Red Pigment-concentrating Hormone Research Articles

REGULATION OF THE RELEASE OF CHROMATOPHOROTROPIC NEUROHORMONES FROM THE ISOLATED EYESTALK OF THE FIDDLER CRAB,UCA PUGILATOR

Electrical stimulation of the isolated eyestalk of Uca pugilator induces the release of several peptides which affect epidermal chromatophores. Thresholds for release of these peptides were different, that for red pigment concentrating hormone (RPCH) being lowest, for black pigment dispersing hormone (BPDH) highest, and for black pigment concentrating hormone (BPCH) intermediate, but no red pigment dispersing hormone(RPDH) was detected at any voltage. Neurotransmitters, known to be present in crustacean central nervous systems, induced chromatophore dose dependent responses. Norepinephrine induced BPDH release, and dopamine induced both RPCH and BPCH release.

Adipokinetic and hyperglycaemic factors of different insect species: Separation with high performance liquid chromatography

Corpus cardiacum extracts from the phasmids, Carausius morosus, Cuniculina impigra, Sipyloidea sipylus, Acrophylla wuelfingi, Eurycantha goliath, Bacillus rossius and Extatosoma tiaratum, from the Orthopterans, Locusta migratoria and Gryllus bimaculatus, from the Dictyopterans, Periplaneta americana, Gromphadorrhina coquereliana and Blaberus craniifer, from the Coleopterans Tenebrio molitor and Pachnoda sp., synthetic adipokinetic hormone and synthetic crustacean red pigment-concentrating hormone (RPCH) were injected into locusts, cockroaches and ligated stick insects as bioassay systems for adipokinetic and hyperglycaemic substances, respectively. The locust and cockroach bioassay gave positive results with all corpus cardiacum material tested (however the lipid response in locusts upon injection of T. molitor corpus cardiacum extract was very poor). The stick insect bioassay was quite specific for stick insect corpus cardiacum material; only corpus cardiacum extracts from a few other species ( G. bimaculatus, P. americana, G. coquereliana and Pachnoda sp.) showed weak activity. All other extracts, including synthetic adipokinetic hormone and RPCH, failed to induce a response. Separations of corpus cardiacum extracts from L. migratoria, P. americana, T. molitor, C. morosus and S. sipylus were achieved on reversed-phase high-performance liquid chromatography (RP-HPLC). Locust corpus cardiacum extract showed two absorbance peaks with adipokinetic activity, adipokinetic hormones I and II. The peaks with hyperglycaemic activity from P. americana corpus cardiacum extracts had different retention times to those of locust adipokinetic hormones I and II. Stick insect corpus cardiacum extracts revealed also 2 absorbance peaks with adipokinetic activity, the major one co-eluting with RPCH. The active compound from corpus cardiacum extracts of T. molitor appeared to elute close to locust adipokinetic hormone I.

Synthesis of shrimp red pigment-concentrating hormone analogs and their biological activity in locusts.

Two analogs of the red pigment-concentrating hormone (RPCH) have been synthesized by the solid-phase method: [Thr6]-RPCH (I) and [Tyr4, Thr6]-RCPH (II). Analog I has the same amino acid composition as the second adipokinetic hormone (AKH-II) isolated from locust corpora cardiaca. Bioassay for lipid-mobilizing activity in adult male locusts gave the following increases in hemolymph lipid content: AKH-I, 3.5; I, 2.4; II, 2.9. The biological response shown by I lends support to the conclusion that its sequence is that of the presumptive AKH-II. Replacement of Phe in position 4 by Tyr does not reduce the adipokinetic response.

