Immuno-derived opioids as mediators of the immuno-enhancing and anti-stress action of melatonin.

The pineal neurohormone melatonin was shown to stimulate the release of opioid peptides from activated CD4+, T lymphocytes. These immuno-derived opioids or "lymphomorphins" crossreact with anti-beta-endorphin and anti-metenkephalin antisera, bind to opioid receptors in the thymus and are the mediators of the immunoenhancing and anti-stress action of endogenous and/or exogenous melatonin. These findings proved the existence of a novel immunoneuroendocrine physiological mechanism which may be related for the long range maintenance of the immune homeostasis in spite of the unavoidable stress and/or infectious events occurring during the span of life in a normal unprotected environment.     

The involvement of pineal gland and melatonin in immunity and aging. I. Thymus-mediated, immunoreconstituting and antiviral activity of thyrotropin-releasing hormone

Circadian, continued treatment with melatonin during the dark cycle produces changes in the blood level of thyroid hormones in aging mice. Thyroid-stimulating hormone (TSH) and thyrotropin-releasing hormone (TRH) antagonize the involution of the thymus produced by prednisolone. This effect of TRH does not seem thyroid dependent. TRH restores antibody production in non-responder athymic nude mice but does not exert this effect in neonatally thymectomized mice. Moreover, this activity does not correlate with thyroxine levels. TRH exerts a powerful protective effect in mice challenged with encephalomyocarditis (EMC) virus. Presumably pineal melatonin exerts its varied regulatory functions via hypothalamic TRH.     

The pineal neurohormone melatonin stimulates activated CD4+, Thy-1+ cells to release opioid agonist(s) with immunoenhancing and anti-stress properties

In previous studies we showed that in mice the pineal gland modulates the immune response via the circadian synthesis and release of melatonin. Exogenous melatonin proved also to exert immunoenhancing effects and to counteract completely the immunologic effect of acute stress. Melatonin was active only in vivo, in mice primed with T-dependent antigens and its effects on the primary antibody response and thymus weight were abolished by the specific opioid antagonist naltrexone. Here we demonstrate that physiologic concentrations of melatonin stimulate, in vitro, activated L3T4+ (CD4+) cells to release opioid agonist(s) that can reproduce in vivo the immunoenhancing and anti-stress effects on thymus cellularity and antibody production of melatonin and compete with specific binding of [3H]naloxone to mouse brain membranes. Similar results were obtained when mitogen-activated human immunocompetent cells were incubated with melatonin. In the human model the results were, however, less consistent than those obtained with murine cells, in that only four out of ten blood donors provided cells that were responsive to melatonin. This finding elucidates the mechanism of a novel immuno-neuroendocrine connection with relevant implications for our understanding of the neuroendocrine factors that may influence the immune response in vivo in normal and stressful situations. In addition, it opens new perspectives in a wide range of research fields.     

Anti-stress role of the melatonin-immuno-opioid network: evidence for a physiological mechanism involving T cell-derived, immunoreactive beta-endorphin and MET-enkephalin binding to thymic opioid receptors.

Our previous work showed that the pineal neurohormone melatonin induces activated T lymphocytes to release opioid peptides with immunoenhancing and anti-stress properties. Here we present evidence that these peptides crossreact with anti-beta-endorphin and anti-met-enkephalin antisera, and bind specifically to thymic opioid receptors. Furthermore, the same antisera injected in prednisolone treated mice prevented the normal recovery of thymus cellularity and of the capacity to mount a primary antibody response against T-dependent antigens. Surgical pinealectomy, i.e. inhibition of endogenous melatonin and absence of antigen activation negated the effect of such antisera demonstrating the physiological relevance of this melatonin-immuno-opioids network. It is proposed that function of this network may be that of driving a correct immune recovery after the depression caused by the elevated corticosteroids level associated with immune responses and/or stressful situations.     

Attenuation of reserpine-induced catalepsy by melatonin and the role of the opioid system

Several reports have indicated that melatonin influences motor activity in animals and humans. Melatonin has been reported to attenuate the rigidity and tremor of Parkinson's disease. Some of the behavioral effects (e.g., analgesic and anticonvulsant properties) of melatonin have been reported to be mediated through interactions with the endogenous opioid peptides. We investigated the effect of melatonin on reserpine-induced catalepsy in the rat and, additionally, examined whether this effect is modified by opioid peptides. Melatonin was found to attenuate markedly the duration of reserpine-induced catalepsy. These effects were potentiated by administration of the opiate agonist nalbuphine hydrochloride, while naloxone partially reversed the catalepsy reducing effect of melatonin. These findings suggest that the motor effects of melatonin may involve critical interactions with opioid peptides, and support the postulated reciprocal interactions between melatonin and opioid peptides that previously have been demonstrated for the analgesic and anticonvulsant properties of melatonin.     

