Influence of pesticides and neuroactive amines on cAMP levels of two-spotted spider mite (Acari: Tetranychidae)

The interaction of several formamidine pesticides, chlordimeform (CDM), N-demethylchlordimeform (DCDM) and amitraz with octopamine receptor(s) and the resulting enhancement of cyclic-AMP (cAMP) production in vitro were investigated in the two-spotted spider mite, Tetranychus urticae Koch. DCDM and amitraz clearly stimulated the production of cAMP when added to a homogenate of the spider mite. Among various biogenic amines tested, octopamine and synephrine were most active but dopamine (DA) and 5-hydroxytroptamine showed only marginal potency to elevate cAMP production. An additivity study was devised to find whether these formamidines interact with the same target site as octopamine. The results indicate that all these chemicals act on the same receptor which functions to transduce the signal of certain biogenic amines to elevate the intracellular cAMP level. Phentolamine (PH) and propranolol (PR) showed an antagonistic effect against the portion of cAMP production which was elevated by DCDM. Among pesticides tested, deltamethrin, fenvalerate, DDT and benzenehexachloride showed no such effect, whereas dicofol, chlorobenzilate, parathion and aldicarb showed slight stimulatory effects on cAMP production. Both DCDM and octopamine cause an increase in the phosphorylation of proteins that are also phosphorylated by exogenous cAMP-dependent protein kinase. The results of pharmacological characterization studies confirmed the overall theory that the agonistic effects of formamidines are expressed primarily through the octopamine-sensitive adenylate cyclase.     

Biogenic amines modulate pulse rate in the dorsal blood vessel of Lumbriculus variegatus

The biogenic amines are widespread regulators of physiological processes, and play an important role in regulating heart rate in diverse organisms. Here, we present the first pharmacological evidence for a role of the biogenic amines in the regulation of dorsal blood vessel pulse rate in an aquatic oligochaete, Lumbriculus variegatus (Müller, 1774). Bath application of octopamine to intact worms resulted in an acceleration of pulse rate, but not when co-applied with the adenylyl cyclase inhibitor MDL-12,330a. The phosphodiesterase inhibitor theophylline mimicked the effects of OA, but the polar adenosine receptor antagonist 8(p-sulphophenyl)theophylline was significantly less potent than theophylline. Pharmacologically blocking synaptic reuptake of the biogenic amines using the selective 5-HT reuptake blocker fluoxetine or various tricyclic antidepressants also accelerated heart rate. Depletion of the biogenic amines by treatment with the monoamine vesicular transporter blocker reserpine dramatically depressed pulse rate. Pulse rate was partially restored in amine-depleted worms after treatment with octopamine or dopamine, but fully restored following treatment with serotonin. This effect of 5-HT was weakly mimicked by 5-methoxytryptamine, but not by α-methylserotonin; it was completely blocked by clozapine and partially blocked by cyproheptadine. Because they are known to orchestrate a variety of adaptive behaviors in invertebrates, the biogenic amines may coordinate blood flow with behavioral state in L. variegatus.     

Effect of various insecticides on the larval growth and biogenic amine levels of Tribolium castaneum Herbst

Various insecticides reduced larval growth of the red flour beetle (tribolium castaneum Herbst) and various biogenic amines, including octopamine (OA), dopamine (DA), serotonin (5-HT), epinephrine (E), norepinephrine (NE), their precursors and metabolites in the insects were measured by high-performance liquid chromatography coupled with electrochemical detection. Tyrosine occurred in the highest concentration followed by OA, tryptophan and 3,4-dihydroxymandelic acid (DOMA). Tyramine (a precursor of OA in the biosynthetic pathway), synephrine (N-methyl OA), DA, 5-HT, E, NE and their related substances occurred in extremely low quantities compared with OA. The insects were stressed by various insecticides, which resulted in a dramatic change of biogenic amine levels: the OA levels increased, whereas the levels of other biogenic amines and related substances decreased.     

