So far, it has been difficult to generate embryonic stem (ES) cell from early stage preimplantation embryos of buffalo. These ES cells will be more helpful for efficient embryo cloning and generation of body cells as they are more primitive than inner cell mass (ICM)-derived ES cells. The present study was conducted to find the effect of lipopolysaccharide (LPS), melatonin (<em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>, a pineal gland product), and citral (3,7-dimethyl-2,6-octadienal and a retinoic acid synthesis blocker) on establishment of primary ES cell colonies, the comparative size of the ES cell colonies, and expression of pluripotent genes during extended period of culture in buffalo. Zona-free eight-cell stage in vitro fertilization (IVF) embryos were cultured in ES cell medium supplemented with none (media I as control), LPS (media II), citral melatonin (media III), or melatonin (media IV). The multiplication of blastomere leading to ES cell colony formation and expression of pluripotent genes were assessed up to day 20 of culture. The primary colony formation, the comparative size of the ES cell colonies, and expression of pluripotent genes in these colonies were better in the medium supplemented with melatonin in all days of culture. Within melatonin supplementation, the colony size was comparatively larger on day 8 and day 12 of culture. Further, with this supplementation, the Oct-4 and <em>N</em>anog expression was comparatively higher on all days of culture. The results indicated that supplementation of melatonin helped in the formation of better primary ES cell colony as well as in the maintenance of pluripotency. The results also indicated that primary colonies developed on day 8 to day 12 of culture may be better for passaging them for establishment of ES cell line from early stage preimplantation IVF embryos of in buffalo.

The i<em>n</em> vivo a<em>n</em>d i<em>n</em> vitro effects of melato<em>n</em>i<em>n</em> (<em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>) o<em>n</em> lipid peroxidatio<em>n</em> of lo<em>n</em>g chai<em>n</em> polyu<em>n</em>saturated fatty acids (PUFA) located i<em>n</em> rat liver microsomes were determi<em>n</em>ed. The effect of i<em>n</em>traperito<em>n</em>eal admi<em>n</em>istratio<em>n</em> of melato<em>n</em>i<em>n</em> (10 mg/kg weight/24 hr) o<em>n</em> ascorbate-Fe++ i<em>n</em>duced lipid peroxidatio<em>n</em> of isolated rat liver microsomes was first exami<em>n</em>ed. The ascorbate i<em>n</em>duced light emissio<em>n</em> i<em>n</em> hepatic microsomes was i<em>n</em>hibited by melato<em>n</em>i<em>n</em> treatme<em>n</em>t [co<em>n</em>trol group: 10.714 +/- 0.894; melato<em>n</em>i<em>n</em> group: 3.162 +/- 0.<em>5</em>1<em>5</em>, cou<em>n</em>ts per mi<em>n</em>ute (cpm) x 10(-<em>5</em>)]. Sig<em>n</em>ifica<em>n</em>t differe<em>n</em>ces i<em>n</em> the co<em>n</em>te<em>n</em>t of arachido<em>n</em>ic C20:4 <em>n</em>-6 a<em>n</em>d docosahexae<em>n</em>oic acid C22:6 <em>n</em>-3 were observed whe<em>n</em> co<em>n</em>trol microsomes were lipid peroxidized i<em>n</em> the prese<em>n</em>ce of ascorbic acid. These cha<em>n</em>ges were less pro<em>n</em>ou<em>n</em>ced i<em>n</em> liver microsomes isolated from melato<em>n</em>i<em>n</em> treated rats. I<em>n</em> vitro assays showed that after i<em>n</em>cubatio<em>n</em> of rat liver microsomes i<em>n</em> a<em>n</em> ascorbate-Fe++ system, at 37 degrees C for 210 mi<em>n</em>, the total cpm origi<em>n</em>ated from light emissio<em>n</em> (chemilumi<em>n</em>esce<em>n</em>ce) was fou<em>n</em>d to be lower i<em>n</em> those membra<em>n</em>es i<em>n</em>cubated i<em>n</em> the prese<em>n</em>ce of melato<em>n</em>i<em>n</em>. The fatty acid compositio<em>n</em> of total lipids isolated from rat liver microsomes was substa<em>n</em>tially modified whe<em>n</em> subjected to <em>n</em>o<em>n</em>e<em>n</em>zymatic lipid peroxidatio<em>n</em> with a co<em>n</em>siderable decrease of docosahexae<em>n</em>oic acid 22:6 <em>n</em>-3 a<em>n</em>d arachido<em>n</em>ic acid 20:4 <em>n</em>-6. The i<em>n</em>hibitio<em>n</em> of the lipid peroxidatio<em>n</em> process evaluated as chemilumi<em>n</em>esce<em>n</em>ce (total cpm at selected times) was melato<em>n</em>i<em>n</em> co<em>n</em>ce<em>n</em>tratio<em>n</em> depe<em>n</em>de<em>n</em>t. Melato<em>n</em>i<em>n</em>, at a co<em>n</em>ce<em>n</em>tratio<em>n</em> 1.2 mm, i<em>n</em>hibited almost completely the lipid peroxidatio<em>n</em> process. Arachido<em>n</em>ic a<em>n</em>d docosahexae<em>n</em>oic acids were more affected tha<em>n</em> docosape<em>n</em>tae<em>n</em>oic acid duri<em>n</em>g the lipid peroxidatio<em>n</em> process. <em>N</em>ot all fatty acids were equally protected after the additio<em>n</em> of melato<em>n</em>i<em>n</em> to the i<em>n</em>cubatio<em>n</em> medium. Our results i<em>n</em>dicate that melato<em>n</em>i<em>n</em> may act i<em>n</em> vivo a<em>n</em>d i<em>n</em> vitro as a<em>n</em> a<em>n</em>tioxida<em>n</em>t protecti<em>n</em>g lo<em>n</em>g chai<em>n</em> PUFA prese<em>n</em>t i<em>n</em> rat liver microsomes from the deleterious effect by a selective mecha<em>n</em>ism that reduces the loss of docosahexae<em>n</em>oic a<em>n</em>d arachido<em>n</em>ic acids.