Chemical Properties and Physiological Actions of Crustacean Chromatophorotropins

The crustacean pigment-translocating hormones, the red pigment-concentrating hormone (RPCH), an octapeptide, and the light-adapting distal retinal pigment hormone (DRPH), an octadecapeptide, are the first invertebrate neurohormones to be fully characterized. Studies with both purified and synthetic hormones show that, in certain decapods, RPCH is a general pigment-concentrating hormone (PCH), affecting the pigments of all kinds of chromatophores (erythrophores, xanthophores, leucophores and melanophores); the DRPH seems to serve not only light-adapting function, but also act as a general chromatophore pigment-dispersing hormone (PDH). The two hormones thus function as antagonists when regulating the color-adaptation of the decapod crustaceans. PCH activity is widely distributed within the arthropod endocrine systems. The first characterized insect neurohormones, the locust adipokinetic hormones (AKH I and AKH II), show close structural similarities to the crustacean hormone, indicating a common evolution of some of the arthropod neurohormones. Physiological studies of the three hormones (RPCH, AKH I, and AKH II) and their synthetic analogs show that they crossreact, i.e., they all exhibit pigment-concentrating activity when tested on decapod crustaceans, adipokinetic activity when tested on locusts, and hyperglycemic activity when tested on cockroaches, although each of the hormones is more potent in its own system. Structure-function studies show, however, that quite different binding-site requirements exist for the hormones in activating their receptors on the various target tissues. The physiological specificity in their action therefore seems to depend on a differential evolution of the hormone receptors.

Regulation of Release and Mode of Action of Crustacean Chromatophorotropins

Pigment movements in crustacean chromatophores are regulated by pigment-concentrating and pigment-dispersing neurosecretory hormones. The release of these hormones from the neurosecretory system is thought to be controlled by neural signals. Although the exact cellular sources of chromatophorotropins and the nature of synaptic input to the neurosecretory system remain unresolved, recent pharmacological studies on fiddler crabs indicate that certain biogenic amines contribute to the release of some of the chromatophorotropins. The current evidence suggests that: 5-hydroxytryptamine stimulates the release of RPDH (red pigment dispersing hormone), dopamine triggers the release of RPCH (red pigment concentrating hormone), and norepinephrine stimulates the release of BPDH/MDH (black pigment/melanin dispersing hormone). Nothing is known of the regulation of release of leucophorotropins and xanthophorotropins. Like most other peptide hormones, crustacean chromatophorotropins are thought to exert their effect by first binding to receptors on the cell surface and then triggering a series of events leading to cellular responses, but the details are far from clear. Recent work suggests that hormone-induced pigment movements may be modulated/mediated by alterations in cyclic nucleotide levels and/or by changes in Ca2+ distribution or flux. Both cyclic nucleotides and Ca2+ may modulate motility by regulating the integrity and/or function of cytoskeletal elements, but the details of this mechanism and the precise roles of cytoskeletal elements in the pigment-translocating process remain unresolved.

Invertebrate neuropeptide hormones.

The development of a long-term research program on the neurosecretory hormones of arthropods is described. The purification and full characterization of the first invertebrate neurohormones, the red pigment-concentrating hormone (RPCH) and the distal retinal pigment hormone (DRPH) demonstrated that they are peptides, an octapeptide and an octadecapeptide, respectively. Physiological function studies with the pure hormones and their synthetic preparations showed that the RPCH acts as a general pigment-concentrating hormone (PCH), and that the DRPH, in addition to its light-adaptive function, also constitutes a general pigment-dispersing hormone (PDH). In the regulation of the color-adaptation of the animals, the two hormones act as antagonists. The chromatophorotropic activities are widely distributed within the arthropod neuroendocrine systems. Purification of the pigment-concentrating activities from the locust corpora cardiaca lead to the isolation and characterization of the first insect neurohormones, the adipokinetic hormones (AKH I and AKH II). These two hormones, AKH I being a decapeptide and AKH II being an octapeptide, are close structural analogs to the crustacean PCH, demonstrating a common evolution of arthropod neurohormones. The hormones of this PCH-family all cross-react, but structure-function studies of the hormones show that quite different parts of their structure are involved in their binding to the various receptors.

Effects of monoamines and monoaminergic agents on migration of the red pigment in chromatophores of the shrimp, Macrobrachium ohione

1. The effects of 5-hydroxytryptamine (5-HT) and the 5-hydroxytryptaminergic agents fenfluramine, fluoxetine and 5-MeODMT, and of dopamine (DA) and the dopaminergic agents ADTN, apor-morphine and piribedil on the disposition and translocation of the red pigment in chromatophores of the shrimp, Macrobrachium ohione, were determined. 2. 5-HT evoked dose-dependent red pigment dispersion whereas DA produced dose-related concentration of the red pigment, but only in intact shrimp. Neither of these monoamines affected red pigment migration in vitro. 3. The data obtained using drugs known to act selectively in mammals via 5-hydroxytryptaminergic and dopaminergic mechanisms on the translocation of the red pigment from the chromatophores both in vitro and in vitro lead to the hypothesis that 5-HT and DA serve as neurotransmitters triggering release of red pigment-dispersing hormone and red pigment-concentrating hormone, respectively, in the shrimp, Macrobrachium ohione.