An Inhibitor of Acetolactate Synthase from a Microbe

Such stress factors as mechanical (vibration), thermal (unfavorable temperature), optical (light), and starvation reduced the larval growth of the red flour beetle (Triholium castaneum Herbst). Various biogenic amines, including octopamine (OA), dopamine (DA), serotonin (5-HT), epinephrine (E), norepinephrine (NE), their precursors, and metabolites, in whole-body T. castaneum were measured by high-performance liquid chromatography coupled with electrochemical detection (ECD). Tyrosine occurred in the highest concentration, followed by OA, tryptophan, and 3,4-dihydroxymandelic acid. The amount of OA was much higher than that of tyramine (a precursor of OA in the biosynthetic pathway) and of synephrine (N-methyl OA). DA, 5-HT, E, NE, and their related substances occurred in extremely low quantities compared with OA. Insects were stressed by vibrating at 1, 10, 100, or 1000Hz, optically under a 24-h light (15W, 50Hz) photoperiod, thermally by changing the incubation temperature from an initial value of 30°C, or by starvation, which resulted in dramatic changes of levels of biogenic amines, including OA.     

Testing the Effects of Biogenic Amines and Alternative Topical Solvent Types on the Behavioral Repertoire of Two Web‐Building Spiders

Investigating the proximate mechanism underlying behavior has long been a primary goal of ethologists. With the recent focus on behavioral correlations (e.g., behavioral syndromes), a fundamental question arises regarding what mechanisms may drive positive relationships between functionally distinct behaviors. Understanding these mechanisms can inform critical questions, such as why distinct behaviors are often correlated, and can also lend insights into how behavioral similarities are conserved across species. Arthropods provide an idea system to study these questions as the biogenic amines, such as octopamine and serotonin, have long been known to mediate a large number of behaviors such as aggression, dominance, escape behavior, and others. Behaviorists have several ways in which they can manipulate these biogenic amines in vivo, including injection, feeding, and topical application. Topical application is potentially an optimal method for manipulating amine concentrations, given that it is minimally invasive and relatively precise. Here, we investigate the role of biogenic amines, octopamine, and serotonin, in mediating a range of similar and often correlated behaviors in two distantly related species of spiders, the western black widow, Latrodectus hesperus, and the funnel‐web spider, Agelenopsis pennsylvanica. Additionally, we investigate the behavioral effects associated with three commonly used carrier solvents: dimethylformamide (DMF), dimethyl sulfoxide (DMSO), and acetone. We demonstrate that biogenic amines are involved both in mediating several behaviors, and also potentially the links between different behaviors. We also show that the same behaviors in different species are not affected by biogenic amines in the same manner, suggesting differences in the underlying neurophysiological mechanisms governing behavior. Furthermore, we discovered large and pervasive effects associated with both carrier solvents, thus calling into question the potential usefulness and interpretability of topical application methods in behavioral research.     

In vivo stimulation of ovarian development in the red swamp crayfish,Procambarus clarkii (Girard), by 5-hydroxytryptamine

Summary

The possibility that biogenic amines affect ovarian development in the red swamp crayfish, Procambarus clarkii, was investigated. Females were administered 15 μg/g body weight (bw) of norepinephrine, dopamine, 5-hydroxytryptamine (5-HT) or octopamine on days 1, 5 and 10 and were sacrificed on day 15. Crayfish given 5-HT showed significant increases in ovarian index (30.5%) and oocyte size (34.0%) over the concurrent controls, while norepinephrine, dopamine and octopamine did not significantly affect either the ovarian index or oocyte size. Significantly more labeling by ¹⁴C-leucine of ovarian proteins was found in ovaries of crayfish that were injected with 5-HT in vivo, but when ovarian lobes from crayfish that had not been injected with 5-HT were incubated in vitro with 5-HT added to the incubation medium, no significant change in the level of incorporation of ¹⁴C-leucine into ovarian proteins occurred.