Effects of melatonin on orphanin FQ/nociceptin-induced hyperalgesia in mice

The pain modulatory properties of melatonin (MT) are generally recognized but the detail of the interaction between melatonin and opioid system in pain regulation is not fully understood. The present study was undertaken to investigate the modulatory effect of melatonin (MT) on the hyperalgesic effect of Orphanin FQ/Nociceptin (OFQ/NC, NC), a member of opioid peptide family. Intracerebroventricular (i.c.v.) administration of NC (10 microg/mouse) induced significant hyperalgesic effect in tail-flick test in mice; i.c.v. (5, 10, 50 microg/mouse) or intraperitoneal (i.p.) (5, 10, 50 mg/kg) co-injection of melatonin dose-dependently reversed NC-induced hyperalgesia and showed a profound analgesic effect. The antihyperalgesia effect of MT could be significantly antagonized by i.c.v. co-injection of luzindole (10 microg/mouse) (an antagonist of MT receptor) or naloxone (10 microg/mouse) (antagonist of traditional opioid receptor). Taken together, all the results suggested that MT could produce a luzindole and naloxone sensitive reversing effect on NC-induced hyperalgesia at supraspinal and peripheral level in mice. The augmentation effect of MT on the traditional opioid system may be one of the mechanisms of this antihyperalgesia action induced by MT. The present work will help to elucidate the mechanism of the pain modulation effect of MT, and also will help to represent new interesting modulating therapeutic targets for the relief of pain.     

The effects of oral melatonin on the autonomic function in healthy subjects.

We investigated the influence of melatonin on the human autonomic functions by measuring muscle sympathetic nerve activity (MSNA). Five healthy male volunteers took 3 mg of melatonin, and their plasma melatonin concentration, blood pressure, heart rate, and burst rate of MSNA were then recorded. The peak level of melatonin concentration showed a marked interindividual variation. Blood pressure was reduced significantly, while heart rate and burst rate of MSNA did not change significantly. These results indicate that melatonin has a hypotensive effect on blood pressure, and the central cardiovascular regulatory mechanism such as lowering of the baroreflex setpoint would be involved in the effect.     

Effect of melatonin replacement on serum hormone rhythms in a patient lacking endogenous melatonin.

A potentially confounding variable inherent in studies designed to examine the effect of melatonin administration in humans is the presence of an endogenous melatonin rhythm in the experimental subjects. The effects of exogenous melatonin administration on serum hormone rhythms was recently examined in a male patient who lacked detectable circulating levels of endogenous melatonin. The patient's pineal gland had been destroyed five years previously in the course of treatment for a pineal astrocytoma. On three separate occasions, over approximately a one-year period, the patient was given daily oral melatonin replacement (2 mg/day, 1 mg/day and 0.5 mg/day). These experiments were designed to assess the effects of exogenous melatonin on serum growth hormone, prolactin, cortisol and testosterone rhythms. Analysis of blood samples collected every 2-4 hours periods both before and during melatonin replacement revealed that the exogenous melatonin rhythm was associated with improvements in self-reported sleep and mood ratings. Melatonin administration produced robust nocturnal peaks in serum growth hormone and prolactin levels immediately following ingestion of the hormone, while serum cortisol and testosterone rhythms were not influenced. These results suggest that melatonin may modulate the coordination and enhancement of selected biological rhythms in man.     

Abnormal nocturnal melatonin secretion and disordered sleep in abstinent alcoholics.

BACKGROUND: Alcoholic patients show prominent disturbance of sleep as measured by electroencephalogram, with difficulties in the onset and maintenance of sleep. Given the role of melatonin in the regulation of the sleep-wake cycle, this study examined the relationship between nocturnal expression of melatonin and sleep in alcoholics as compared with control subjects. METHODS: Alcoholic patients (n = 11) and comparison control subjects (n = 10) underwent all-night polysomnography and serial blood sampling every 30 min from 10:00 PM to 6:30 AM for measurement of circulating levels of melatonin and cortisol. RESULTS: Coupled with prolonged sleep latency, alcoholics showed lower levels of melatonin during the early part of the night and a delay in the onset of the nocturnal plateau or peak value of melatonin as compared with control subjects. The nocturnal delay of melatonin correlated with prolonged sleep latency. Circulating levels of cortisol were lower during the early part of the night and higher in the late part of night in the alcoholics as compared with the control subjects. CONCLUSIONS: A delay in the nocturnal rise of melatonin may contribute to disordered sleep in chronic alcoholics, with implications for the use of melatonin in the treatment of insomnia in recovering alcoholics.     

Reversal of morphine tolerance and dependence by melatonin: possible role of central and peripheral benzodiazepine receptors.