The mass spectra of the four tryptamine derivatives, <em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em> (melatonin), <em>N</em>-<em>acetyl</em>-<em>5</em>-hydroxytryptamine (<em>N</em>-<em>acetyl</em>-serotonin), <em>N</em>,<em>N</em>-dimethyl-<em>5</em>-hydroxtryptamine (bufotenine) and <em>N</em>,<em>N</em>-dimethyl-<em>5</em>-<em>methoxytryptamine</em> (O-methylbufotenine), with specifically labeled [D4] aminoethyl sidechains have been measured. Comparison of these spectra with those of the unlabeled compounds enable the major fragmentations of the compounds to be defined.

Melatonin (<em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>) and serotonin <em>N</em>-<em>acetyl</em>transferase (<em>N</em>AT), a key regulatory enzyme in melatonin synthesis, are present in the adults and larvae of several insect species, as well as in vertebrates. To determine when melatonin and <em>N</em>AT first appear in insects ontogenetically, melatonin levels and <em>N</em>AT-like activity were measured in developing eggs of the cricket Gryllus bimaculatus. When the eggs were incubated under a 12-h light/12-h dark (LD) cycle at 24-26 degrees C, melatonin was detected in the egg extracts at all of the developmental stages examined. <em>N</em>AT-like activity was first found in the eggs 3 days after oviposition. From <em>5</em> to 11 days after oviposition, both <em>N</em>AT-like activity and melatonin levels showed significant day/night changes with the high levels occurring during the dark period of the LD cycle. By contrast, significant day/night changes were not detected in eggs just before hatching. To determine more detailed temporal changes, <em>N</em>AT-like activity was assayed in eggs 6 to 7 days after oviposition at 2- or 4-h intervals over a 48-h period. The activity in the eggs clearly exhibited a diurnal rhythm, peaking in the dark period of the LD cycle, and the rhythm persisted in constant darkness. These results suggest that the cricket egg (probably the embryo) synthesizes melatonin, and that its melatonin synthesis may fluctuate with a circadian rhythm. In addition, the results of the present study strongly suggest that a circadian clock controlling <em>N</em>AT activity functions in the cricket at the embryonic stage.

Serotonin (<em>5</em>-HT)-induced stimulation or progesterone (P4) production by bovine luteal cells was characterized with respect to the receptor subtype mediating this response, the steroidogenic response to <em>5</em>-HT metabolites, the role of adenylate cyclase, and the <em>5</em>-HT concentration of bovine luteal tissue. Addition of <em>5</em>-HT (10(-<em>5</em>) M) stimulated the production of P4 (P less than 0.0<em>5</em>) and this stimulation was inhibited by the <em>5</em>-HT antagonist mianserin at a concentration of 10(-<em>5</em>) M (P less than 0.0<em>5</em>), but not at a mianserin concentration of 10(-7) M. Additionally, the response to <em>5</em>-HT could not be inhibited by ketanserin (10(-<em>5</em>) M), a <em>5</em>-HT2 receptor antagonist. Incubation of luteal cells with a specific <em>5</em>-HT1 agonist, (+/-)-8-hydroxydipropylaminotetralin HBr (DPAT) (10(-4) M), stimulated the production of P4 (P less than 0.0<em>5</em>) and this response could not be blocked by mianserin at 10(-7) M or by ketanserin, but was inhibited by mianserin at 10(-<em>5</em>) (P less than 0.0<em>5</em>). The addition of the <em>5</em>-HT metabolite <em>5</em>-<em>methoxytryptamine</em> (<em>5</em>-MTA) stimulated P4 production (P less than 0.0<em>5</em>) and this response could be inhibited by mianserin (10(-<em>5</em>) M, P less than 0.0<em>5</em>). <em>N</em>either, <em>N</em>-<em>acetyl</em>-<em>5</em>-HT nor <em>5</em>-methoxytryptophan significantly affected P4 production. The addition of the phosphodiesterase inhibitor 3-isobutyl-methylxanthine (IBMX, 0.1 mM) potentiated the effects of <em>5</em>-HT and DPAT (P less than 0.0<em>5</em>), but this effect was additive rather than synergistic. In contrast, the addition of luteinizing hormone (10 ng/ml) plus IBMX resulted in a significant synergistic response (P less than 0.0<em>5</em>).(ABSTRACT TRU<em>N</em>CATED AT 2<em>5</em>0 WORDS)

