Evidence for the involvement of a SchistoFLRF-amide-like peptide in the neural control of locust oviduct

Summary The presence of a SchistoFLRFamide-like peptide associated with the oviducts of Locusta migratoria has been shown using sequential reversed-phase high performance liquid chromatography separation coupled with radioimmunoassay and bioassay. The peptide is present in areas of the oviduct which receive extensive innervation, with sixfold less peptide in areas that receive little innervation. Material with FMRFamide-like immunoreactivity (determined by radioimmunoassay) is also present in the oviducal nerve and VIIth abdominal ganglion.SchistoFLRFamide is a potent modulator of contraction of this visceral muscle, inhibiting or reducing the amplitude and frequency of spontaneous contractions, relaxing basal tonus, and reducing the amplitude of neurally-evoked, proctolin-induced, glutamate-induced and high potassium-induced contractions. The FMRFamide-like immunoreactivity within the oviducts which co-elutes with SchistoFLRFamide on two separations is also capable of reducing the amplitude of neurally-evoked and proctolin-induced contractions, and of inhibiting spontaneous contractions and relaxing basal tonus.The effects of SchistoFLRFamide upon this visceral muscle are not abolished by the -adrenergic receptor antagonist phentolamine and do not appear to be mediated by cyclic AMP. Thus the receptors for Schisto-FLRFamide are distinct from those of octopamine which mediate similar physiological effects but which are blocked by phentolamine and which are coupled to adenylate cyclase.The results indicate that SchistoFLRFamide, or a very similar peptide, which has previously been identified as a modulator of locust heart beat, is also associated with visceral muscle of the reproductive system, and may play a neural role in concert with octopamine, at modulating muscular activity.Abbreviations BPP Bovine pancreatic polypeptide - BSA Bovine serum albumin - EJP Excitatory junctional potential - FaRPs FMRFamide-related peptides - FLI FMRFamide-like immuno-reactivity - LMS Leucomyosuppressin - RIA Radioimmunoassay - RP-HPLC Reversed-phase high performance liquid chromatography - TFA Trifluoroacetic acid     

Some pharmacological properties of neuromuscular transmission in the oviduct of the locust,Locusta migratoria

The effects of various pharmacological agents on neurally evoked contractions of the visceral muscles of the oviduct of Locusta migratoria have been examined. The pentapeptide, proctolin, at low concentrations (10^?11 M?10^?10 M), induced an increase in the amplitude of neurally evoked contractions and basal tonus, and induced the appearance and increased the frequency of myogenic contractions. Glutamate, at 10^?4 M, produced a small transient contraction which in some preparations was accompanied by a reduction in amplitude of neurally evoked contractions. Octopamine, at 10^?6 M, reduced the amplitude of neurally evoked contractions and also resulted in a relaxation of the muscles. The octopaminergic effects were inhibited by the α-aminergic antagonist phentolamine. Neurally evoked contractions were unaffected by dopamine, 5-HT or the acetylcholine receptor antagonists atropine and hexamethonium. Acetylcholine increased the amplitude of neurally evoked contractions, but only at the high concentration of 10^?3 M. The possible role of proctolin and glutamate as excitatory neuro-transmitters and the inhibitory action of octopamine is discussed.     

Octopamine action on the spontaneous contractions of the isolated nerve cord of Lumbricus terrestris

Octopamine and synephrine were observed to effect the spontaneous rhythmic contractions displayed by the isolated ventral nerve cord of the earthworm, Lumbricus terrestris. octopamine and synephrine produced dose-dependent significant changes in the frequency, amplitude and basal tonus of the spontaneous contractions. Application of adrenergic receptor antagonists suggested the octopamine receptors to have some similarity to vertebrate alpha 1-adrenergic receptors. The spontaneous contractions were not abolished by tetrodotoxin (TTX) which suggested a myogenic origin for the contraction of the ventral nerve cord sheath muscles. Octopamine, in the presence of TTX, increased the basal tonus and maximum force of the spontaneous contractions.     

