Comparative aspects of the pineal/melatonin system of poikilothermic vertebrates

Abstract: The pineal gland of poikilothermic vertebrates originates as an evagination from the diencephalic roof between the habenular and the posterior commissures, and associates with a parapineal organ to form the so-called pineal complex. The pinealocytes may be photosensitive, secretory or intermediate cells between both. Melatonin, the indoleamine secreted by the pineal, exhibits a circadian secretory rhythm that conveys environmental information to the organism. The peak melatonin secretion occurs during the night, although there are a few examples of an increase in indoleamine secretion during the day. Melatonin is also synthesized in other sites such as the retina, and it has been found in many invertebrates and unicellular organisms. The rhythmic secretory pattern of melatonin is responsible for many biological rhythms exhibited by lower vertebrates. These rhythms are abolished by pinealectomy in some species, but not in others, suggesting the existence of an extra-pineal pacemaker. The photoperiod and the temperature (especially in reptiles) are the main environmental factors affecting the secretory rhythm of melatonin. Poikilothermic vertebrates exhibit a circadian rhythmic color change, with nocturnal blanching, usually related to melatonin secretion. In amphibians, melatonin exhibits a potent skin lightening activity. However, in fishes and reptiles the melatonin effects vary with the species, the developmental stage, and the pigment cell location. Melatonin also exerts inhibitory or excitatory activity on the amphibian reproductive system, regulation of circadian locomotory activity in reptiles, and modulation of the amphibian metamorphosis. Melatonin has also a modulatory effect on the response of target cells to different hormones and high concentrations or prolonged exposure to the indoleamine may cause autodesensitization in various tissues. Binding sites of melatonin have been detected in the central nervous system and peripheral tissues of various vertebrates. The relative potencies of melatonin analogues demonstrated two subtypes of melatonin receptors (ML-1 and ML-2). A transmembrane melatonin receptor has been cloned from Xenopus laevis melanophores; it belongs to the family of the G protein-coupled receptors and exhibits 85% homology with the mammalian nervous system receptor. Melatonin binding sites in the nucleus of many cell types and its potent intracellular anti-oxidant action suggest mechanisms of action other than through the G-protein coupled receptor.     

Melatonin as an organoprotector in the stomach and the pancreas

Melatonin was thought to originate primarily from the pineal gland and to be secreted during the night, but recent studies revealed that gastrointestinal (GI) tract presents another, many times larger, source of melatonin that contributes significantly to the circulating concentration of this indole. Melatonin may exert a direct effect on GI tissues but its major influence on GI organs seems to occur indirectly, via the brain-gut axis including peripheral receptors, sensory afferent (vagal or sympathetic) pathways and central nervous system (CNS) acting on these organs via autonomic efferents and neuromediators. This article reviews and updates our experience with the fascinating molecule, as related to GI organs, with special focus on secretory activity of the stomach and pancreas and the maintenance of their tissue integrity. In addition to being released into the circulation, melatonin is also discharged into the gut lumen and this appears to be implicated in the postprandial stimulation of pancreatic enzyme secretion, mediated by melatonin-induced release of cholecystokinin, acting through entero-gastro-pancreatic reflexes. Although exerting certain differences in the mechanism of action on gastric and pancreatic secretory activities, melatonin derived from its precursor L-tryptophan, exhibits similar highly protective actions against the damage of both the stomach and the pancreas and accelerates the healing of chronic gastric ulcerations by stimulating the microcirculation and cooperating with arachidonate metabolites such as prostaglandins, with nitric oxide released from vascular endothelium, and/or sensory nerves and with their neuropeptides such as calcitonin gene related peptide. The beneficial effects of melatonin results in gastro- and pancreato-protection, prevents various forms of gastritis and pancreatitis through the activation of specific MT2-receptors and scavenges reactive oxygen species (ROS). Melatonin counteracts the increase in the ROS-induced lipid peroxidation and preserves, at least in part, the activity of key anti-oxidizing enzymes such as superoxide dismutase. It is proposed that melatonin should be considered as the agent exerting an important role in prevention of gastric and pancreatic damage and in accelerating healing of gastric ulcers.     