Melatonin (<em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>) and its immediate precursor <em>N</em>-<em>acetyl</em> serotonin in the metabolism of tryptophan are free radical scavengers that have been found to protect against non-enzymatic lipid peroxidation in many experimental models. By contrast, little is known about the antioxidant ability of these indoleamines against <em>N</em>ADPH enzymatic lipid peroxidation. The light emission produced by rat-liver microsomes, expressed as total cpm during 180 min of incubation at 37 degrees C, was two-fold greater in the presence of ascorbate (0.4mM) when compared with <em>N</em>ADPH (0.2 mM). Maximal peaks of light emission produced by microsomes lipid peroxidized with ascorbic-Fe(2+) or <em>N</em>ADPH and expressed as cpm were 3<em>5</em>4,208 (at 60 min) and 13<em>5</em>,800 (at 1<em>5</em> min), respectively. During non-enzymatic lipid peroxidation a decrease of total chemiluminescence (inhibition of lipid peroxidation) was observed when increasing concentrations of melatonin were added to liver microsomes. The protective effect was concentration-dependent. The inhibition observed in light emission was coincident with the protection of the most PUFAs. Preincubation of microsomes with <em>N</em>-<em>acetyl</em> serotonin reduced these changes very dramatically. Thus, in the presence of both antioxidants (0.36, 0.7<em>5</em>, 1.<em>5</em> mM), light emission percent inhibition during non-enzymatic (ascorbate-Fe(2+)) lipid peroxidation of rat liver microsomes was for melatonin: 6.12, 16.20, 34.88 and for <em>N</em>-<em>acetyl</em> serotonin: 8<em>5</em>.10, 88.48, 84.4 respectively. The incubation of rat liver microsomes in the presence of <em>N</em>ADPH (0.36, 0.7<em>5</em>, 1.<em>5</em> mM) produce a sudden increase of chemiluminescence that gradually increased and reached a maximal value at about 1<em>5</em> min; however, <em>N</em>-<em>acetyl</em> serotonin reduced these changes very efficiently.

Melatonin (Mel), or <em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>, is a circadian hormone that can diffuse through all the biological membranes thanks to its amphiphilic structure, also overcoming the blood-brain barrier and placenta. Although Mel has been reported to exhibit strong antioxidant properties in healthy tissues, studies carried out on tumor cultures gave a different picture of its action, often describing Mel as effective to trigger the cell death of tumor cells by enhancing oxidative stress. Based on this premise, here Mel effect was investigated using a tumor cell line representative of the human alveolar rhabdomyosarcoma (ARMS), the most frequent soft tissue sarcoma affecting childhood. For this purpose, Mel was given either dissolved in ethanol (EtOH) or dimethyl sulfoxide (DMSO) at different concentrations and time exposures. Cell viability assays and ultrastructural observations demonstrated that Mel was able to induce a dose- and time-dependent cell death independently on the dissolution solvent. Microscopy analyses highlighted the presence of various apoptotic and necrotic patterns correlating with the increasing Mel dose and time of exposure. These findings suggest that Mel, triggering apoptosis in ARMS cells, could be considered as a promising drug for future multitargeted therapies.

OBJECTIVE

Melatonin (<em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>) (MT) is a hormone that acts as an antioxidant. It is produced by the pineal gland and within the retina; its release is blocked by light entering the eye. We examined whether MT daytime levels differ between pseudophakic patients with age-related macular degeneration (ARMD) and pseudophakic subjects without any ocular pathology of the same age.

METHODS

A prospective, cross-sectional, observational study was performed. Pseudophakic patients of the same age group were included. Patients underwent complete ophthalmic examinations and blood sampling between 08:00 and 10:00 hr. MT daytime value in the serum was the main outcome measure.

RESULTS

Sixty-nine pseudophakic patients were included. Fifty patients with exudative and non-exudative ARMD were in the study group while 19 patients were controls. Patients with ARMD had significantly higher daytime levels of MT (P = 0.003). There were significant differences in MT daytime levels between the exudative and non-exudative forms (P = 0.009). MT values also correlated with the best-corrected visual acuity (r = -0.28<em>5</em>, P = 0.019).

CONCLUSIONS

These data indicate that pseudophakic patients with ARMD produce more MT during the day compared to pseudophakic subjects without ARMD. This may be caused by the reduced visual acuity in patients with ARMD, whereby less light reaches the photoreceptors, allowing MT secretion to continue during the day. Because MT also acts as an antioxidant and daytime levels are higher in patients with ARMD, these results might be interpreted as a rescue factor.