The 5-HT receptor blocker LY53857 (25 μg/g bw) retarded ovarian development. The 5-HT releaser fenfluramine and the 5-HT potentiator fluoxetine (both 15 μg/g bw) were also used. Crayfish given fenfluramine, fluoxetine, fenfluramine plus 5-HT or fluoxetine plus 5-HT showed significant increases of ovarian index (24.0–102.8%), oocyte size (20.0–87.4%) and in vitro ¹⁴C-leucine labeling of ovarian proteins (30.6–123.6%) over the concurrent controls. The ovaries of crayfish that received the 5-hydroxytryptaminergic neurotoxin 5,6-dihydroxytryptamme (10 μg/g bw) did not show any significant change as compared with the initial control. These findings are consistent with the hypothesis that 5-HT, which is present in the central nervous system of Procambarus clarkii, exerts its stimulatory effect on the ovary of this crayfish indirectly by triggering release of the ovary-stimulating hormone.     

The role of biogenic amines in the control of blood glucose level in the decapod crustacean,Carcinus maenas L.

1. Administration of biogenic amines into intact Carcinus maenas induces dose- and timedependent elevation of hemolymph glucose level.2. Removal of the neurosecretory centre containing the crustacean hyperglycemic hormone (CHH) by ablation of the eyestalks did not induce hypoglycemia.3. Injection of dopamine (DA) into eyestalkless crabs showed no hyperglycemic effect, while serotonin (5-HT), epinephrine (E), norepinephrine (NE), and octopamine (OA) elevated glucose levels.4. The dopaminergic effect was significantly reduced by administration of trifluoperazine (TFP).5. 5-HT and OA were found to be strong elevators of glucose levels, while the other biogenic amines had moderate effects only.6. The results indicate, that DA exerts its hyperglycemic effect by stimulating the release of CHH from the eyestalk neurosecretory centre. Elevation of hemolymph glucose level by 5-HT, OA, E, and NE, occurs independently of CHH.     

5-Hydroxytryptamine stimulates fluid secretion in locust malpighian tubules independently of cAMP

5-Hydroxytryptamine (5-HT) stimulates fluid secretion by semi-isolated Malpighian tubules of Locusta in a dose-dependent manner. The threshold of stimulation is between 10(-8) and 10(-7) M 5-HT; maximal activation occurs at doses greater than 10(-6) M. Relative to the activation induced by diuretic hormone (storage lobe extracts), 5-HT increases the rate of fluid secretion by only 65%. Phentolamine, the alpha-adrenergic blocker, failed to inhibit either DH or 5-HT stimulated secretion. Diuretic hormone raises the levels of intracellular of cAMP, and activates adenylate cyclase in plasma membrane preparations of Locusta Malpighian tubules. 5-HT (10(-4) M) has no effect in either assay system. Thus 5-HT can stimulate fluid secretion independently of cAMP. A hypothetical model for hormone stimulated fluid secretion by Locusta Malpighian tubules, involving dual-receptor activation, is proposed. Other biogenic amines, including octopamine, adrenalin, dopamine, synephrine and the formamidine chlordimeform were tested for their ability to stimulate fluid secretion. Only dopamine showed a weakly stimulatory effect.     

Amphetamine and Reserpine Deplete Brain Biogenic Amines and Alter Blow Fly Feeding Behavior