Mice, melatonin and the circadian system

Melatonin effects are discussed by reviewing results from mice with intact or disrupted melatonin signaling. Melatonin, the neuroendocrine hand of the clock produced in the pineal gland during night, acts upon two receptor subtypes. Melatonin receptors are found in the suprachiasmatic nuclei (SCN), hypophysial pars tuberalis (PT) and adrenal gland. In SCN, melatonin interacts with PACAP, a neuropeptide of the retinohypothalamic tract. Moreover, melatonin acts on the SCN to modulate the activity of the sympathetic nervous system. Melatonin is not required to maintain rhythmic clock gene expression in SCN. By contrast, the rhythmic clock gene expression in PT depends on a melatonin signal interacting with adenosine. Melatonin may also affect clock gene protein levels in the adrenal cortex and influence adrenal functions. In conclusion, melatonin may serve the synchronization of peripheral oscillators by interacting with other neuroactive substances. A stress-reducing potency of melatonin needs to be explored in further studies.     

A brief survey of pineal gland-immune system interrelationships.

The present paper summarizes evidence that support the hypothesis of the existence of bilateral interactions between pineal gland and the immune system. Both in vivo and in vitro experiments show that the pineal gland, via its hormone melatonin, enhances immune function. Mechanisms involved in this immunostimulatory effect are not well understood, but some evidence suggests the existence of specific binding sites for melatonin on immune cells. Moreover, the release of opioid peptides and interleukin-2 by T-helper cells may also participate in this mechanism by activating, at least natural killer activity and antibody-dependent cellular cytotoxicity. Some immune signals, i.e., gamma-interferon, may be involved in regulating pineal function, thereby representing a regulatory mechanism in the opposite direction. The physiological and clinical significance of these data remains to be studied.     

mt1 Melatonin receptor in the primate adrenal gland: inhibition of adrenocorticotropin-stimulated cortisol production by melatonin

The pineal hormone melatonin participates in circadian, seasonal, and reproductive physiology. The presence of melatonin binding sites in human brain and peripheral tissues is well documented. However, in the mammalian adrenal gland, low-affinity melatonin binding sites have been detected only in the rat by some but not all authors. Conflicting evidence for a regulatory role of melatonin on adrenal cortisol production, prompted us to investigate this possibility in a New World primate, the capuchin monkey. Expression of melatonin receptors in the adrenal cortex was demonstrated through pharmacological characterization and autoradiographic localization of 2-[125I]iodomelatonin binding sites (dissociation constant = 96.9 +/- 15 pM; maximal binding capacity = 3.8 +/- 0.4 fmol/mg protein). The mt1 identity of these receptors was established by cDNA sequencing. Melatonin treatment of dispersed cells and explants from adrenal gland did not affect basal cortisol production. However, cortisol production stimulated by 100 nM ACTH was significantly inhibited by low melatonin concentrations (0.1-100 nM); this inhibitory effect was reversed by the mt1/MT2 melatonin antagonist luzindole. Melatonin also inhibited dibutyril-cAMP-stimulated cortisol production, suggesting that melatonin acts through a cAMP-independent signaling pathway. The present data demonstrate that the primate adrenal gland cortex expresses functional mt1 melatonin receptors and shows that melatonin inhibits ACTH-stimulated cortisol production.     

Melatonin modulates secretion of growth hormone and prolactin by trout pituitary glands and cells in culture

In Teleost fish, development, growth, and reproduction are influenced by the daily and seasonal variations of photoperiod and temperature. Early in vivo studies indicated the pineal gland mediates the effects of these external factors, most probably through the rhythmic production of melatonin. The present investigation was aimed at determining whether melatonin acts directly on the pituitary to control GH and prolactin (PRL) secretion in rainbow trout. We show that 2-[125I]-iodomelatonin, a melatonin analog, binds selectively to membrane preparations and tissue sections from trout pituitaries. The affinity was within the range of that found for the binding to brain microsomal preparations, but the number of binding sites was 20-fold less than in the brain. In culture, melatonin inhibited pituitary cAMP accumulation induced by forskolin, the adenyl cyclase stimulator. Forskolin also induced an increase in GH release, which was reduced in the presence of picomolar concentrations of melatonin. At higher concentrations, the effects of melatonin became stimulatory. In the absence of forskolin, melatonin induced a dose-dependent increase in GH release, and a dose-dependent decrease in PRL release. Melatonin effects were abolished upon addition of luzindole, a melatonin antagonist. Our results provide the first evidence that melatonin modulates GH and PRL secretion in Teleost fish pituitary. Melatonin effects on GH have never been reported in any vertebrate before. The effects result from a direct action of melatonin on pituitary cells. The complexity of the observed responses suggests several types of melatonin receptors might be involved.     