Reproduction in most fish is seasonal or periodic, and the spawning occurs in an appropriate season to ensure maximum survival of the offspring. The sequence of reproductive events in an annual cycle is largely under the control of a species-specific endogenous timing system, which essentially relies on a well-equipped physiological response mechanism to changing environmental cues. The duration of solar light or photoperiod is one of the most predictable environmental signals used by a large number of animals including fish to coordinate their seasonal breeding. In vertebrates, the pineal gland is the major photoneuroendocrine part of the brain that rhythmically synthesizes and releases melatonin (<em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>) into the circulation in synchronization with the environmental light-dark cycle. Past few decades witnessed an enormous progress in understanding the mechanisms by which melatonin regulates seasonal reproduction in fish and in other vertebrates. Most studies emphasized hormonal actions of melatonin through its high-affinity, pertussis toxin-sensitive G-protein (guanine nucleotide-binding protein)-coupled receptors on the hypothalamus-pituitary-gonad (HPG) axis of fish. However, the discovery that melatonin due to its lipophilic nature can easily cross the plasma membrane of all cells and may act as a potent scavenger of free radicals and stimulant of different antioxidants added a new dimension to the idea explaining mechanisms of melatonin actions in the regulation of ovarian functions. The basic concept on the actions of melatonin as an antioxidant emerged from mammalian studies. Recently, however, some new studies clearly suggested that melatonin, apart from playing the role of a hormone, may also be associated with the reduction in oxidative stress to augment ovarian functions during spawning. This review thus aims to bring together the current knowledge on the role of melatonin as a hormone as well as an antioxidant in the control of fish reproduction and shape the current working hypotheses supported by recent findings obtained in carp or based on knowledge gathered in mammalian and avian species. In essence, this review highlights potential actions of melatonin as a hormone in determining temporal pattern of spawning and as an antioxidant in regulating oocyte maturation at the downstream of HPG axis in fish.

Melatonin (<em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>) is a highly pleiotropic hormone with antioxidant, antiproliferative, oncolytic and neuroprotective properties. Here, we present evidence that the <em>N</em>-<em>acetyl</em> side chain plays a key role in melatonin's antiproliferative effect in HT22 and sw-13<em>5</em>3 cells, but it does so at the expense of antioxidant and neuroprotective properties. Removal of the <em>N</em>-<em>acetyl</em> group enhances the antioxidant and neuroprotective properties of the indole, but it can lead to toxic methamphetamine-like effects in several cell lines. Inhibition of <em>N</em>FkB mimicked melatonin's antiproliferative and antioxidant effects, but not neuroprotection. Our results strongly suggest that neuroprotective and antiproliferative effects of melatonin rely on different parts of the molecule and are likely mediated by different mechanisms. We also predict that melatonin metabolism by target cells could determine whether melatonin inhibits cell proliferation, prevents toxicity or induces cell death (e.g. apoptosis or autophagy). These observations could have important implications for the rational use of melatonin in personalized medicine.

Melatonin (<em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>), an indole hormone, regulates a variety of important central and peripheral actions related to circadian rhythms and reproduction through its specific receptor subtypes (Mel-1a, Mel-1b and Mel-1c). However, the expression profile of melatonin receptor genes (MT<em>N</em>R1A, MT<em>N</em>R1B and MT<em>N</em>R1C) in ovarian hierarchical follicles of geese remains to be clarified. In this study, the expression level of melatonin receptors in small white follicle (SWF), small yellow follicle (SYF), the largest follicle (F1), second largest (F2), third largest (F3), fourth largest (F4), fifth largest (F<em>5</em>), and postovulatory follicle (POF) in the Sichuan white goose were examined using quantitative real-time PCR (qRT-PCR). The results showed that the expression levels of MT<em>N</em>R1A, MT<em>N</em>R1B and MT<em>N</em>R1C initially increased and later decreased. The highest levels of gene expression of these receptor subtypes were observed in F<em>5</em> or F4 in all examined follicles. Furthermore, the expression of MT<em>N</em>R1A and MT<em>N</em>R1B mR<em>N</em>A was significantly greater in SYF compared with SWF (P<0.0<em>5</em>), but MT<em>N</em>R1C was absent in SWF. The expression of MT<em>N</em>R1A, MT<em>N</em>R1B and MT<em>N</em>R1C mR<em>N</em>A was significantly greater in F<em>5</em> compared with SYF (P<0.0<em>5</em>), and the expression of MT<em>N</em>R1A and MT<em>N</em>R1C mR<em>N</em>A was higher in F1 compared with POF (P<0.0<em>5</em>). In addition, the oestrogen concentration in SWF, SYF, F4, F3, F2, F1 and POF was measured using ELISA. The oestrogen concentration and melatonin receptor expression both were initially observed to increase and subsequently decrease. The oestrogen concentration in F4 and F3 was highest in all examined samples and was 1318.2pg/g and 1318.1pg/g, respectively. These results suggest that the melatonin receptor may be involved in the activation of the SWF and SYF to allow the SWF and SYF to develop into the subsequent follicles. Furthermore, follicles and the expression of the melatonin receptors may be regulated by the secretion of the oestrogen.

The aim of this study was to synthesize selective ligands for melatoninergic subtype receptors that could elucidate the physiological role of melatonin (<em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>, 1). So, we first investigated the role of a nitro substituent in the 4-, 6-, or 7-position of the indole heterocycle. Comparatively to melatonin, its analogues that nitrated in the 6- or 7-position (6 and 22) lose MT(3) but retain good MT(1) and MT(2) affinities, whereas the 4-nitro isomer (<em>5</em>) shows very high affinity (nanomolar) and selectivity for the MT(3) binding sites. <em>N</em>-Methylation of the indole nucleus of compound <em>5</em> potentiates these effects and affords the most potent and selective MT(3) ligand (17). The 2-iodo derivatives (12 and 10) of compounds <em>5</em> and 17 have also been synthesized to evaluate their binding profile with a view to further develop MT(3) selective radioligands.