Modulation of tension production by proctolin in the dorsal longitudinal muscles of the cricket,Teleogryllus oceanicus

High-frequency electrical stimulation (～20 Hz) of the lateral nerve in abdominal segments of the cricket, Teleogryllus oceanicus, caused an increase in tonus of the abdominal dorsal longitudinal muscle (DLM). This effect persisted for 1–5 min following stimulation. Application of the pentapeptide proctolin (threshold 1–10 nM) mimicked the increase in muscle tonus produced by electrical stimulation. Individual twitches were unaffected or slightly reduced by proctolin. Low-frequency electrical stimulation (<7 Hz) of the lateral nerve counteracted a previously induced increase in muscle tonus, apparently by activation of an inhibitory motoneuron. γ-Aminobutyric acid (GABA) mimicked the effect of low-frequency stimulation and reduced muscle tonus. Octopamine, in concentrations of ≤0.1 mM, was inactive on the abdominal DLM when stimulated at low frequencies (0.5–2 Hz). Application of proctolin to the metathoracic DLM caused an increase in twitch amplitude but had little effect on basal tonus. In conjunction with the previously described responses of the metathoracic DLM to octopamine, these results show that the serially homologous abdominal and metathoracic DLMs have dissimilar responses to the modulators proctolin and octopamine.     

Stimulatory effects of male accessory-gland extracts on the myogenicity and the adenylate cyclase activity of the oviduct of Locusta migratoria

The effects of male accessory-gland extracts on the myogenic contractions and the adenylate cyclase activity of the oviduct of Locusta migratoria have been examined. The extracts stimulated first the frequency and the amplitude, then the tonus of the oviduct contractions in dose-dependent and reversible ways. They also stimulated the adenylate cyclase activity of oviduct disrupted-cell preparations. Extracts of opalescent glands (one of the 15 male accessory glands) gave similar results with only a quantitative difference. The tonus response is probably independent of the adenylate cyclase activity because octopamine and forskolin did not mimic this effect, and also because phentolamine was unable to inhibit the effect.Frequency and mainly amplitude responses can be induced through an adenylate cyclase-dependent receptor as shown by the similitude of actions with octopamine and forskolin. However, since the effects on the adenylate cyclase activity of octopamine and the accessory-gland extracts were cumulative, we concluded that these compounds are acting on two discrete types of receptors. All these results suggest that male accessory-gland secretions directly act upon the oviduct, in one case through adenylate cyclase-dependent receptors.     

A neurohormonal role for serotonin in the control of locust oviducts

Serotonin increases the frequency and amplitude of spontaneous contractions and leads to an increase in the basal tonus of the locust oviducts. These effects were dose-dependent and were seen on both the non-innnervated and innervated portion of the oviducts. Vertebrate type serotonin agonists and antagonists were used and the profile shows that the receptors on the non-innervated and innervated portion of the oviducts are more similar to 5-HT3 receptors than to either 5-HT1 or 5-HT2 receptors. No serotonin was found associated with the oviducts or the innervation to the oviducts using immunohistochemistry and HPLC coupled to electrochemical detection, suggesting a neurohormonal role for serotonin in the control of locust oviducts.     

Evidence for a possible neurotransmitter/neuromodulator role of tyramine on the locust oviducts

Visualization of the tyraminergic innervation of the oviducts was demonstrated by immunohistochemistry, and the presence of tyramine was confirmed using high-performance liquid chromatography coupled to electrochemical detection. Oviducts incubated in high-potassium saline released tyramine in a calcium-dependent manner. Stimulation of the oviducal nerves also resulted in tyramine release, suggesting that tyramine might function as a neurotransmitter/neuromodulator at the locust oviducts. Tyramine decreased the basal tension, and also attenuated proctolin-induced contractions in a dose-dependent manner over a range of doses between 10(-7) and 10(-4) M. Low concentrations of tyramine attenuated forskolin-stimulated cyclic AMP levels in a dose-dependent manner. This effect was not blocked by yohimbine. High concentrations of tyramine increased basal cyclic AMP levels of locust oviducts in a dose-dependent manner; however, the increases in cyclic AMP were only evident at the highest concentrations tested, 5 x 10(-5) and 10(-4) M tyramine. The tyramine-induced increase in cyclic AMP shared a similar pharmacological profile with the octopamine-induced increase in cyclic AMP. Tyramine increased the amplitude of excitatory junction potentials at low concentrations while hyperpolarizing the membrane potential by 2-5 mV. A further increase in the amplitude of the excitatory junction potentials and the occurrence of an active response was seen upon washing tyramine from the preparation. These results suggest that tyramine can activate at least three different endogenous receptors on the locust oviducts a putative tyramine receptor at low concentrations, a different tyramine receptor to inhibit muscle contraction, and an octopamine receptor at high concentrations.     

