Study of the effects of the casein derived bitter tastant on the melanophores in milieu with the melatonin receptors

The present study was undertaken to ascertain whether the casein derived bitter tastant Cyclo (Leu-Trp) [CLT] has an affinity or not for the particular receptors of the pineal hormone, melatonin, on the melanophores of a major carp Labeo rohita (Ham.). The bitter tastant CLT, in the dose range of 3.34?×?10^?16 M to 3.34?×?10^?4 M, has induced an aggregatory effect but not in a dose dependent manner. Binding of CLT with the receptors may vary at different concentrations. Denervation of the melanophores has shown a complete inhibition of the CLT mediated aggregation. Prazosin has partially inhibited the aggregatory effect of CLT. Moreover, the bitter tastant’s response is mediated through the α₂ adrenoceptors only at particular dose ranges. The MT₁ and MT₂ melatonin receptor antagonist luzindole and the MT₂ specific antagonist K185 have perfectly blocked the aggregatory effects of CLT. We have found that the CLT mediated aggregatory effect is dependent upon the release of neurotransmitters and the two subtypes of melatonin (MT) receptors (MT₁ and MT₂) possess a perfect affinity towards the bitter tastant CLT. Our study demands a need to further make a clinical research on the effects of bitter tastants on the physiology of the biological rhythm maintaining hormone melatonin.     

Analyzing the responses of saccharin in context with melatonin receptors on the melanophores of the fish Labeo rohita (Ham.)

The hormone melatonin regulates the biological clock and assist in various other physiologies of vertebrates. Present work is intended to check the affinity of saccharin towards the melatonin receptors and the possible role of saccharin interference in the melatonin physiology. The present in vitro study is based on the working model of isolated scale melanophores in the dorso-lateral region of Labeo rohita. The pigment cells were incubated in the agonist and the antagonists within a limited time frame and subsequently their Melanophore Size Index (MSI) were calculated. The inferences were drafted through the observed signal transduction upshots in pigment translocations within the melanophores. Saccharin, in a wide dose range, has consistently induced a concentration-related aggregation similar to the aggregatory effect as shown by melatonin on the melanophores. Binding of saccharin with the receptors and eliciting its aggregatory effect is partially dependent on the release of neurotransmitters. The aggregatory effects were found to be significantly blocked by luzindole, K185, and prazosin, which are the potent melatonin receptor blockers, at the higher concentrations of saccharin. Hence, all the three subtypes of melatonin receptors viz. MT?, MT?, and MT? are participating in saccharin-mediated aggregations. Blocking by neomycin shows that Ca2? ions are very crucial in dispensing the aggregatory effect of the sweetener. This research demands that an intensive and careful thorough study should be made about saccharin, specifically its effects upon melatonin physiology, before its unwarranted use as the food ingredients for human use.     

Evidence for the presence of novel β-melatonin receptors along with classical α-melatonin receptors in the fish Rasbora daniconius (Ham.)

Abstract

The effects of melatonin (MT) were examined on the isolated scale melanophores from dorso-lateral (D-L) and band regions of a tropical fish Rasbora daniconius. Our study primarily aimed for further depiction of the signaling receptors involved in MT mediated pigment translocations in the fish. Melanophore Size Index (MSI) was employed as a recording parameter for the responses of melanophores to MT and various antagonists. MT has induced aggregation as well as dispersion in D-L region and aggregation in band region melanophores during summer season. During winter, MT-induced responses were only of aggregatory type in D-L region, while in the band region there was an increase in the sensitivity. The responses of the melanophores to MT were reversible. The aggregation of innervated melanophores induced by MT on the D-L and band regions was partially mediated through the neurotransmitters released under the influence of MT and partially by the specific MT receptors. Luzindole and K185 have completely blocked the aggregatory responses of D-L and band region melanophores. Aggregatory receptors may be of the conventional α-MT type. Dispersion of D-L and band region melanophores induced by MT in the presence of various antagonists and on denervated band region could be the result of activation of β-MT receptors of dispersive nature. Presence of α and β MT receptors is thus indicated in this fish melanophores.     