Procedures were developed for the determination of 17 circulating amines using gas chromatography-negative ion chemical ionisation mass spectrometry. The amines were quantified against their appropriate deuterated isotopomers. The mean concentrations and ranges of catecholamines and trace amines were high compared with previous studies. In comparison with nonhypertensives, plasma from hypertensives had higher concentrations of the following amines: noradrenaline (t = 4.0%); normetanephrine (t = 6.1%) and metanephrine (t = 1.9%). There were no significant differences between <em>5</em>HT levels in plasma from hypertensives and controls. The following trace amines could be detected in variable amounts in plasma: p-tyramine, m-tyramine, p-octopamine, m-octopamine, p-synephrine, m-synephrine, and salsolinol. The trace amines melatonin, <em>N</em>-<em>acetyl</em> <em>5</em>HT, tryptamine, 6-hydroxymelatonin and <em>5</em>-<em>methoxytryptamine</em> could not be detected in plasma with limits of detection lying in the range 20-100 pg ml-1.

6-Methoxy-1,2,3,4-tetrahydro-β-carboline (pinoline) and <em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em> (melatonin) are both structurally related to <em>5</em>-hydroxytryptamine (serotonin). Here we describe the design, synthesis, and characterization of a series of melatonin rigid analogues resulting from the hybridization of both pinoline and melatonin structures. The pharmacological evaluation of melatonin-pinoline hybrids comprises serotonergic and melatonergic receptors, metabolic enzymes (monoamine oxidases), antioxidant potential, the in vitro blood-brain barrier permeability, and neurogenic studies. Pinoline at trace concentrations and 2-<em>acetyl</em>-6-methoxy-1,2,3,4-tetrahydro-β-carboline (2) were able to stimulate early neurogenesis and neuronal maturation in an in vitro model of neural stem cells isolated from the adult rat subventricular zone. Such effects are presumably mediated via serotonergic and melatonergic stimulation, respectively.

Aging is characterized by a progressive deterioration in physiological functions and metabolic processes. The loss of cells during aging in vital tissues and organs is related to several factors including oxidative stress and inflammation. Skeletal muscle degeneration is common in elderly people; in fact, this tissue is particularly vulnerable to oxidative stress since it requires large amounts of oxygen, and thus, oxidative damage is abundant and accumulates with increasing age. Melatonin (<em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>) is a highly efficient scavenger of reactive oxygen species and it also exhibits beneficial anti-inflammatory and anti-aging effects. This study investigated the susceptibility of rat L6 skeletal muscle cells to an induced oxidative stress following their exposure to hydrogen peroxide (<em>5</em>0 μM) and evaluating the potential protective effects of pre-treatment with melatonin (10 nM) compared to the known beneficial effect of alpha-lipoic acid (300 μM). Hydrogen peroxide-induced obvious oxidative stress; it increased the expression of tumour necrosis factor-alpha and in turn promoted nuclear factor kappa-B and overrode the endogenous defence mechanisms. Conversely, pre-treatment of the hydrogen peroxide-exposed cells to melatonin or alpha-lipoic acid increased endogenous antioxidant enzymes, including superoxide dismutase-2 and heme oxygenase-1; moreover, they ameliorated significantly oxidative stress damage and partially reduced alterations in the muscle cells, which are typical of aging. In conclusion, melatonin was equally effective as alpha-lipoic acid; it exhibited marked antioxidant and anti-aging effects at the level of skeletal muscle in vitro even when it was given in a much lower dose than alpha-lipoic acid.

Melatonin (<em>N</em>-<em>Acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>) is a hormone secreted mainly by the pineal gland or epiphyse and in smaller amounts by the retina. It is biosynthesized from tryptophan, the critical enzymatic step depends upon <em>N</em>-<em>Acetyl</em>-transferase (<em>N</em>AT). The circadian rhythm of melatonin is the same in man and all the laboratory animals studied until now with nocturnal plasma concentrations 3-10 times greater than during daytime. The secretion and release of melatonin depend upon a large number of exogenous and endogenous factors as e.g. sex, age, pubertal stage, menstrual cycle, drugs, season.... Light is the major regulating factor which acts through the retino-hypothalamic tract. Melatonin is considered as a transducer of the light signal forwarding to the organism the information about day length (relative length of day and night). It is a time-clue provider used by the organism to adapt itself to its environment.

Melatonin, or <em>N</em>-<em>acetyl</em> <em>5</em>-<em>methoxytryptamine</em>, a neurohormone produced in the pineal gland during periods of darkness, plays a key role in the regulation of circadian and seasonal biological rhythms. In mammals, specific MT1 and MT2 receptors are located in the central nervous system, mainly in suprachiasmatic nuclei, and also in a number of peripheral sites. Besides its chronobiotic action on light-dependant functions, such as sleep/waking alternance or seasonal depression, melatonin exerts modulatory effects on immune, endocrine and metabolic functions. However, its short half-life and extensive metabolism lead to a poor bioavailability. This prompted to search for metabolically stable analogs displaying new and innovative properties. The S 20098 compound, a melatoninergic agonist, has proven potent antidepressive and anxiolytic actions. The S 20928 compound, a melatonin antagonist, was shown to enhance basal metabolism and reduce weight gain. Thus, both of these melatonin derivatives open perspectives for the development of innovative therapeutic agents in the fields of depression and obesity.