Roles of 5-hydroxytryptamine and dopamine as neurotransmitters eliciting release of erythrophorotropic hormones in the fiddler crab, [formula omitted

This article reviews the endocrinological, pharmacological and biochemical evidence ascribing neurotransmitter roles for 5-hydroxytryptamine (5-HT, serotonin) in eliciting the release of red pigment-dispersing hormone (RPDH) and for dopamine (DA) in stimulating the release of red pigment-concentrating hormone (RPCH) in the fiddler crab, Uca pugilator . 5-HT produces red pigment dispersion in intact crabs, but only indirectly. Likewise, DA evokes red pigment concentration in vivo but it has no effect on red chromatophores (erythrophores) in isolated legs. The data obtained with 5-HT and DA agonists and antagonists on red pigment translocation in vivo and in vitro , are consistent with their neurotransmitter candidacies in evoking the release of these erythrophorotropic hormones.

Hormonal activation of protein kinase and lipid mobilization in the locust fat body in vitro

Cyclic nucleotide-activated protein kinase in locust fat body is elevated by extracts containing adipokinetic hormone (AKH) from locust corpora cardiaca or by synthetic redpigment-concentrating hormone (RPCH) which also exhibits lipid-mobilizing activity. Maximal activation is first attained in vitro 90 sec after RPCH is added to the incubation medium and 3.5 min after addition of AKH extract. This activation reflects the increase in endogenous cyclic nucleotides. Incubation of fat body in the presence of exogenous cyclic nucleotides activates lipase, and RPCH stimulation ultimately elevates the level of transported diacylglycerol 2.3-fold. RPCH stimulates fat body lipase in vivo.

Activation of fat body glycogen phosphorylase in Locusta migratoria by corpus cardiacum extract and synthetic adipokinetic hormone

Between 10 and 20 per cent of the total glycogen phosphorylase in the fat body of mature Locusta migratoria of both sexes is in the active form. Injection of an aqueous corpus cardiacum (CC) extract results in a rapid activation: within 2 min the level of active phosphorylase is significantly increased and full activation is reached within 10 to 20 min. As little as 0.002 CC gland equivalents stimulate fat body glycogen phosphorylase significantly and maximum activation is obtained with 0.05 CC gland equivalents. From experiments with known quantities of injected synthetic adipokinetic hormone (SAKH), it appears that this hormone cannot account for all the activation. This is supported by results obtained when extracts of carefully isolated storage lobes are injected; at the dose used here these have no adipokinetic activity, but activate fat body phosphorylase. Furthermore, when locusts are ‘stressed’ by rotation, although no adipokinetic hormone is released, an activation of phosphorylase occurs. Starvation causes also an increase in the active form of the enzyme. The fat body receptor sites of the locust recognise also the crustacean red pigment concentrating hormone (RPCH), whose structure closely resembles that of the locust adipokinetic hormone, leading to activation of the phosphorylase. However, RPCH is about 2.5–5 times less potent than SAKH. Crude CC extracts of a stick insect ( Carausius morosus), a cockroach ( Periplaneta americana) and the tobacco hornworm ( Manduca sexta) activate locust fat body phosphorylase, although this last extract has no effect on lipid elevation. On the other hand, CC extracts of the death's head hawk moth ( Acherontia atropos) and purified crustacean hyperglycaemic hormone from a crayfish ( Orconectes limosus) have no effect.