Study of the effects of the casein derived bitter tastant on the melanophores in milieu with the melatonin receptors

The present study was undertaken to ascertain whether the casein derived bitter tastant Cyclo (Leu-Trp) [CLT] has an affinity or not for the particular receptors of the pineal hormone, melatonin, on the melanophores of a major carp Labeo rohita (Ham.). The bitter tastant CLT, in the dose range of 3.34×10(-16) M to 3.34×10(-4) M, has induced an aggregatory effect but not in a dose dependent manner. Binding of CLT with the receptors may vary at different concentrations. Denervation of the melanophores has shown a complete inhibition of the CLT mediated aggregation. Prazosin has partially inhibited the aggregatory effect of CLT. Moreover, the bitter tastant's response is mediated through the α2 adrenoceptors only at particular dose ranges. The MT1 and MT2 melatonin receptor antagonist luzindole and the MT2 specific antagonist K185 have perfectly blocked the aggregatory effects of CLT. We have found that the CLT mediated aggregatory effect is dependent upon the release of neurotransmitters and the two subtypes of melatonin (MT) receptors (MT1 and MT2) possess a perfect affinity towards the bitter tastant CLT. Our study demands a need to further make a clinical research on the effects of bitter tastants on the physiology of the biological rhythm maintaining hormone melatonin.     

On the role of histamine receptors in regulating pigmentary responses in Oreochromis mossambicus melanophores

Purpose: The present work was carried out to reveal the involvement of histamine receptors at the neuro-melanophore junction of teleost, Oreochromis mossambicus.

Methods: The isolated scale melanophores were assayed using the mean melanophore size index and their responses were recorded in presence of various concentrations of histamine along with H₁ and H₂ receptor specific agonists and antagonist and potentiator compound 48/80.

Results: Melanophores showed high sensitivity to histamine and its specific agonists. Histamine caused a dose-dependent pigment aggregation, whereas 2-(2-Pyridyl) ethylamine (PEA), a specific H₁R agonist also caused aggregation in a similar manner. Conversely, amthamine, a specific H₂R agonist resulted in pigment dispersion. The effects were antagonized by mepyramine; specific H₁R antagonist and ranitidine a specific H₂R antagonist.

Conclusion: It is concluded that O. mossambicus melanophores have both H₁ and H₂ receptors which mediate melanophore aggregation and dispersion respectively. Compound 48/80 augmented the melanin-aggregating and dispersing effects of PEA and amthamine. It is suggested that the effect of histamine is directly mediated through H1 and H2 receptors, whereas H1Rs may be predominantly involved in the aggregatory responses.     

β-Adrenergic Receptor Subtypes in Melanophores of the Marine Gobies Tridentiger trigonocephalus and Chasmichthys gulosus

The subtype of β-adrenergic receptors in melanophores of the marine gobies Tridentiger trigonocephalus and Chasmichthys gulosus was studied. Pigment of denervated melanophores in isolated, split caudal fins was preliminarily aggregated by incubating the specimens in a physiological saline containing 10 μM phentolamine and 30–100 μM verapamil or 2–10 nM melatonin, and the responses of the melanophores to a β-adrenergic agonist added to the incubating medium were recorded photoelectrically. The β-adrenergic agonists noradrenaline, adrenaline, isoproterenol, salbutamol and, dobutamine were all effective in evoking a dispersion of melanophore pigment in the presence of phentolamine and verapamil or melatonin. The pigment-dispersing effect of noradrenaline (β₁-selective agonist) was inhibited by metoprolol (β₁-selective antagonist), propranolol, and butoxamine. Whereas, the effect of salbutamol (β₂-selective agonist) was hardly inhibited by metoprolol, though it was considerably inhibited by propranolol and ICI-118551. It was estimated that β₁- and β₂-adrenergic receptors coexist at ratios of 8.6:91.4, in the melanophore of Tridentiger trigonocephalus, and 25:75, in the melanophore of Chasmichthys gulosus, through the analyses of Hofstee plots of the effects of the β-adrenergic drugs. It was suggested that the relation between the pigment-dispersing effect of a β-adrenergic agonist on the melanophores and the concentration of the drug follows mass action kinetics, when the effect is mainly caused by the activation of β₂-adrenergic receptors of the melanophores. However, when it is mainly caused by the activation of β₁-adrenergic receptors of the melanophores, the relation does not follow mass action kinetics.     