Melatonin "<i><em>N</em></i>-<em>Acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>" (MT) has recently been considered as a new plant growth regulator with multiple physiological functions. Although many previous studies have confirmed that exogenous applied-MT can alleviate the deleterious effects of drought stress in many plant species, most of these studies were exclusive on seeds, seedlings, and young plants for a short period of their life cycles. Therefore, the knowledge of using MT as a potential promising agricultural foliar application to improve crop productivity and quality is still insufficient under adverse open field conditions. In this study, we investigated the effect of MT as a foliar application at 0, 20, and 40 ppm on tomato plants that were grown in the open field under the long term of optimal and deficit irrigation conditions. The results indicated that exogenous MT significantly enhanced plant growth, chlorophyll and activities of antioxidant enzymes, including ascorbate peroxidase (APX), catalase (CAT), and peroxidase (POX). This improvement was associated with a marked reduction in proline and soluble sugars. In addition, applied-MT worked as a protective agent against oxidative damage by reducing the cellular content of toxic substances such as H₂O₂ and malondialdehyde (MDA). Similarly, MT-treated plants showed greater total fruit yield with improving its quality attributes like total soluble solids (TSS), ascorbic acid, and lycopene. Generally, the highest significant fruit yield either under well-watered (13.7%) or water deficit (37.4%) conditions was achieved by the treatment of 20 ppm MT. These results indicate that exogenous MT played an essential role in enhancing tomato tolerance to deficit irrigation and could be recommended as a promising agricultural treatment under such conditions.

Keywords: Melatonin; Solanum lycopersicum L.; antioxidant enzymes; drought; quality attributes.

OBJECTIVE

To examine the effects of <em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em> (melatonin) on radiation-induced inner ear damage.

METHODS

An experimental animal model.

METHODS

Forty rats were randomized into five groups, as follows: 1) melatonin and then radiotherapy group (n = 8), which received intraperitoneal (i.p.) melatonin (<em>5</em> mg/kg) followed by irradiation 30 minutes later; 2) radiotherapy and then melatonin group (n = 8), which received irradiation with i.p. melatonin (<em>5</em> mg/kg) 30 minutes later; 3) melatonin group (n = 8), which received i.p. melatonin (<em>5</em> mg/kg); 4) radiotherapy group (n = 8), which underwent only irradiation; <em>5</em>) and the control group (n = 8), which received i.p. 0.9% <em>N</em>aCl. The medications and irradiation were administered for <em>5</em> days. All rats underwent the distortion product otoacoustic emission (DPOAE) test before and 10 days after the experiment. The middle ears of the rats were excised, and assessment of tissue alterations in the organs of Corti, spiral ganglions, and stria vascularis were compared among the groups.

RESULTS

In the radiotherapy group, the DPOAE amplitudes at frequencies of 4000 to 6000 Hz were significantly decreased when compared with the controls. The DPOAE amplitudes both in the melatonin and then radiotherapy group and the radiotherapy and then melatonin group exhibited better values than they did in the radiotherapy group. Histopathological evidence of damage to the organs of Corti, spiral ganglions, and stria vascularis damage was markedly reduced in both these two groups when compared to the radiotherapy group.

CONCLUSIONS

These results indicate that melatonin may have significant ameliorative effects on cochlear damage secondary to ionizing radiation.

OBJECTIVE

Metabolomics may unravel global metabolic changes in response to environmental exposures and identify important biological pathways involved in the pathophysiology of childhood obesity. Phthalate has been considered an obesogen and contributing to overweight and obesity in children. The purpose of this study is to evaluate changes in urine metabolites in response to the environmental phthalate exposure among overweight or obese children, and to investigate the metabolic mechanisms involved in the obesogenic effect of phthalate on children at puberty.

METHODS

Within the national Puberty Timing and Health Effects in Chinese Children (PTHEC) study, 69 overweight/obese children and 80 normal weight children were selected into the current study according to their puberty timing and WGOC (The Working Group for obesity in China) references. Urinary concentrations of six phthalate monoesters (MMP, MEP, MnBP, MEHP, MEOHP and MEHHP) were measured using API 2000 electrospray triple quadrupole mass spectrometer (ESIMS/MS). Metabolomic profiling of spot urine was performed using gas chromatography-mass spectrometry. Differentially expressed urinary metabolites associated with phthalate monoesters exposure were examined using orthogonal partial least square-discriminant analysis and multiple linear regression models. In addition, the candidate metabolites were regressed to obesity indices with multiple linear regression models and logistic regression models in all subjects.

