflumuron was evaluated in the laboratory on three adult groups (virgin females, virgin males, and fertilized, nongravid females) of the German cockroach, Blattella germanica (L.), through ingestion of treated bait. Novifumuron caused significant ovicidal effects at concentrations ranging from 10 to 5000 ppm after 5-d feeding exposure to virgin and fertilized females. Untreated females produced little or no viable oothecae when mated with virgin males that had previously ingested bait (5-d exposure) with 1000 ppm or 5000 ppm noviflumuron. The highest tested concentration of noviflumuron (5000 ppm) caused 100% ovicidal activity through two ovarian cycles for all three adult groups. Noviflumuron seems to have broader ovicidal activity against B. germanica than reported for other benzoylphenylurea insecticides and can potentially impact cockroach populations through a combination of nymphal mortality and ovicidal activity.

Animals↗

Mechanosensory control of antennal movement by the scapal hair plate in the American cockroach.

The voluntary movement of antennae of blinded cockroaches was examined in the tethered-walking condition. An object of metal plate was presented to a tip of a single antenna in order to induce tactile orientation behavior. Horizontal movements of the antenna before and during the object presentation were analyzed both before and after ablation of a mechanosensory organ, the scapal hair plate (S-HP), at the base of antenna. The resting antennal position shifted outwardly by about 20 degrees after the S-HP ablation. Spontaneous antennal movements in ablated animals became stiff and wider ranged. The tactile object was set at two different horizontal positions, 45 degrees and 90 degrees clockwise to the head, for the right side antenna. The number of contacts in a constant test period was significantly decreased in the tests at 45 degrees after ablation. Trajectories of antennal movements before and after contacts were categorized into four patterns. In the case that an antenna made contact with the object during its abduction (outward) movement, it then passed the object outwardly or withdrew inwardly. These were termed "outward-pass (O-P)" and "outward-withdrawal (O-W)" patterns, respectively. Similarly, contacts during the adduction (inward) movement were divided into "inward-pass (I-P)" or "inward-withdrawal (I-W)" pattern. Significant effects of the S-HP ablation appeared in the tests at 90 degrees : the I-P pattern mostly disappeared and was replaced by the I-W pattern. The results strongly suggest that the S-HP has crucial roles for controlling both spontaneous and stimulated movements of the cockroach antenna.

Animals↗