Melatonin Signaling Controls the Daily Rhythm in Blood Glucose Levels Independent of Peripheral Clocks

Melatonin is rhythmically secreted by both the pineal gland and retina in a circadian fashion, with its peak synthesis occurring during the night. Once synthesized, melatonin exerts its effects by binding to two specific G-protein coupled receptors–melatonin receptor type 1(MT₁) and melatonin receptor type 2(MT₂). Recent studies suggest the involvement of MT₁ and MT₂ in the regulation of glucose homeostasis; however the ability of melatonin signaling to impart timing cues on glucose metabolism remains poorly understood. Here we report that the removal of MT₁ or MT₂ in mice abolishes the daily rhythm in blood glucose levels. Interestingly, removal of melatonin receptors produced small effects on the rhythmic expression patterns of clock genes within skeletal muscle, liver, and adipose tissue. Taken together, our data suggest that the loss of the daily rhythm in blood glucose observed in MT₁^-/- and MT₂^-/- mice does not occur as a consequence of ‘disrupted’ clocks within insulin sensitive tissues. Finally our results highlight a diurnal contribution of melatonin receptor signaling in the daily regulation of blood glucose levels.     

Human Gastroenteropancreatic Expression of Melatonin and Its Receptors MT1 and MT2

Background and Aim

The largest source of melatonin, according to animal studies, is the gastrointestinal (GI) tract but this is not yet thoroughly characterized in humans. This study aims to map the expression of melatonin and its two receptors in human GI tract and pancreas using microarray analysis and immunohistochemistry.

Method

Gene expression data from normal intestine and pancreas and inflamed colon tissue due to ulcerative colitis were analyzed for expression of enzymes relevant for serotonin and melatonin production and their receptors. Sections from paraffin-embedded normal tissue from 42 individuals, representing the different parts of the GI tract (n=39) and pancreas (n=3) were studied with immunohistochemistry using antibodies with specificity for melatonin, MT₁ and MT₂ receptors and serotonin.

Results

Enzymes needed for production of melatonin are expressed in both GI tract and pancreas tissue. Strong melatonin immunoreactivity (IR) was seen in enterochromaffin (EC) cells partially co-localized with serotonin IR. Melatonin IR was also seen in pancreas islets. MT₁ and MT₂ IR were both found in the intestinal epithelium, in the submucosal and myenteric plexus, and in vessels in the GI tract as well as in pancreatic islets. MT₁ and MT₂ IR was strongest in the epithelium of the large intestine. In the other cell types, both MT₂ gene expression and IR were generally elevated compared to MT₁. Strong MT₂, IR was noted in EC cells but not MT₁ IR. Changes in gene expression that may result in reduced levels of melatonin were seen in relation to inflammation.

Conclusion

Widespread gastroenteropancreatic expression of melatonin and its receptors in the GI tract and pancreas is in agreement with the multiple roles ascribed to melatonin, which include regulation of gastrointestinal motility, epithelial permeability as well as enteropancreatic cross-talk with plausible impact on metabolic control.     