Proctolin's role in neurally evoked contractions of the locust oviducts

The effects of proctolin (RYLPT) on neurally evoked contractions of locust oviduct muscle were studied to examine the role of proctolin as a cotransmitter. Increasing the number of stimuli in a burst (from one to 30 stimuli) resulted in an increase in amplitude of contraction of locust oviduct muscle. Proctolin was capable of increasing the amplitude of neurally evoked contractions at lower-stimulus regimes (one- and two-stimulus bursts) but did not do so at higher-stimulus regimes (five- and 10-stimulus bursts). The effects of proctolin were dose dependent within the one- and two-stimulus regimes, with thresholds at 10⁻⁹ M and maxima at 2.5 × 10⁻⁸ M. Addition of proctolin increased the basal tonus and size of a postcontraction relaxation of the oviduct muscle in a dose-dependent manner during all stimulus regimes. However, the effect of proctolin on basal tonus and the postcontraction relaxation was much less at the higher stimulus regimes. Previously, several proctolin analogues have been tested for their ability to antagonize proctolin-induced contractions of the oviduct muscle. Since proctolin is proposed to be a cotransmitter at this neuromuscular junction, one of these analogues, cycloproctolin, was used to antagonize proctolin's effects on neurally evoked contractions. In the presence of the antagonist, the maximum amplitude induced by application of proctolin was decreased by 22.7%, while the proctolin-induced increase in basal tonus was decreased by 45.8%. Finally, the maximum increase in the size of the postcontraction relaxation caused by proctolin was lowered by 32.0%. The results of the present study show that exogenously applied proctolin is an excitant of the oviduct muscle at lower, rather than higher, stimulus regimes, and this latter inaction may be due to the corelease of endogenous proctolin during increased neural stimulation. © 1997 John Wiley & Sons, Inc. J Neurobiol 33: 139–150, 1997     

Interaction between octopamine and proctolin on the oviducts of Locusta migratoria

The biogenic amine octopamine and the pentapeptide proctolin are two important neuroactive chemicals that control contraction of the oviducts of the African locust Locusta migratoria. The physiological responses and signal transduction pathways used by octopamine and proctolin have been well characterized in the locust oviducts and this therefore provides the opportunity to examine the interaction between these two pathways. Octopamine, via the intracellular messenger adenosine 3',5'-cyclic monophosphate (cyclic AMP), inhibits contraction of the oviducts, while proctolin, via the phosphoinositol pathway, stimulates contraction. We have examined the physiological response of the oviducts to combinations of octopamine and proctolin and also looked at how combinations of these affect one of the main intracellular mediators of the octopamine response, namely cyclic AMP. It was found that application of octopamine to the oviducts led to a dose-dependent reduction in tonus of the muscle and also a decrease in the amplitude and frequency of spontaneous phasic contractions. Octopamine-induced relaxation was enhanced in the presence of the phosphodiesterase inhibitor, 3-isobutyl-1-methylxanthine (IBMX). Octopamine was also able to inhibit proctolin-induced contractions of the oviducts in a dose-dependent manner. A 10(-9) M proctolin-induced contraction was inhibited by 83% in the presence of 10(-5) M octopamine, and was completely inhibited in the presence of 10(-5) M octopamine plus 5x10(-4) M IBMX. Octopamine led to a dose-dependent increase in cyclic AMP content as measured by radioimmunoassay. In the presence of 10(-9) M proctolin, this octopamine-induced increase in cyclic AMP was reduced by as much as 60%. Proctolin also caused a dose-dependent decrease in the cyclic AMP elevation produced by 5x10(-6) M octopamine. These results indicate that octopamine and proctolin can antagonize each other's physiological response when added in combination, and that proctolin is able to modulate the response of the oviducts to octopamine by influencing cyclic AMP levels.     