Melatonin and its precursor, L-tryptophan: influence on pancreatic amylase secretion in vivo and in vitro

Melatonin, considered as a main pineal product, may be also synthetized in the gastrointestinal tract from L-tryptophan. Melatonin has been recently shown to affect insulin release and its receptors have been characterized in the pancreas however, the effects of melatonin on the pancreatic enzyme secretion have not been examined. The aim of this study was to investigate the effects of melatonin or L-tryptophan on amylase secretion in vivo in anaesthetized rats with pancreato-biliary fistulas, and in vitro using isolated pancreatic acini. Melatonin (1, 5 or 25 mg/kg) or L-tryptophan (10, 50 or 250 mg/kg) given to the rats as a intraperitoneal (i.p.) bolus injection produced significant and dose-dependent increases in pancreatic amylase secretion under basal conditions or following stimulation of enzyme secretion by diversion of bile-pancreatic juice. This was accompanied by a dose-dependent rise in melatonin plasma level. Stimulation of pancreatic enzyme secretion caused by melatonin or L-tryptophan was completely abolished by vagotomy, deactivation of sensory nerves with capsaicin or pretreatment with CCK1 receptor antagonists (tarazepide or L-364,718). Pretreatment with luzindole, an antagonist of melatonin MT(2) receptor failed to affect melatonin- or L-tryptophan-induced amylase secretion. Administration of melatonin (1, 5 or 25 mg/kg i.p.) or L-tryptophan (10, 50 or 250 mg/kg i.p.) to the rats resulted in the dose-dependent increase of cholecystokinin (CCK) plasma immunoreactivity. Enzyme secretion from isolated pancreatic acini was not significantly affected by melatonin or L-tryptophan used at doses of 10(-8) -10(-5) M. We conclude that exogenous melatonin, as well as that produced endogenously from L-tryptophan, stimulates pancreatic enzyme secretion in vivo while increasing CCK release. Stimulatory effect of melatonin or L-tryptophan on the exocrine pancreas involves vagal sensory nerves and the CCK release by these substances.     

Minireview: Cell protective role of melatonin in the brain

Abstract: In recent years an increasing amount of data has been published involving melatonin in the control of brain function. The pineal gland exerts a depressive influence on CNS excitability. This activity is linked to melatonin, since pharmacological doses of the hormone prevent seizures in several animal models. In addition, melatonin also has analgesic properties in these species. However, the sites and mechanism of melatonin action are not known. A role for the pineal gland and its hormone melatonin as a homeostatic system controlling brain excitability has been proposed, and GABA-containing neurons may be involved in some central action of melatonin. There is evidence supporting a role of melatonin in the regulation of the GABA-benzodiazepine receptor complex, and it appears that melatonin potentiates this inhibitory neurotransmitter system in brain. Melatonin does not bind to GABA or benzodiazepine binding sites themselves, because in vitro binding data showed that melatonin is a weak competitor of benzodiazepine binding in brain membranes at concentrations greater than 10^?5 M. The effect of melatonin on brain activity also involves the participation of corticotropic and opioid peptides, and the existence of an opioid-antiopioid homeostatic system is proposed, with the GABA-benzodiazepine receptor complex as an effector. Moreover, the interaction of melatonin with corticotropic peptides and mitochondrial benzodiazepine receptors may result in a participation of neurosteroids in the control of GABA activity and function. The most recently available data from biochemical and electrophysiological studies support the possibility that the anticonvulsant and depressive effects of melatonin on neuron activity may depend on its antioxidant and antiexcitotoxic roles, acting as a free radical scavenger and regulating brain glutamate receptors. The full characterization of the nuclear melatonin receptor explains the genomic effects of melatonin, opening a new perspective regarding actions and roles of melatonin as a cellular protector.     

Melatonin: a hormone,a tissue factor,an autocoid,a paracoid,and an antioxidant vitamin

Melatonin, a derivative of an essential amino acid, tryptophan, was first identified in bovine pineal tissue and subsequently it has been portrayed exclusively as a hormone. Recently accumulated evidence has challenged this concept. Melatonin is present in the earliest life forms and is found in all organisms including bacteria, algae, fungi, plants, insects, and vertebrates including humans. Several characteristics of melatonin distinguish it from a classic hormone such as its direct, non-receptor-mediated free radical scavenging activity. As melatonin is also ingested in foodstuffs such as vegetables, fruits, rice, wheat and herbal medicines, from the nutritional point of view, melatonin can also be classified as a vitamin. It seems likely that melatonin initially evolved as an antioxidant, becoming a vitamin in the food chain, and in multicellular organisms, where it is produced, it has acquired autocoid, paracoid and hormonal properties.     