Melatonin does not affect the black pigment migration in the crab Neohelice granulata

N-acetyl-5-methoxytryptamine or melatonin is a multifunctional molecule. The main physiological function, at least in vertebrates, is to transduce to the animal the photoperiodic information and regulate rhythmic parameters. But studies have also observed the action of this molecule on pigment migration in ectothermic vertebrates. Thus the aim of this paper was to investigate in vivo and in vitro the influence of melatonin on the pigment migration in melanophores of the crab Neohelice granulate. Injections of melatonin (2 × 10⁻⁹ moles · crab⁻¹) at 07:00 h or 19:00 h did not affect (p > 0.05) the circadian pigment migration of the melanophores in constant darkness. Additionally no significant pigment migration (p > 0.05) was verified in normal and eyestalkless crabs injected with melatonin (10⁻¹⁰–10⁻⁷ moles · crab⁻¹) during the day or night. In the ^{in vitro} assay, the response of melanophores to the pigment-dispersing hormone in eyestalkless crabs injected with melatonin (2 × 10⁻⁹ moles · crab⁻¹) 1 and 12 hours before the observations did not differ (p > 0.05) from the control group (injected with physiological solution). These results suggest that melatonin does not act as a signaling factor for pigment dispersion or aggregation in the melanophores of N. Granulate.     

Melanotropin receptors in the cartilaginous fish,Potamotrygon reticulatus and in the lungfish,Lepidosiren paradoxa

1. P. reticulatus skins in vitro exhibit punctate melanophores, and darken in response to the melanotropins in a dose-related manner.2. The relative potencies (Ac-Nle⁴-alpha-MSH_4–13-NH₂< alpha-MSH = Ac-Cys⁴,Cys¹⁰-alpha-MSH_4–13-NH₂) suggest that the supression of the amino-terminal tripeptide may affect the ideal peptide conformation for interaction with the receptor, which is restored with cyclization of the decapeptide fragment.3. Our data suggest that hormonal control of P. reticulatus pigment cells is mainly exerted by alpha-MSH, as none of the other agonists (isoproterenol, norepinephrine, MCH and melatonin) exhibited pigment dispersing or aggregating activities.4. L. paradoxa scales possess non-innervated epidermal and dermal melanophores, which exhibit stellate shape in vitro.5. Norepinephrine is a weak partial agonist in L. paradoxa epidermal melanophores, in the presence or in the absence of the beta-adrenoceptor blocker propranolol, suggesting that catecholamines do not play a role in the control of pigment cells.6. Both epidermal and dermal melanophore responses to MCH and to its analogue MCH_5–15, and to melatonin were also slow, partial and obtained with high doses of the hormones.7. The relative insensitivity of L. paradoxa melanophores is in accordance with the animal cryptic behavior, since it lives in muddy dark waters, and does not seem to depend on chromatic adaptation for camouflage.     

Melatonin attenuates the acetylcholine-induced contraction in isolated intestine of a teleost fish

The present study investigates the possible direct actions of melatonin (N-acetyl-5-methoxytryptamine) on intestinal motility in goldfish (Carassius auratus) using an in vitro system of isolated intestine in an organ bath engaged to an isometric transducer. The longitudinal strips from goldfish intestine in the organ bath showed a resting spontaneous myogenic rhythmic activity which is not altered by melatonin. The addition of acetylcholine (1 nmol l⁻¹–10 mmol l⁻¹) to the organ bath induces a significant contraction of the intestinal strips in a concentration-dependent manner. The addition of melatonin and its agonist, 2-iodomelatonin, induced a concentration-dependent attenuation of acetylcholine-induced contractile response. The specificity of this effect is tested by the preincubation of the intestine strips in the presence of two melatoninergic antagonists, luzindole (a non-selective MT₁/MT₂ melatonin receptor antagonist) and 4-P-PDOT (preferred antagonist of MT2 receptor subtype), which counteracted the melatonin-induced relaxation in a concentration-dependent manner. Finally, present results demonstrate that this melatoninergic effect on intestinal strips is a process highly dependent on extracellular calcium. In conclusion, this is the first study demonstrating the role of melatonin in the control of gut motility in a non-mammalian vertebrate. The melatonin effects on isolated intestine from goldfish are mediated by melatoninergic membrane receptors, and could suggest a delay in food transit time, supporting its anorectic effect reported on in vivo studies.     