The effects of biogenic amines on color changes of the fiddler crab, Uca pugilator: Further evidence for roles of 5-hydroxytryptamine and dopamine as neurotransmitters triggering release of erythrophorotropic hormones

1. 1. The effects of 5-hydroxytrytryptaminergic potentiators (Fluvoxamine and Fluoxetine), a 5-hydroxytryptaminergic antagonist (LY53857), a dopaminergic agonist (ADTN) and dopaminergic antagonists (Spiroperidol and γ-hydroxybutyrate) on color changes involving the erythrophores of the fiddler crab, Uca pugilator, were determined. 2. 2. The results were consistent with the hypotheses that 5-hydroxytryptamine triggers release of a red pigment-dispersing hormone and that dopamine triggers release of a red pigment-concentrating hormone in Uca pugilator.

Evidence Implicating Calcium as the Second Messenger for Red Pigment-Concentrating Hormone in the Prawn Palaemonetes pugio

Three different agents were examined for their effect on pigment concentration in the large erythrophores of Palaemonetes pugio: guanosine 3',5' cyclic monophosphate (cGMP), ouabain, and the divalent cation ionophore A23187. High concentrations (10 mM) of cGMP in the medium did not cause pigment concentration. High concentrations (1 mg/ml) of ouabain failed to inhibit pigment concentration induced by either the ionophore A23187 or the red pigment-concentrating hormone. The ionophore A23187, in Ca⁺⁺-containing saline, caused pigment concentration in the large erythrophores of three internal organs: ventral nerve cord, intestine, and ovary. When erythrophores whose pigment had been concentrated by the ionophore were exposed to Ca⁺⁺-free/EGTA-containing saline, pigment dispersion resulted. Erythrophores whose pigment was concentrated by long-term exposure to a white background did not disperse their pigment when treated with the ionophore in Ca⁺⁺-free/EGTA saline. The red pigment-concentrating hormone responded to Ca⁺⁺-free/EGTA saline by an initial concentration of pigment, followed by redispersion. The activity of the red pigment-concentrating hormone was greatly inhibited in saline whose calcium had been replaced with lanthanum. On the basis of the data above, it is tentatively proposed that the red pigment-concentrating hormone, upon binding to its receptor, behaves as a Ca⁺⁺ ionophore.

Preparation of an 125I-labeled red pigment-concentrating hormone analog

The red pigment-concentrating hormone (RPCH) from crustacea is an octapeptide with the following structure: pGlu-Leu-Asn-Phe-Ser-Pro-Gly-Trp-NH 2, [4-Tyr]RPCH has been synthesized and was 4 times as active as RPCH in the Leander assay. Iodination of the hormone analog using chloramin-T, lactoperoxidase, solid-state lactoperoxidase, and thallium chloride was carried out and the results were compared. 125I-labeled [4-Tyr]RPCH was prepared using a modificated chloramin-T method and was purified from unlabeled and diiodinated peptide by chromatography on Sephadex LH-20.

Characterization of a second peptide with adipokinetic and red pigment-concentrating activity from the locust corpora cardiaca

A new peptide with both adipokinetic activity in the locust and red pigment-concentrating activity in the shrimp, can be readily separated from the adipokinetic hormone in extracts of corpora cardiaca (CC) from Schistocerca americana gregaria by the use of gel chromatography and the Leander adspersus bioassay. The new peptide accounts for 20% of the total biological activity in locust CC, and it is located principally in the CC glandular lobe. The amino acid composition of the pure peptide is: Asp, Thr, Ser, Glu, Gly, Leu, Phe, Trp. This composition is similar to, but not identical with the red pigment-concentrating hormone of the shrimp.

Structure-function studies on red pigment-concentrating hormone, II. The significance of the C-terminal tryptophan amide.

The significance of the C-terminal tryptophan residue of the red pigment-concentrating hormone (RPCH: Glu-Leu-Asn-Phe-Ser-Pro-Gly-Trp-NH2) regulating the blanching of the crustacean chromatophores has been investigated. RPCH and a number of analogues that differ only in the C-terminal part of the hormone, have been synthesized and assayed for biological activity on the shrimp Leander adspersus. It has been shown that the indole skeleton of tryptophan is an absolute requirement for the biological activity of the hormone. To provide maximum response the tryptophan must be blocked as the amide. The activity of synthetic [Tyr4]RPCH and adipokinetic hormone (AKH) purified from Schistocerca gregaria has been compared with the activity of synthetic RPCH.