HPLC with electrochemical detection was used to determine the levels of p-hydroxyphenylethanolamine (octopamine), 3,4-dihydroxyphenylethylamine (dopamine), and 5-hydroxytryptamine (5-HT) in the brains of control, reserpine, and d-amphetamine-treated blow flies, Phormia regina Meigen. Parallel studies were carried out to assess the effects of the two drugs on fly feeding behavior, measured as mean acceptance threshold: the minimum sucrose concentration to which the average fly in a population will respond by proboscis extension when its tarsi contact the solution. In saline-injected control flies, all three amines were found at levels of approximately 2 pmol/brain. Thirty minutes after injection with d-amphetamine (12 micrograms/fly), brain octopamine was depleted by 85%, whereas dopamine and 5-HT were depleted by 70%. Reserpine (5 micrograms/fly) caused 70% depletion of dopamine and greater than 90% depletion of both octopamine and 5-HT 24 h after injection. However, the effect of reserpine was much slower in onset (hours versus minutes) and more persistent (days versus hours) than was the effect of d-amphetamine. With either drug, the time course of amine depletion closely matched the time course of the increase in feeding threshold observed in drug-treated flies. These results suggest that CNS pools of the biogenic amines, octopamine, dopamine, and 5-HT are important in governing blow fly responsiveness to food stimuli.     

Brain biogenic amines and reproductive dominance in bumble bees (Bombus terrestris)

To begin to explore the role of biogenic amines in reproductive division of labor in social insects, brain levels of dopamine, serotonin, and octopamine were measured in bumble bee (Bombus terrestris) workers and queens that differ in behavioral and reproductive state. Levels of all three amines were similar for mated and virgin queens. Young workers that developed with or without a queen had similar amine levels, but in queenright colonies differences in biogenic amine levels were associated with differences in behavior and reproductive physiology. Dominant workers had significantly higher octopamine levels compared with workers of lower dominance status but of similar size, age, and ovary state. High dopamine levels were associated with the last stages of oocyte development irrespective of worker social status and behavior. These results suggest that biogenic amines are involved in behavioral and physiological aspects of regulation of reproduction in bumble bees. Accepted: 10 December 1999     

Cloning and expression pattern of a putative octopamine/tyramine receptor in antennae of the noctuid moth <Emphasis Type="Italic">Mamestra brassicae</Emphasis>

In insects, biogenic amines have been shown to play an important role in olfactory plasticity. In a first attempt to decipher the underlying molecular mechanisms, we report the molecular cloning and precise expression pattern of a newly identified octopamine/tyramine-receptor-encoding gene in the antennae of the noctuid moth Mamestra brassicae (MbraOAR/TAR). A full-length cDNA has been obtained through homology cloning in combination with rapid amplification of cDNA ends/polymerase chain reaction; the deduced protein exhibits high identities with previously identified octopamine/tyramine receptors in other moths. In situ hybridization within the antennae has revealed that MbraOAR/TAR is expressed at the bases of both pheromone-sensitive and non-sensitive olfactory sensilla and in cells with a neurone-like shape. In accordance with previous physiological studies that have revealed a role of biogenic amines in the electrical activity of the receptor neurones, our results suggest that biogenic amines (either octopamine or tyramine) target olfactory receptor neurones to modulate olfactory coding as early as the antennal level.     

Developmental Changes in Biogenic Amine Levels in the Central Nervous System and the Haemolymph of the Eastern Death's Head Hawk Moth,Acherontia styx (Lepidoptera: Sphingidae)

In an effort to understand the role of biogenic amines in insect development, changes in the levels of octopamine (OA), dopamine (DA), epinephrine (E), norepinephrine (NE), and serotonin (5-HT) in the brain, the optic lobes and the haemolymph of different developmental stages of Acherontia styx were analyzed using HPLC with electrochemical detector. In the brain, OA was the most abundant monoamine. DA, OA, and E levels in larvae peaked around the wandering stage (W). A dramatic increase in DA, 5-HT, and E levels was observed in the brain of the adult as compared to the pupal stage. NE, however, was not detected in the brain of most stages of the insect, except in the brain of 20-day-old pupae and adults. A 3-fold increase in OA levels was observed in the optic lobes of the adult as compared to late pupal stage. No changes were observed for E, DA, and 5-HT. NE was not detected in the optic lobes. In the haemolymph of 5^th instar larvae, OA was also the most abundant amine. Both DA and OA peaked prior to the onset of the W stage. In contrast, E and NE concentrations decreased with development of the 5^th instar larvae. 5-HT was not detected in the haemolymph. Finally, different profiles for amine levels were observed for the two forms of the 5^th instar larvae (green vs brown). These results are interpreted in the light of the role of biogenic amines and their relation to development in the nervous system of lepidopteran insects.     