Presynaptic effects of melatonin on norepinephrine release and uptake in rat pineal gland

The effect of melatonin injection on norepinephrine (NE) turnover rate in rat pineal gland was estimated from the decline of tissue NE levels after the injection of the tyrosine hydroxylase inhibitor alpha-methyl-p-tyrosine. The administration of a single injection of 300 micrograms/Kg of melatonin at the beginning of the scotophase induced, 3 hr later, a significant decrease of pineal NE turnover. The possible direct effect of melatonin on pineal NE release was examined in vitro. Exposure of rat pineal explants previously loaded with 3H-NE to 10(-8)-10(-6) M melatonin decreased significantly 3H-NE release triggered by 60 mM K+. This activity of melatonin was revealed only in pineals excised at night (0000 and 0400, i.e., at the fourth or eighth hours of darkness) and not in those excised in the middle (1400) or late light phase of the daily photoperiod (2000). Melatonin did not modify the spontaneous pineal 3H-NE efflux. Melatonin decreased 3H-NE uptake at a low NE concentration (0.5 microM) in a dose-dependent manner (IC50 identical to 10(-10) M). A kinetic analysis of the pineal NE uptake process indicated that melatonin augmented both Vmax and Km of transmitter uptake. These results suggest that endogenously released melatonin may be a regulatory signal for rat pineal sympathetic synapses.     

Evidence of immune system melatonin production by two pineal melatonin deficient mice, C57BL/6 and Swiss strains

Abstract:  We evaluated two pineal melatonin deficient mice described in the literature, i.e., C57BL/6 and Swiss mice, as animal models for studying the immunomodulatory action of melatonin. Plasma melatonin levels in C57BL/6 and Swiss strains were detectable, but lower than levels in control C3H/HENHSD mice. Since these strains are suppose to be pineal melatonin deficient an extrapineal melatonin synthesis may contribute to plasma levels. Regarding cells and tissues from the immune system, all of them were found to synthesize melatonin although at low levels. N-acetyltransferase (AANAT) mRNA was also amplified in order to analyze the alternative splicing between exons 3–4 described for pineal C57BL/6 mice which generates an inclusion of a pseudoexon of 102 bp. For the pineal gland, both the wild type and the mutant isoforms were present in all mice strains although in different proportions. We observed a predominant wild type AANAT mature RNA in thymus, spleen and bone marrow cells. Peripheral blood mononuclear cells (PBMC) culture shown an evident AANAT amplification in all strains studied. Although the bands detected were less intense in melatonin deficient mice, the amplification almost reached the control cell intensity after stimulation with phytohemaglutinin (PHA). In summary, melatonin detection and AANAT mRNA expression in inbred and outbred mice clearly indicate that different cells and tissues from the immune system are able to synthesize melatonin. Thus, the pineal defect seems not to be generalized to all tissues, suggesting that other cells may compensate the low pineal melatonin production contributing to the measurable plasma melatonin level.     

Inherited changes in concentrations of retinal and serum melatonin in the chicken

Concentrations of melatonin in the retina, serum, and pineal gland were studied in genetically blind chicks carrying an autosomal recessive mutation, rc, characterized by the degeneration of photoreceptors in the retina after hatching. Blind homozygous (rc/rc) and sighted heterozygous (Rc+/rc) chicks were housed under 12:12 light:dark cycles. They were decapitated at 4, 6, 8, and 10 weeks of age at midlight and middark. Retinas, pineal glands, and serum samples were collected, and the resultant tissue melatonin was extracted and determined by radioimmunoassay. Retinal and pineal melatonin were also identified and quantified by gas chromatography-mass spectrometry (GC-MS). Good correlations were demonstrated between the values obtained by GC-MS and levels of quantified by radioimmunoassay. In all the tissues studied, there were age-related changes and diurnal variations in melatonin levels with high levels in the dark period. Melatonin levels in the retina and serum of rc/rc chicks were also significantly lower than those of Rc+/rc control birds. However, storages of melatonin in the pineal gland were similar between the two groups of chicks studied. These results suggest that (1) retinal melatonin is synthesized in the photoreceptor, (2) the phototransduction process which produces neural signals (i.e., electroretinogram) may be different from the phototransduction process which initiated the rhythmic melatonin synthesis and production in the retina, (3) the inherited degeneration of retinal photoreceptors with lower retinal melatonin levels correlates with an inherited abnormality of the pineal melatonin synthesis and/or secretion resulting in lower serum melatonin levels (pleiotropism), (4) levels of pineal melatonin (an indicator of the rate of synthesis and/or storage) and that of serum melatonin (an indicator of the rate of release) may not be directly correlated, and (5) the chicken pineal secretes melatonin not only by simple diffusion but also from a bound pool of melatonin in the gland.     