Tyramine as a possible neurotransmitter/neuromodulator at the spermatheca of the African migratory locust, Locusta migratoria

Tyramine-like immunoreactivity was identified in neurons of the VIIIth abdominal ganglion and in axons projecting to the spermatheca of adult females of Locusta migratoria. Tyramine-like immunoreactive processes were also found throughout all regions of the spermatheca and tyramine-like immunoreactive bipolar or multipolar neurons were present on the spermathecal sac. HPLC coupled with electrochemical detection revealed more tyramine than octopamine present in spermathecal tissue. Electrical stimulation of the ventral ovipositor nerve resulted in a significant increase in calcium-dependent release of tyramine from the spermatheca. Both tyramine and octopamine increase the frequency and basal tonus of spermathecal contractions in a dose-dependent manner, with octopamine having a lower threshold. When tyramine is applied along with a half maximal octopamine dose, there is an additive effect on contractions of the spermatheca with slight synergistic effects at lower doses of tyramine. High concentrations of tyramine (10(-4)M) stimulated increases in cyclic AMP levels of the spermatheca; an effect blocked by phentolamine. Phentolamine has a higher affinity (and thus a lower IC(50) value congruent with5.6x10(-8)M) than yohimbine (IC(50) congruent with1.1x10(-4)M) in reducing tyramine-induced spermathecal contractions. Taken together, these results suggest that tyramine may be a co-transmitter with octopamine at the spermatheca, with both neuroactive chemicals acting on an octopamine receptor.     

Pharmacological characterization of the response of the leech pharynx to acetylcholine

In this study we document the sensitivity of the leech pharynx to acetylcholine and begin to characterize the acetylcholine receptor mediating this response by examining the effects of selective cholinergic agonists and antagonists on the contractile behavior of the pharynx. The order of potency derived from the EC50 of each agonist was (+/-)epibatidine > acetylcholine (in the presence of physostigmine) > McN A-343 > carbachol > nicotine. However, when response amplitude was considered, the order of potency to the tested agonists was (+/-)epibatidine > nicotine > McN A-343 > carbachol > acetylcholine. Acetylcholine-induced contractions of the pharynx were antagonized by d-tubocurarine, but not by alpha-bungarotoxin, alpha-conotoxin M1, or mecamylamine. Application of high concentrations of hexamethonium (1 mM) augmented the acetylcholine-induced contractions. However, this augmentation was apparently due to inhibition of acetylcholinesterase by hexamethonium. The muscarinic antagonist atropine produced complex actions and apparently acted as a mixed agonist/antagonist. Atropine by itself produced an increase in basal tonus and increased the frequency and amplitude of phasic contractions. Atropine increased the peak tension of the acetylcholine-induced response; however, it reduced the amplitude of both the acetylcholine-induced increase in basal tonus and integrated area. Based on the pharmacological profile of the pharyngeal acetylcholine response, we conclude that the acetylcholine receptor mediating the response is a nicotinic receptor. However, the responsiveness of the pharynx to muscarinic agents diverges from that of a classical nicotinic receptor.     

Crustacean cardioactive peptide is a modulator of oviduct contractions in Locusta migratoria

Crustacean cardioactive peptide (CCAP) stimulates the contractions of locust oviducts. CCAP increased the basal tonus and increased the frequency and amplitude of phasic contractions, as well as the amplitude of neurally-evoked oviduct contractions in a dose-dependent manner. Oviducts from Vth instar larvae and adult locusts aged 10 days or less, were more sensitive to CCAP than oviducts from adult locusts aged 12 days or more. This may be indicative of a differential expression of number or subtypes of CCAP receptors on the oviducts at different ages, and may be related to reproductive functions or to functions of CCAP on the oviducts during ecdysis. The oviducts appear more sensitive to CCAP when compared with previously published reports of CCAP actions on the hindgut. CCAP actions on the amplitude of neurally-evoked contractions of the oviducts are similar to those of proctolin, however, the oviducts are more sensitive to CCAP. No CCAP-like immunoreactive structures were discovered in the nerves innervating the oviducts, or on the oviducts themselves, confirming the previously published suggestion (Dircksen et al., 1991) that CCAP acts as a neurohormone at the oviducts. Cells showing CCAP-like immunoreactivity were discovered in the fat body associated with the oviducts and represent a potential source of CCAP, along with CCAP released from the transverse nerve and perivisceral organs.     