Color changes induced by peptide derivatives of lysergic acid in the fiddler crab, UCA PANACEA

1. Ergocornine, α-ergocryptine and 2-Br-α-ergocryptine (bromocryptine, CB-154) evoke erythrophoric pigment concentration in intact fiddler crabs. 2. Dopamine evokes erythrophoric pigment concentration in intact fiddler crabs. DA was more effective in eliciting this response than the ergot alkaloids. 3. The ergot alkaloids act synergistically with DA in eliciting erythrophoric pigment concentration. 4. The ergot alkaloids and DA do not cause erythrophoric pigment concentration in vitro. Their actions in vivo appear to be mediated by the release of a red pigment-concentrating hormone from the neuroendocrine system.

Structure-function studies on red pigment-concentrating hormone; the significance of the terminal residues.

The significance of the terminal residues of the red pigment-concentrating hormone (RPCH: Glu-Leu-Asn-Phe-Ser-Pro-Gly-Trp-NH2) for its blanching effect on crustacean chromatophores has been investigated. The chemical synthesis of the following small analogues, starting from the C-terminal amino acid are described: Ac-Trp-NH2, Boc-Gly-Trp-NH2, Ac-Gly-Trp-NH2, Gly-Trp-NH2, Ac-Pro-Gly-Trp-NH2, Glu-Trp-NH2, Glu-Pro-Gly-Trp-NH2, Ac-Pro-Gly and Glu-Pro-Gly-NH2. Assay of the biological activity of the various synthetic compounds in the shrimp Leander adspersus has established that only the C-terminal tryptophan residue is indispensable for the blanching effect of the hormone, although elongation of the chain length improves its potency.

EVIDENCE FOR AN ADIPOKINETIC FUNCTION OF THE RPCH ACTIVITY PRESENT IN THE DESERT LOCUST NEUROENDOCRINE SYSTEM

1. Red pigment concentrating hormone-like activity (RPCA) has been demonstrated and quantified in the head or cephalic neuroendocrine system of several insects. Schistocerca gregaria adults contained the highest levels of RPCA.2. RPCA was found in all post-embryonic life cycle stages of S. gregaria, and seems to be present in equal quantities in both males and females. Adult locusts contain significantly more RPCA than do immature animals.3. RPCA is concentrated in the locust CC glandular lobe hut is also present in the CC storage lobe, brain and hemolymph of both mature and immature animals. Significant variations in CC RPCA content have been demonstrated in both mature and immature stages.4. Synthetic red pigment concentrating hormone (RPCH) is adipokinetic in both intact and neck-ligatured S. gregaria adults and in intact immature animals. Doses of about 6 ng and 50 ng, respectively, result in minimal and maximal adipokinetic responses. RPCH selectively elevates hemolymph 1.2-diglycerides.5. RPCH in la...

Relative potencies of locust adipokinetic hormone and prawn red pigment-concentrating hormone in insect and crustacean systems

The molar activities of locust adipokinetic hormone (AKH) and prawn red pigment-concentrating hormone (RPCH) in mobilising lipids in locusts and on red pigment concentration in prawns are compared. AKH is five times more potent than RPCH in the locust system, whereas RPCH is four times more active than AKH on prawn erythrophores. These two hormones are equally effective in concentrating black pigments in the shrimp, Crangon. RPCH is more potent than AKH in causing hyperglycaemia in cockroaches, through relatively large doses of both hormones are needed to elicit this response. AKH, in lower doses, is also able to induce hyperglycaemia in the crab, Portunus. The close structural similarity of AKH and RPCH clearly account for their activity in each other's systems. However, it is not known whether the additional effects observed here reflect structural resemblances among other arthropod neurohormones.

Antagonistic actions of dopamine and 5-hydroxytryptamine on color changes in the fiddler crab, Uca pugilator

1. 1. Dopamine (DA) produces pigment concentration in the erythrophores of intact fiddler crabs, but has no effect on the erythrophores in isolated legs. 2. 2. DA antagonizes the red pigment-dispersing action of 5-hydroxytryptamine. 3. 3. It is hypothesized that DA triggers release of red pigment-concentrating hormone.