The Presence and Localization of Melatonin Receptors in the Rat Aorta

Melatonin is involved in blood pressure modulation in rats and humans. Some of the effects of melatonin are presumably mediated via two G-protein-coupled receptors (MT₁ and MT₂), but the distribution of MT₁ and MT₂ in the cardiovascular system remains to be explored comprehensively. We investigated the expression of both the receptors in the rat aorta on mRNA level by RT-PCR and real time RT-PCR as well as on protein level via western blotting and immunofluorescence microscopy. We verified MT₁ mRNA expression in the rat aorta and demonstrated the absence of MT₂ mRNA in this vessel type. MT₁ receptors were confirmed also at the protein level, and surprisingly they were preferentially localized to the tunica adventitia. Since no daily changes in MT₁ mRNA expression were detected, we suppose that the circadian changes in circulating melatonin concentrations are sufficient to mediate circadian effects of melatonin in the aorta. The localization of MT₁ in the tunica adventitia suggests an influence of melatonin on vasa vasorum function and signal transduction in the aorta wall.     

MT2-like melatonin receptor modulates amplitude receptor potential in visual cells of crayfish during a 24-hour cycle

Retinular photoreceptors are structures involved in the expression and synchronization of the circadian rhythm of sensitivity to light in crayfish. To determine whether melatonin possesses a differential effect upon the receptor potential (RP) amplitude of retinular photoreceptors circadian time (CT)-dependent, we conducted experiments by means of applying melatonin every 2 h during a 24-hour cycle. Melatonin with 100 nM increased RP amplitude during subjective day to a greater degree than during subjective night. To determine whether MT₂ melatonin receptors regulate the melatonin-produced effect, we carried out two experiments, circadian times (CTs) 6 and 18, which included the following: (1) application of the MT₂ receptor selective agonist 8-M-PDOT and antagonist DH97, and (2) the specific binding of [¹²⁵I]-melatonin in eyestalk membranes. The amount of 10 nM of 8-M-PDOT increased RP amplitude in a similar manner to melatonin, and 1 nM DH97 abolished the increase produced by melatonin and 8-M-PDOT. Binding of [¹²⁵I]-melatonin was saturable and specific. Scatchard analysis revealed an affinity constant (K_d) of 1.1 nM and a total number of binding sites (B_max) of 6 fmol/mg protein at CT 6, and a K_d of 1.46 nM and B_max of 7 fmol/mg protein at CT 18. Our results indicate that melatonin increased RP amplitude of crayfish retinular photoreceptors through MT₂-like melatonin receptors. These data support the idea that melatonin is a signal of darkness for the circadian system in crayfish retinular cells.     

Role of the MT1 and MT2 melatonin receptors in mediating depressive‐ and anxiety‐like behaviors in C3H/HeN mice

Melatonin is a neurohormone primarily synthesized by the pineal gland following a circadian rhythm with a high level during the night and a low level during the day. Alterations in the synthesis and secretion of melatonin have been reported in various mood disorders, including major depressive disorder. However, the role of endogenous melatonin in the pathophysiology of depressive disorder is unclear. Melatonin primarily acts through two G protein‐coupled receptors, termed MT₁ and MT₂. The present study investigated the effect of genetic deletion of the MT₁ and/or MT₂ receptors on tests associated with depression‐ and anxiety‐like behaviors in C3H/HeN mice. Deletion of the MT₁ and/or MT₂ receptors caused a deficit in hedonic and social interaction behavior, and increased anxiety‐like behavior. It is likely that dysregulations of the MT₁ and/or MT₂ melatonin receptors could be involved in the pathophysiology of depression and anxiety.     