The oviduct musculature of the horsefly, Tabanus sulcifrons, and its response to 5-hydroxytryptamine and proctolin

ABSTRACT. A delicate lace-like membrane covers the ovaries of Tabanus sulcifrons. These membranes were found to contain muscle fibres that provide the organs with motile properties. The lateral oviducts consist of a single layer of longitudinal muscles that form a structural syncytium by means of an extensive anastomosis of the fibres comprising it. The common oviduct is composed of two muscle layers, an outer sheath of circular muscle and an inner substratum of longitudinal muscle. Both of these layers showed evidence of a structural syncytium. When isolated in saline, the oviduct was spontaneously active and gave a simple phasic pattern of contraction. Such muscle preparations were sensitive to both 5-hydroxytryptamine (5HT) and the insect myotropic peptide, proctolin. Excitation was generally indicated by a rise in muscle tonus or an increase in the frequency and amplitude of individual phasic contractions, or all three characteristics. The threshold for activation with 5HT was variable, ranging from as low as 4 × 10^-9M to 1 × 10^-7M. Proctolin evoked a noticeable increase in the tonus of most oviducts at 10^-10M. However, several preparations responded to as little as 3 × 10^-11M proctolin.     

Octopamine and 5‐hydroxytryptamine mediate hemocytic phagocytosis and nodule formation via eicosanoids in the beet armyworm,Spodoptera exigua

Octopamine and 5‐hydroxytryptamine (5‐HT) have been known to mediate cellular immune responses, such as hemocytic phagocytosis and nodule formation, during bacterial invasion in some insects. In addition, eicosanoids also mediate these cellular immune reactions in various insects, resulting in clearing the bacteria circulating in the hemolymph. This study investigated a hypothesis on signal cross‐talk between both types of immune mediators in the beet armyworm, Spodoptera exigua, which had been observed in the effect of eicosanoids on mediating the cellular immune responses. In response to bacterial infection, octopamine or 5‐HT markedly enhanced both hemocytic phagocytosis and nodule formation in S. exigua larvae. Their specific antagonists, phentolamine (an octopamine antagonist) or ketanserin (a 5‐HT antagonist) suppressed both cellular immune responses of S. exigua. These effects of biogenic monoamines on the immune mediation were expressed through eicosanoids because the inhibitory effects of both antagonists were rescued by the addition of arachidonic acid (a precursor of eicosanoid biosynthesis). Furthermore, the stimulatory effects of both monoamines on the cellular immune responses were significantly suppressed by different inhibitors acting at their specific levels of eicosanoid biosynthesis. Taken together, this study suggests that octopamine and 5‐HT can mediate hemocytic phagocytosis and nodule formation through a downstream signal pathway relayed by eicosanoids in S. exigua. © 2009 Wiley Periodicals, Inc.     

Octopamine modulates a central pattern generator associated with egg-laying in the locust,Locusta migratoria

Egg-laying in Locusta migratoria involves the control of a variety of complex behavioural patterns including those that regulate digging of the oviposition hole and retention of eggs during digging. These two behavioural patterns are under the control of central pattern generators (CPGs). The digging and egg-retention CPGs are coordinated and integrated with overlapping locations of neural substrate within the VIIth and VIIIth abdominal ganglia of the central nervous system (CNS). In fact, the egg-retention CPG of the VIIth abdominal ganglion is involved in both egg-retention and protraction of the abdomen during digging. The biogenic amine, octopamine, has peripheral effects on oviduct muscle, relaxing basal tension of the lateral and upper common oviduct and enabling egg passage. Here we show that octopamine also modulates the pattern of the egg-retention CPG by altering the motor pattern that controls the external ventral protractor of the VIIth abdominal segment. There is no change in the motor pattern that goes to the oviducts. Octopamine decreased the frequency of the largest amplitude action potential and decreased burst duration while leading to an increase in cycle duration and interburst interval. The effects of octopamine were greatly reduced in the presence of the α-adrenergic blocker, phentolamine, indicating that the action of octopamine was via a receptor. Thus, octopamine orchestrates events that can lead to oviposition, centrally inhibiting the digging behavior and peripherally relaxing the lateral and common oviducts to enable egg-laying.     