Adaptive Value of Genes Controlling the Level of Biogenic Amines in Drosophila

The effects of mutations that change the level of biogenic amines (octopamine and dopamine) on stress-reactivity and fitness of Drosophila adults are considered. It has been shown that (1) the biogenic amines represent an important but not triggering factor of the development of stress reaction; (2) under normal conditions, reproduction is regulated by genes that control dopamine metabolic pathways (indirectly via regulation of the juvenile hormone level). Under unfavorable conditions, reproduction is regulated by genes that control octopamine metabolism; (3) heat-stress adaptation depends on expression of genes controlling the background level of dopamine.     

THE UPTAKE OF BIOGENIC MONOAMINES BY THE ISOLATED AURICLE OF THE SNAIL (HELIX POMATIA)

—The uptake of [³H]5HT, [³H]dopamine, [³H]noradrenaline and [³H]octopamine into the auricle of Helix pomatia was studied. When tissues were incubated at 25°C in media containing radioactive amines, tissue:medium ratios of about 49:1, 14:1 and 5:1 for 5-HT, dopamine, noradrenaline, and octopamine respectively were obtained after a 20–30 min incubation time. Tissues incubated at 25°C in media containing radioactive amines for 20–30 mins showed that almost all (96%) the radioactivity was present as unchanged [³H]5-HT, [³H]dopamine, [³H]octopamine or [³H]noradrenaline. The high tissue:medium ratios for 5-HT and dopamine, but not for noradrenaline and octopamine, showed saturation kinetics which were dependent upon temperature and sodium ions. From the Lineweaver–Burk plots, two uptake mechanisms for 5-HT at 25°C were resolved; the high affinity uptake process having a K_m1 value of 6.0 ± 10^?8m and a V_m1 value of 0.115 nmol/g/min while the lower affinity process had a K_m2 value of 1.04 ± 10^?6m and a V_m2 value of 0.66nmol/g/min. At 0°C a single uptake mechanism for 5-HT occurred which gave a K_m value of 5.02 ± 10^?8m and a V_m value of 0.0165 nmol/g/min. In the case of dopamine, the Lineweaver–Burk plot at 25°C showed a single uptake process with values for K_m and V_m of 1.55 ± 10^?7m and 0.086 nmol/g/min respectively. This process did not function at 0°C. The effect of various agents and ions upon the accumulation processes for all amines was also studied, and the data indicate that the same neurons probably accumulate more than one amine type. It is concluded that 5-HT and dopamine uptake in the auricle is a mechanism for inactivating these substances at 25°C and that an uptake mechanism for 5-HT also functions at 0°C. The results are discussed from the point of view of 5-HT's being the cardioexcitatory substance in the snail heart.     

Hormonal Effects on the in vitro Larval Growth of the Swine Intestinal Roundworm,Ascaris suum

Summary

Several biogenic amines and insect juvenile hormone III were tested in a growth bioassay of the parasitic nematode, Ascaris suum. Compounds (1 to 1000μzmol) were placed in culture with third-stage larvae for 24 hr, larvae were then rinsed several times, and larval cultures were returned to incubators for 6 more days. By this time, larvae had developed to the fourth-stage. The larvae were fixed in hot formalin, and their lengths were measured. Epinephrine and norepinephrine oxidized and were nematocidal under these culture conditions. Histamine and serotonin had no effect on length of the larvae. Octopamine (10–50μmol) exposure resulted in a significant dose-dependent increase in length. When incubated with octopamine (10μzmol) for 7 days, larvae grew more slowly than controls without octopamine (P< 0.05). Juvenile hormone III stimulated a dose-dependent (0 to 10μmol) increase in length after a 24 hr exposure. No synergism was detected between juvenile hormone III and ecdysone when co-incubated with larvae. These results indicate that Ascaris larvae are growth-insensitive to low concentrations of biogenic amines of host origin. Conversely, biological transmitters of invertebrate origin are potent stimulators of larval growth.     