Distribution of MT1 Melatonin Receptor Promoter-Driven RFP Expression in the Brains of BAC C3H/HeN Transgenic Mice

The pineal hormone melatonin activates two G-protein coupled receptors (MT₁ and MT₂) to regulate in part biological functions. The MT₁ and MT₂ melatonin receptors are heterogeneously distributed in the mammalian brain including humans. In the mouse, only a few reports have assessed the expression of the MT₁ melatonin receptor expression using 2-iodomelatonin binding, in situ hybridization and/or polymerase chain reaction (PCR). Here, we described a transgenic mouse in which red fluorescence protein (RFP) is expressed under the control of the endogenous MT₁ promoter, by inserting RFP cDNA at the start codon of MTNR1a gene within a bacterial artificial chromosome (BAC) and expressing this construct as a transgene. The expression of RFP in the brain of this mouse was examined either directly under a fluorescent microscope or immunohistochemically using an antibody against RFP (RFP-MT₁). RFP-MT₁ expression was observed in many brain regions including the subcommissural organ, parts of the ependyma lining the lateral and third ventricles, the aqueduct, the hippocampus, the cerebellum, the pars tuberalis, the habenula and the habenula commissure. This RFP-MT₁ transgenic model provides a unique tool for studying the distribution of the MT₁ receptor in the brain of mice, its cell-specific expression and its function in vivo.     

Melatonin Signaling Modulates Clock Genes Expression in the Mouse Retina

Previous studies have shown that retinal melatonin plays an important role in the regulation of retinal daily and circadian rhythms. Melatonin exerts its influence by binding to G-protein coupled receptors named melatonin receptor type 1 and type 2 and both receptors are present in the mouse retina. Earlier studies have shown that clock genes are rhythmically expressed in the mouse retina and melatonin signaling may be implicated in the modulation of clock gene expression in this tissue. In this study we determined the daily and circadian expression patterns of Per1, Per2, Bmal1, Dbp, Nampt and c-fos in the retina and in the photoreceptor layer (using laser capture microdissection) in C3H-f+/+ and in melatonin receptors of knockout (MT₁ and MT₂) of the same genetic background using real-time quantitative RT-PCR. Our data indicated that clock and clock-controlled genes are rhythmically expressed in the retina and in the photoreceptor layer. Removal of melatonin signaling significantly affected the pattern of expression in the retina whereas in the photoreceptor layer only the Bmal1 circadian pattern of expression was affected by melatonin signaling removal. In conclusion, our data further support the notion that melatonin signaling may be important for the regulation of clock gene expression in the inner or ganglion cells layer, but not in photoreceptors.     

Melatonin Receptor-Mediated Inhibition of Cyclic AMP Accumulation in Chick Retinal Cell Cultures

Abstract: Melatonin receptors were characterized in cultured neurons and photoreceptors prepared from chick embryo retina. Cultured cells contained high-affinity 2-[¹²⁵I]iodomelatonin binding sites (K_D = 41.6 pM), similar to those in intact retina. The effects of melatonin and related indoles on cyclic AMP accumulation were examined. Melatonin (10^?7M) had no effect on basal or K⁺-stimulated cyclic AMP accumulation, but inhibited forskolin-stimulated cyclic AMP accumulation by approximately 50%. Melatonin inhibited forskolin-stimulated cyclic AMP accumulation in the presence or absence of the cyclic nucleotide phosphodiesterase inhibitor 3-isobutyl-1-methylxanthine, suggesting an effect on cyclic AMP synthesis rather than degradation. Half-maximal inhibition was observed at 5.9 × 10^?10M melatonin. The relative order of potency among melatonin analogues was 2-iodomelatonin > melatonin ≈ 6-chloromelatonin ≥ 6-hydroxymelatonin > N-acetylserotonin ≈ 5-methoxytryptophol > serotonin. The EC₅₀ value for inhibition of cyclic AMP accumulation by 2-iodomelatonin (36.7 pM) was comparable to the K_D value for binding of the radioligand, suggesting that the binding sites represent functional receptors. The inhibitory effect of melatonin was antagonized by the putative melatonin antagonists luzindole, N-acetyltryptamine, and N-(2,4-dinitrophenyl)-5-methoxytryptamine, with estimated K_B values of 0.12, 0.17, and 1 µM, respectively. At a concentration of 10 µM, N-(2,4-dinitrophenyl)-5-methoxytryptamine significantly inhibited forskolin-stimulated cyclic AMP accumulation when added alone; at 30 µM, luzindole and N-acetyltryptamine also had significant inhibitory effects. The inhibitory effect of melatonin was blocked by pretreatment with pertussis toxin. The results of this study indicate that melatonin receptors on retinal cells are coupled via inhibitory G proteins to cyclic AMP accumulation. Thus, some of the effects of melatonin on retinal physiology may be related to regulation of cyclic nucleotide metabolism.     