Multiplicity of chemical mechanisms of regulation of muscle contractions in <Emphasis Type="Italic">Lymnaea stagnalis</Emphasis> L.

Effects of acetylcholine, octopamine, and their antagonists, as well as of glutamic acid were studied on contractions of dorsal longitudinal muscle of the mollusc Lymnaea stagnalis L., evoked by electrical stimulation of n. cervicalis inferior. Acetylcholine and octopamine increased amplitude of contractions evoked by applications at concentrations about 10^–8 M and decreased at concentrations higher than 10^–7 M. Glutamic acid produced only inhibitory effect on the contraction amplitude, which appeared at concentrations beginning from 10^–9 M and higher. The acetylcholine antagonists atropine and d-tubocurarine also decreased amplitude of evoked contractions. Their blocking effect rose with increase of their concentrations in the range from 10^–9 M to 10^–5 M. Specificity of the effect of the studied substances was established in experiments with a combined application of the transmitters and their antagonists. The obtained results indicate multiplicity of chemical mechanisms of regulation of contractions of the dorsal longitudinal muscle in L. stagnalis.Translated from Zhurnal Evolyutsionnoi Biokhimii i Fiziologii, Vol. 41, No. 1, 2005, pp. 44–50.Original Russian Text Copyright © 2005 by Kononenko, Zhukov.     

Role of Potassium Channels in the Effects of Hydrogen Sulfide on Contractility of Gastric Smooth Muscle Cells in Rats

The effect of sodium hydrosulfide (NaHS), a hydrogen sulfide (H₂S) donor, on spontaneous contractile activity of rat gastric smooth muscle cells was analyzed. Experiments were conducted on gastric stripes under conditions of isometric contraction. It was shown that NaHS has a biphasic effect on spontaneous contractile activity, increasing tonic tension and the amplitude of phasic contractions within the first minutes since application. This initial phase is followed by a decrease in amplitude, basal tone, and frequency of spontaneous contractions. The inhibitory effect of NaHS was dose-dependent at concentrations from 10 to 600 μM. Preliminary application of tetraethylammonium and 4-aminopirydine, inhibitors of voltage-gated and calciumactivated potassium channels, prevented a NaHS-induced initial increase in basal tone and phasic contraction amplitude. Activation of ATP-dependent potassium channels (KATP-channels) by diazoxide prevented in part a NaHS-induced decrease in basal tone and amplitude of spontaneous contractions. Glibenclamide, an inhibitor of K_ATP-channels, decreased the inhibitory effect of NaHS on amplitude, basal tone and frequency of spontaneous contractions. It was concluded that in rat gastric smooth muscles the excitatory effect of H₂S is mediated by the inhibition of voltagegated and calcium-activated potassium channels, while its inhibitory effect involves the activation of K_ATP-channels.     

Neuromuscular modulation in Limulus by both octopamine and proctolin

Both octopamine and proctolin potentiate nerve-evoked skeletal muscle contractions in the horseshoe crab, Limulus. The threshold concentration for octopamine was 10^?9 to 10^?8M, while for proctolin it was 3 × 10^?9M. Norepinephrine and dopamine produced effects similar to octopamine but at higher thresholds; tyramine and serotonin were ineffective. Octopamine caused significant increases in amplitudes of excitatory postsynaptic potentials (epsps) of muscle fibers, but had little effect on muscle fiber input resistance or membrane potential. Also, octopamine did not affect depolarization of muscle fibers and subsequent contraction due to the direct action of exogenously applied glutamate. These results suggest that octopamine potentiates nerve-evoked contractions primarily by facilitating release of neuromuscular transmitter. At concentrations above 10^?7M, however, octopamine sometimes caused muscle spikes in response to motoneuron stimulation, a finding that suggests that octopamine may also have some postsynaptic action. Proctolin potentiated the muscle contractions evoked by glutamate but had little effect on glutamate-evoked muscle fiber depolarization, muscle fiber input resistance, or membrane potential. Thus, proctolin appears to act directly on skeletal muscle to enhance contractility. The proctolin-induced potentiations of contraction were sometimes accompanied by modest increases in epsp amplitude, so that unlike lobster skeletal and Limulus cardiac neuromuscular preparations, proctolin may have a secondary direct synaptic effect. Both octopamine and proctolin have been found in Limulus cardiac ganglion. This potential access to the hemolymph and the relatively low threshold concentrations needed for physiological action suggest that octopamine and proctolin could function as hormonal modulators of neuromuscular function in Limulus.     