RESULTS

Compared with <em>n</em>ormal weight childre<em>n</em>, higher levels of M<em>n</em>BP were detected i<em>n</em> uri<em>n</em>ary samples of childre<em>n</em> with overweight a<em>n</em>d obesity. After adjusti<em>n</em>g for co<em>n</em>fou<em>n</em>ders i<em>n</em>cludi<em>n</em>g chro<em>n</em>ological age, ge<em>n</em>der, puberty o<em>n</em>set, daily e<em>n</em>ergy i<em>n</em>take a<em>n</em>d physical activity a<em>n</em>d socio-eco<em>n</em>omic level, positive associatio<em>n</em> remai<em>n</em>ed betwee<em>n</em> uri<em>n</em>ary M<em>n</em>BP co<em>n</em>ce<em>n</em>tratio<em>n</em> a<em>n</em>d childhood overweight/obesity [OR = 1.<em>5</em>86, 9<em>5</em>% CI:1.043,2.412]. We observed elevated M<em>n</em>BP co<em>n</em>ce<em>n</em>tratio<em>n</em> was sig<em>n</em>ifica<em>n</em>tly correlated with i<em>n</em>creased levels of mo<em>n</em>osteari<em>n</em>, 1-mo<em>n</em>opalmiti<em>n</em>, stearic acid, itaco<em>n</em>ic acid, glycerol 3-phosphate, <em>5</em>-<em>methoxytryptamine</em>, kyotorphi<em>n</em>, 1-methylhyda<em>n</em>toi<em>n</em>, d-ala<em>n</em>yl-d-ala<em>n</em>i<em>n</em>e, pyrrole-2-carboxylic acid, 3,4-Dihydroxyphe<em>n</em>ylglycol, a<em>n</em>d butyraldehyde. Mea<em>n</em>while, i<em>n</em>creased M<em>n</em>BP co<em>n</em>ce<em>n</em>tratio<em>n</em> was also sig<em>n</em>ifica<em>n</em>tly correlated with decreased levels of lactate, glucose 6-phosphate, d-fructose 6-phosphate, palmitic acid, 4-acetamidobutyric acid, l-glutamic acid, <em>n</em>-<em>acetyl</em>-l-phe<em>n</em>ylala<em>n</em>i<em>n</em>e, imi<em>n</em>odiacetic acid, hydroxyproli<em>n</em>e, pipecoli<em>n</em>ic acid, l-or<em>n</em>ithi<em>n</em>e, <em>n</em>-<em>acetyl</em>-l-glutamic acid, gua<em>n</em>osi<em>n</em>e, cytosi<em>n</em>, a<em>n</em>d (s)-ma<em>n</em>delic acid i<em>n</em> the <em>n</em>ormal weight subjects. The observatio<em>n</em>s i<em>n</em>dicated that M<em>n</em>BP exposure was related to global uri<em>n</em>e metabolic ab<em>n</em>ormalities characterized by disrupti<em>n</em>g argi<em>n</em>i<em>n</em>e a<em>n</em>d proli<em>n</em>e metabolism a<em>n</em>d i<em>n</em>creasi<em>n</em>g oxidative stress a<em>n</em>d fatty acid reesterificatio<em>n</em>. Amo<em>n</em>g the metabolic markers related to M<em>n</em>BP exposure, 1-methylhyda<em>n</em>toi<em>n</em>, pyrrole-2-carboxylic acid a<em>n</em>d mo<em>n</em>osteari<em>n</em> were fou<em>n</em>d to be positively correlated with obesity i<em>n</em>dices, while hydroxyproli<em>n</em>e, l-or<em>n</em>ithi<em>n</em>e, a<em>n</em>d lactate were <em>n</em>egatively associated with overweight/obesity i<em>n</em> childre<em>n</em>.

CONCLUSIONS

Our results suggested that the disrupted arginine and proline metabolism associated with phthalate exposure might contribute to the development of overweight and obesity in school-age children, providing insights into the pathophysiological changes and molecular mechanisms involved in childhood obesity.

Gastric cancer accounts 8% of the total cancer cases leading to 10% of total cancer deaths worldwide. The indoleamine <em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>, better known as melatonin, is the principal hormone produced by the pineal gland. Recently, it has been well documented some anti-cancer roles of melatonin in some malignancies as breast and colon cancer; as well as some its protective roles in the GI tract that have been known as free radical scavenger, antimitogenic and apoptotic properties. According to the anti-cancer effects of melatonin, wide distribution of this neurohormone in GI tract and some proposed physiologic and pharmacologic roles for this neurohormone and following our previous study which has shown expression of MT2 receptor in gastric adenocarcinoma, this study initially scheduled to determine the expression of melatonin receptor MT1 in tissue samples of adenocarcinoma cancer patients. A total of 10 gastric adenocarcinoma patients and 10 normal individuals were examined for MT1 gene expression by real-time PCR. Additionally, for screening of different alleles of MT1 in our samples, the SSCP-PCR procedure was developed. Our results have shown interestingly high expression for MT1 receptor in cancer and marginal cancer groups comparing with normal group. Our findings also have shown that a remarkable association between MT1 receptor mR<em>N</em>A levels and grade in individuals over age <em>5</em>0. PCR-SSCP analysis results showed a variation between individuals which may be effective on their gene expression patterns. According to our knowledge, for the first time this study evaluated the expression of MT1 receptor gene in gastric adenocarcinoma tissues which consistent with our previous study but with some difference in comparisons between kind of tissue expression and difference in polymorphisms. Moreover, these results show the defending role of melatonin in the GI system.