Synthesis of new N-(arylcyclopropyl)acetamides and N-(arylvinyl)acetamides as conformationally-restricted ligands for melatonin receptors

N-(Arylcyclopropyl)acetamides and N-(arylvinyl)acetamides or methyl ureas have been prepared as constrained analogues of melatonin. The affinity of these new compounds for chicken brain melatonin receptors and recombinant human MT₁ and MT₂ receptors was evaluated using 2-[¹²⁵I]-iodomelatonin as radioligand. Strict ethylenic or cyclopropyl analogues of the commercialized agonist agomelatine (Valdoxan®) were equipotent to agomelatine in binding bioassays. However, the ethylenic analogue was more effective than the cyclopropyl one in the melanophore aggregation bioassay, but was still less potent than the disubstituted 2,7-dimethoxy-naphtalenic compounds.     

O-linked melatonin dimers as bivalent ligands targeting dimeric melatonin receptors

A series of dimeric melatonin analogues 3a-e obtained by connecting two melatonin molecules through the methoxy oxygen atoms with spacers spanning 16–24 atoms and the agomelatine dimer 7 were synthesized and characterized in 2-[¹²⁵-I]-iodomelatonin binding assays, bioluminescence resonance energy transfer (BRET) experiments, and in functional cAMP and β-arrestin recruitment assays at MT₁ and MT₂ receptors. The binding affinity of 3a-e generally increased with increasing linker length. Bivalent ligands 3a-e increased BRET signals of MT₁ dimers up to 3-fold compared to the monomeric control ligand indicating the simultaneous binding of the two pharmacophores to dimeric receptors. Bivalent ligands 3c and 7 exhibited important changes in functional properties on the G_i/cAMP pathway but not on the β-arrestin pathway compared to their monomeric counterparts. Interestingly, 3c (20 atoms spacer) shows inverse agonistic properties at MT₂ on the G_i/cAMP pathway. In conclusion, these findings indicate that O-linked melatonin dimers are promising tools to develop signaling pathway-based bivalent melatonin receptor ligands.     

5-HT receptor subtypes as key targets in mediating pigment dispersion within melanophores of teleost,Oreochromis mossambicus

The presence of distinct class of 5-HT receptors in the melanophores of tilapia (Oreochromis mossambicus) is reported. The cellular responses to 5-HT (5-hydroxytryptamine), 5-HT₁, and 5-HT₂, agonists on isolated scale melanophores were observed with regard to pigment translocation within the cells. It was found that 5-HT exerted rapid and strong concentration dependent pigment granule dispersion within the melanophores. The threshold pharmacological dose of 5-HT that could elicit a measurable response was as low as 4.7 × 10^? 12 M/L. Selective 5-HT₁ and 5-HT₂ agonists, sumatriptan and myristicin were investigated and resulted in dose-dependent pigment dispersion. The dispersing effects were effectively antagonized by receptor specific antagonists. It is suggested that 5-HT-induced physiological effects are mediated via distinct classes of receptors that possibly participate in modulation of pigmentary responses of the fish.