Fluctuation of Blood Melatonin Concentrations With Age: Result of Changes in Pineal Melatonin Secretion, Body Growth, and Aging

Melatonin in the systemic circulation of rats fluctuates with age, and the causes for such changes were investigated. Male rats (aged 7 days, 16 days, 18 days, 20 days, 30 days, 48 days, 60 days, and greater than 17 months) were adapted under a lighting regime of 12L:12D for at least 7 days. Pineals and blood samples from the trunk or confluens sinuum were collected in the dark period. Melatonin in tissues was extracted, identified, and determined by gas chromatography-mass spectrometry (GC-MS) and/or radioimmunoassay. Tissue melatonin levels obtained by radioimmunoassay correlated closely with those quantified by GC-MS. Thus, the melatonin radioimmunoassay used is a reliable assay method for melatonin in the plasma and pineal of the rat. Plasma melatonin in the confluens sinuum of rats exhibited episodic release superimposed on a basal release pattern. It was suggested that there are two pools of melatonin in the pineal gland, a readily releasable pool and a bound pool. The mean plasma levels of melatonin in the confluens sinuum of rats increased with age with the highest level recorded at 60 days old and declined to a lower level at greater than 17 months old. The above age-related changes, being similar to the alterations in pineal melatonin levels with growth and aging, suggest that, under our experimental conditions, levels of pineal melatonin increase or decrease with its secretory rate. In developing rats, the age-related increase in the rate of secretion of pineal melatonin as reflected by increases in melatonin levels in the confluens sinuum or pineal melatonin content before adulthood is different from the changes in melatonin levels in the systemic circulation which showed an early developmental rise, followed by an active period and then a prepubertal decline. However, when the body weight was taken into consideration, changes in the levels of pineal melatonin content per 100 gm body weight or the calculated blood melatonin levels (plasma melatonin in the confluens sinuum/body:head ratio) correlated well with the fluctuation of serum melatonin in the systemic circulation. Thus, the developmental changes in the concentrations of melatonin in the general circulation are the result of 1) changes in the rate of pineal melatonin secretion and 2) increase in the dilution factor because of increase in body size. In old rats, levels of plasma melatonin in the confluens sinuum and pineal melatonin content decreased indicating a decline in the rate of pineal melatonin secretion.(ABSTRACT TRUNCATED AT 400 WORDS)     

The effect of food deprivation on brain and gastrointestinal tissue levels of tryptophan, serotonin, 5-hydroxyindoleacetic acid, and melatonin

In order to investigate the effect of food deprivation on the levels of indoles in the brain and the gastrointestinal tissues, we have determined tissue levels of tryptophan (TRP), serotonin (5-HT), 5-hydroxyindoleacetic acid (5-HIAA), and melatonin in the brain and the gastrointestinal tract (GIT) of mice on ad libitum diet as well as in mice deprived of food for 24 and 48 hr. The reduction of food intake 1) had no effect on TRP levels in the brain, but increased TRP concentrations in the stomach and the gut, especially in the colon; 2) decreased 5-HT levels in the brain, but increased values in the stomach and the intestines; 3) decreased 5-HIAA levels in the brain, but increased them in the stomach and the intestines; 4) did not change 5-HT conversion to 5-HIAA in the brain, stomach, and the jejunum, but increased the conversion in the ileum and colon and; 5) increased melatonin levels in all tissues investigated, particularly in the stomach and the brain. The changes of indole levels induced by food deprivation were compared to their known function in the brain and the individual segments of the GIT. A possible serotonin-melatonin antagonism in the brain and GIT function is considered.     

Pineal gland interface between the photoperiodic environment and the endocrine system.