Melatonin receptors (MT₁ and MT₂) transduce inhibitory signaling by melatonin (<em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>), which is associated with sleep induction and circadian rhythm modulation. Although recently reported crystal structures of ligand-bound MT₁ and MT₂ elucidated the basis of ligand entry and recognition, the ligand-induced MT₁ rearrangement leading to G_i-coupling remains unclear. Here we report a cryo-EM structure of the human MT₁-G_i signaling complex at 3.3 Å resolution, revealing melatonin-induced conformational changes propagated to the G-protein-coupling interface during activation. In contrast to other G_i-coupled receptors, MT₁ exhibits a large outward movement of TM6, which is considered a specific feature of G_s-coupled receptors. Structural comparison of G_i and G_s complexes demonstrated conformational diversity of the C-terminal entry of the G_i protein, suggesting loose and variable interactions at the end of the α<em>5</em> helix of G_i protein. These notions, together with our biochemical and computational analyses, highlight variable binding modes of Gα_i and provide the basis for the selectivity of G-protein signaling.

The levels of <em>N</em>-<em>acetyl</em>seroto<em>n</em>i<em>n</em> a<em>n</em>d melato<em>n</em>i<em>n</em> (<em>n</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>) i<em>n</em> the rat brai<em>n</em> were studied. Male rats were housed u<em>n</em>der a photoperiod of 12L:12D. After 2 weeks, the rats were sacrificed i<em>n</em> light or i<em>n</em> dark at 4-hour i<em>n</em>tervals rou<em>n</em>d the clock. Brai<em>n</em> melato<em>n</em>i<em>n</em> a<em>n</em>d <em>N</em>-<em>acetyl</em>seroto<em>n</em>i<em>n</em> were extracted by chloroform at greater tha<em>n</em> pH 10 a<em>n</em>d ethyl acetate at less tha<em>n</em> pH 3 respectively a<em>n</em>d qua<em>n</em>tified by radioimmu<em>n</em>oassay. There was a circadia<em>n</em> rhythm of brai<em>n</em> melato<em>n</em>i<em>n</em> (F less tha<em>n</em> 0.0<em>5</em>) with a <em>n</em>adir at 1600 h a<em>n</em>d a ze<em>n</em>ith at 0400 h. Brai<em>n</em> <em>N</em>-<em>acetyl</em>seroto<em>n</em>i<em>n</em>, u<em>n</em>like brai<em>n</em> melato<em>n</em>i<em>n</em>, demo<em>n</em>strated <em>n</em>o diur<em>n</em>al variatio<em>n</em>. Effects of pi<em>n</em>ealectomy o<em>n</em> brai<em>n</em> <em>N</em>-<em>acetyl</em>seroto<em>n</em>i<em>n</em> a<em>n</em>d melato<em>n</em>i<em>n</em> were also studied. Male rats were housed as <em>n</em>oted. After 2 weeks, they were pi<em>n</em>ealectomized or shamoperated. The a<em>n</em>imals were killed i<em>n</em> light or i<em>n</em> dark 1 week after operatio<em>n</em>. The levels of brai<em>n</em> melato<em>n</em>i<em>n</em> at 2400 h a<em>n</em>d 0400 h were sig<em>n</em>ifica<em>n</em>tly lowered (P less tha<em>n</em> 0.0<em>5</em>) followi<em>n</em>g pi<em>n</em>eal removal. Pi<em>n</em>ealectomy, however, has <em>n</em>o observable effect o<em>n</em> the level of brai<em>n</em> <em>N</em>-<em>acetyl</em>seroto<em>n</em>i<em>n</em>. Our fi<em>n</em>di<em>n</em>gs suggest that 1) there is a diur<em>n</em>al rhythm of brai<em>n</em> melato<em>n</em>i<em>n</em>, 2) the elevatio<em>n</em> of brai<em>n</em> melato<em>n</em>i<em>n</em> i<em>n</em> the dark period is depe<em>n</em>de<em>n</em>t o<em>n</em> the pi<em>n</em>eal, 3) brai<em>n</em> <em>N</em>-<em>acetyl</em>seroto<em>n</em>i<em>n</em> has <em>n</em>o diur<em>n</em>al variatio<em>n</em>, a<em>n</em>d 4) brai<em>n</em> <em>N</em>-<em>acetyl</em>seroto<em>n</em>i<em>n</em> is <em>n</em>ot affected by pi<em>n</em>ealectomy.

A simple capillary GC-MS method for the determination of melatonin from tablets is proposed. Melatonin (<em>N</em>-<em>acetyl</em>-<em>5</em>-<em>methoxytryptamine</em>) was determined by gas chromatography-mass spectrometry after extraction from ground tablets with ethyl acetate and derivatization with <em>N</em>-methyl-<em>N</em>-trimethylsilyl-heptafluorobutyramide (MTSHFBA). Splitless injection offers good reproducibility with a standard deviation of +/- 2%. The proposed method was applied to analyse the melatonin content from three different tablet formulations.