Larvae of the small white butterfly,Pieris rapae,express a novel serotonin receptor

The biogenic amine serotonin ( 5‐hydroxytryptamine, 5‐HT) is a neurotransmitter in vertebrates and invertebrates. It acts in regulation and modulation of many physiological and behavioral processes through G‐protein‐coupled receptors. Five 5‐HT receptor subtypes have been reported in Drosophila that share high similarity with mammalian 5‐HT_1A, 5‐HT_1B, 5‐HT_2A, 5‐HT_2B, and 5‐HT₇ receptors. We isolated a cDNA (Pr5‐HT₈) from larval Pieris rapae, which shares relatively low similarity to the known 5‐HT receptor classes. After heterologous expression in HEK293 cells, Pr5‐HT₈ mediated increased [Ca²⁺]_i in response to low concentrations (< 10 nM) of 5‐HT. The receptor did not affect [cAMP]_i even at high concentrations (> 10 μM) of 5‐HT. Dopamine, octopamine, and tyramine did not influence receptor signaling. Pr5‐HT₈ was also activated by various 5‐HT receptor agonists including 5‐methoxytryptamine, (±)‐8‐Hydroxy‐2‐(dipropylamino) tetralin, and 5‐carboxamidotryptamine. Methiothepin, a non‐selective 5‐HT receptor antagonist, activated Pr5‐HT₈. WAY 10635, a 5‐HT_1A antagonist, but not SB‐269970, SB‐216641, or RS‐127445, inhibited 5‐HT‐induced [Ca²⁺]_i increases. We infer that Pr5‐HT₈ represents the first recognized member of a novel 5‐HT receptor class with a unique pharmacological profile. We found orthologs of Pr5‐HT₈ in some insect pests and vectors such as beetles and mosquitoes, but not in the genomes of honeybee or parasitoid wasps. This is likely to be an invertebrate‐specific receptor because there were no similar receptors in mammals.

     

Octopamine enhances phagocytosis in cockroach hemocytes: Involvement of inositol trisphosphate

Octopamine and 5-hydroxytryptamine (5-HT) were previously shown to affect phagocytosis in cockroach hemocytes through unidentified receptor-mediated events. In the present study, we examined the ability of 5-HT and octopamine to enhance inositol trisphosphate (IP₃) production using hemocyte membranes of the American cockroach, Periplaneta americana. Octopamine enhanced IP₃ production with a maximal peak at 100 nM. Similarly, 5-HT enhanced IP₃ production with a maximal effect at 10 nM. The effects of 5-HT and octopamine are not additive, suggesting that both are working through the same receptor. Phentolamine, a general octopamine antagonist, blocked the effects of octopamine and 5-HT, while a mammalian 5-HT₂ antagonist that blocks 5-HT-sensitive receptors in insect peripheral tissue, ketanserin, did not. A pharmacological profile indicates that the receptor is similar to an octopamine₁-type. Octopamine at 1 μM increased phagocytosis in cockroach hemocytes exposed to Staphylococcus aureus in vitro, and this effect was mimicked by IP₃ (10 μM). The octopamine-treated hemocytes were shown to increase IP₃ production in the latter stage of phagocytosis. Adult cockroaches exposed to an LD₅₀ dose of S. aureus in conjunction with either 0.1 mM octopamine or the octopamine₁ agonist, clonidine, had higher survival rates compared to saline-treated cockroaches. Correspondingly, the octopamine₁ antagonist, chlorpromazine, partially blocked the octopamine-mediated increase in cockroach survival. © 1994 Wiley-Liss, Inc.