The photoperiodic message that the pineal gland conveys to the organism is encoded in the circadian melatonin rhythm. Melatonin is a ubiquitously acting hormone that mediates seasonal changes in reproduction in nonhuman mammals and may have reproductive consequences in humans as well. Additionally, melatonin may relate to the function o f the immune system, hormone-responsive tumor growth, circadian rhythm disturbances, and a number of other processes.     

Melatonin for the treatment of irritable bowel syndrome

Irritable bowel syndrome(IBS)is a common disorder characterized by recurrent abdominal pain or discomfort,in combination with disturbed bowel habits in the absence of identifiable organic cause.Melatonin(Nacetyl-5-methoxytryptamine)is a hormone produced by the pineal gland and also large number by enterochromaffin cells of the digestive mucosa.Melatonin plays an important part in gastrointestinal physiology which includes regulation of gastrointestinal motility,local anti-inflammatory reaction as well as moderation of visceral sensation.Melatonin is commonly given orally.It is categorized by the United States Food and Drug Administration as a dietary supplement.Melatonin treatment has an extremely wide margin of safety though it may cause minor adverse effects,such as headache,rash and nightmares.Melatonin was touted as a potential effective candidate for IBS treatment.Putative role of melatonin in IBS treatment include analgesic effects,regulator of gastrointestinal motility and sensation to sleep promoter.Placebo-controlled studies in melatonin suffered from heterogeneity in methodology.Most studies utilized 3 mg at bedtime as the standard dose of trial.However,all studies had consistently showed improvement in abdominal pain,some showed improvement in quality of life of IBS patients.Melatonin is a relatively safe drug that possesses potential in treating IBS.Future studies should focus on melatonin effect on gut mobility as well as its central nervous system effect to elucidate its role in IBS patients.     

Distribution, function and physiological role of melatonin in the lower gut

Melatonin is a hormone with endocrine, paracrine andautocrine actions. It is involved in the regulation of multiple functions, including the control of the gastroin-testinal (GI) system under physiological and pathophys-iological conditions. Since the gut contains at least 400times more melatonin than the pineal gland, a reviewof the functional importance of melatonin in the gutseems useful, especially in the context of recent clinicaltrials. Melatonin exerts its physiological effects throughspecific membra...     

Properties of the melatonin-generating system of the sailfin molly, Poecilia velifera

The properties of the melatonin-generating system of a tropical teleost, the sailfin molly (Poecilia velifera), were investigated in vitro in a series of experiments using static or perifusion culture techniques. The properties examined included photic entrainment, circadian rhythmicity under continuous light (LL) and continuous darkness (DD), functionality of the melatonin-generating system at birth, and presence of multiple circadian oscillators in the molly pineal. Pineal glands or skull caps with the pineal gland firmly attached were dissected from adult and new-born fishes, respectively, and placed into static or perifusion culture at constant temperature (27 degrees C) depending upon the experiment. Melatonin release in samples was quantified by RIA. Rhythmic melatonin release was observed from isolated adult pineals under 12L:12D and 14L:10D, with low amounts of melatonin released during the light and high amounts during the dark. Melatonin release was inhibited by LL. However, under DD, melatonin release was robust and rhythmic with a circadian period (Tau) that ranged between 21.3 and 27.0 h (n = 21). Pineals from new-born (1-day old) mollies released melatonin rhythmically under a light:dark cycle and DD in both static and perifusion culture. Melatonin release from half and quarter pineals of adult mollies under DD was robust and rhythmic with circadian periods that ranged between 22.5 and 29.0 h (n = 19). Taken together, these data show that the molly pineal is photosensitive, fully functional from birth, and contains multiple circadian oscillators (at least four) regulating melatonin production.     

Pineal melatonin synthesis and release are not altered throughout the estrous cycle in female rats

Melatonin times reproduction with seasons in many photoperiodic mammalian species. Whether sexual hormones reflect on melatonin synthesis is still debated. The aim of this work was to study, using a large panel of technical approaches, whether the daily profile of pineal melatonin synthesis and release varies with the estrous cycle in the female rat. The mRNA levels and enzyme activities of the melatonin synthesizing enzymes, arylalkylamine N-acetyltransferase and hydroxyindole-O-methyltransferase were similar at the four stages of the rat estrous cycle. The endogenous release of melatonin, followed by transpineal microdialysis during six consecutive days in cycling female rats, displayed no significant variation during this interval. Taken together, the present results demonstrate that there is no regular fluctuation in the pineal metabolism leading to melatonin synthesis and release throughout the estrous cycle in female rats.