Biogenic amines and DOPA in the central nervous system of decapod crustaceans

The biogenic amines serotonin (5-HT), dopamine (DA), noradrenaline (NA), octopamine (OA) and the amino acid dihydroxyphenylalanine (DOPA) were identified and measured in the brain and the eyestalks of five decapod crustacean species using high pressure liquid chromatography (HPLC) with electrochemical detection. The amounts fall within 0.01-1.1 micrograms/g or 0.17-60 pmoles, and OA is the dominating amine in most species. THe DOPA levels in many of the species varied considerably between different measurements. It is concluded that the biogenic amines and DOPA are ubiquitous in the central nervous system of decapod crustaceans and the presence of NA and DOPA increases the number of presumed neurotransmitter/modulator candidates in the crustacean nervous system.     

Octopamine and serotonin have opposite effects on antipredator behavior in the orb-weaving spider, <Emphasis Type="Italic">Larinioides cornutus</Emphasis>

In this study, we experimentally elevated levels of octopamine and serotonin in an orb-weaving spider, and observed the effects on the antipredator behavior thanatosis (death feigning), activity level, and running speed. We found that octopamine significantly shortened the duration of thanatosis, and its effect wore off over 24 h. We also found that serotonin significantly lengthened thanatosis, but in this case, the effect persisted for over 24 h. Neither octopamine nor serotonin affected the general activity or running speed of the spiders. To our knowledge, this is the first study to directly explore the role of biogenic amines on a specific antipredator behavior in spiders. Given that spiders must be both aggressive toward prey, yet wary of predators, we believe that this system will be an outstanding model to explore connections between behavioral ecology and neurochemistry.     

Effect of larval density on whole-body biogenic amine levels of Tribolium freemani hinton

1. Various biogenic amines, their precursors and metabolites in gregarious and solitary forms of the flour beetle Tribolium freemani Hinton larvae were measured using high-performance liquid chromatography coupled with electrochemical detection.2. Octopamine was much more abundant than dopamine, serotonin, epinephrine, norepinephrine and related substances, suggesting that octopamine may play much more important roles in growth and behaviour of T. freemani than other biogenic amines and related substances.3. Octopamine levels were higher in gregarious forms than solitary forms of T. freemani larvae, and the relation between octopamine content, hormone cycles and motility is discussed.     

Place memory formation in Drosophila is independent of proper octopamine signaling

The biogenic amines play a critical role in establishing memories. In the insects, octopamine, dopamine, and serotonin have key functions in memory formation. For Drosophila, octopamine is necessary and sufficient for appetitive olfactory memory formation. Whether octopamine plays a general role in reinforcing memories in the fly is not known. Place learning in the heat-box associates high temperatures with one part of a narrow chamber, and a cool, strongly preferred temperature with the other half of the chamber. The cool-temperature-associated chamber half could provide a rewarding stimulus to a fly, and thus a place memory is composed of an aversive and rewarded memory component. The role of octopamine in place memory was thus tested. Using a mutation in the tyramine beta hydroxylase (TβH[M18]) and blocking of evoked synaptic transmission in the octopamine (and tyramine) neurons labeled with a tyramine decarboxylase-2 (TDC2) gene regulatory elements we found that reinforcement of place memories is independent of normal octopamine signaling. Thus, reinforcing mechanisms in Drosophila have specialized systems in the formation of specific memory types.