Possible reversal by melatonin of morphine-induced tolerance and dependence was studied in mice. A 10-day repeated injection regimen was followed to induce morphine tolerance and dependence. Co-administration of melatonin (1-10 mg/kg, i.p.) with morphine (10 mg/kg, s.c.) during the induction phase (day 1 to 9) reversed the development of opioid tolerance and dependence tested on 10th day. On the other hand acute administration of melatonin (1-10 mg/kg) on the 10th day, ie. during the expression phase of morphine dependence, it reduced the incidence of naloxone-induced withdrawal jumps without affecting the tolerance to analgesic effect. Co-administration of flumazenil (2 mg/kg, i.p.), a central benzodiazepine (BZ) receptor antagonist had no effect on melatonin response, whereas peripheral antagonist for BZ receptor PK11195 (2 mg/kg, i.p.) significantly reversed the attenuating effect of melatonin on physical dependence both during induction and expression phase of morphine tolerance and dependence. These observations suggest that melatonin reverses development of tolerance and dependence to morphine, and this action possibly involved peripheral benzodiazepine receptors.     

Features of melatonin catabolism in chicks

Melatonin is a neurohormone that is involved in biorhythm synchronization that is produced by the pineal gland and retina. We have studied the production of the kynurenine products, N-acetyl-N-formyl-5-methoxykynurenamine (AFMK) and N-acetyl-5-methoxykynurenamine (AMK), during catabolism of melatonin in chicks (Gallus gallus). Analyses of blood serum samples, retinas, and pineal glands were performed using the HPLC method with fluorometric and electrochemical detection. Considerable amounts of melatonin were found in the pineal gland, retina, and blood serum, although endogenous AFMK and AMK were not found in these tissues. Nevertheless, after intraperitoneal injection of melatonin at a dose of 10 mg/kg, these compounds were found in the blood serum, and AMK was found in the retina. Analysis of AFMK and AMK pharmacodynamics in the blood serum showed that their peak concentrations were 4.8 ng/ml and 2 ng/ml, respectively; the peaks were observed within less than 5 minutes after melatonin injection for AFMK and in 20 minutes after melatonin injection for AMK. The amount of AMK in the retina also reached its peak (60 pg/retina) in 20 minutes. The concentration of both metabolites decreased to the detection limit within one hour. Thus, melatonin may be oxidized via the kynurenine pathway, and its oxidation products, AFMK and AMK, have short half lives. The total amount of both metabolites after melatonin injection was approximately 0.04% of its amount in the blood serum and retina, thus, this pathway of melatonin oxidation is not the major one.     

Pineal melatonin and sensory symptoms in Parkinson disease

Sensory symptoms have been reported in 40–60% of patiens with Parkinson's disease, and in at least 10% of patients these symptoms precede the onset of the motor disorder. The pathophysiology of these symptoms remains unknown. Diminished brain serotonin concentration has been reported to be associated with sensory symptoms. Serotonin metabolism is regulated by pineal melatonin. The secretory activity of the pineal gland may be diminished in Parkinson's disease. In experimental animals pineal melatonin has been shown to exert analgesic effects by interacting with opiate receptors. In addition, since opioid peptides mediate the analgesic effects of melatonin, decreased opioid peptide functions in Parkinson's disease may be associated with disruption of the “fine-tuning” pain modulatory functions of melatonin and possibly indirectly facilitate the emergence of sensory symptoms.

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Sommario Nel 40–60% dei pazienti affetti da morbo di Parkinson sono stati descritti sintomi sensoriali e nel 10% di essi questi sintomi precedono l'inizio dei disturbi motori. La patofisiologia di questi sintomi è, per ora, sconosciuta. è stata però descritta una diminuzione della concentrazione della serotonina cerebrale associata ai sintomi sensoriali. Il metabolismo della serotonina è regolato dalla melatonina pineale: perciò l'attività secernente della ghiandola pineale può essere diminuita nel morbo di Parkinson. Nelle esperienze sugli animali la melatonina pineale risulta esercitare un'azione analgesica in quanto interagisce coi recettori oppioidi. Inoltre, piché i peptidi oppioidi mediano gli effetti analgesici della melatonina, la riduzione degli effetti dei peptidi oppioidi nel morbo di Parkison può essere associato colla scomparsa delle funzioni modulatorie della melatonina sul dolore e ciò può facilitare indirettamente la comparsa di sintomi sensoriali.

     

Immunomodulation by melatonin: its significance for seasonally occurring diseases

Melatonin is not only synthesized by the pineal gland but also in many other organs and tissues of the body, particularly by lymphoid organs such as the bone marrow, thymus and lymphocytes. Melatonin participates in various functions of the body, among which its immunomodulatory role has assumed considerable significance in recent years. Melatonin has been shown to be involved in the regulation of both cellular and humoral immunity. Melatonin not only stimulates the production of natural killer cells, monocytes and leukocytes, but also alters the balance of T helper (Th)-1 and Th-2 cells mainly towards Th-1 responses and increases the production of relevant cytokines such as interleukin (IL)-2, IL-6, IL-12 and interferon-gamma. The regulatory function of melatonin on immune mechanisms is seasonally dependent. This fact may in part account for the cyclic pattern of symptom expression shown by certain infectious diseases, which become more pronounced at particular times of the year. Moreover, melatonin-induced seasonal changes in immune function have also been implicated in the pathogenesis of seasonal affective disorder and rheumatoid arthritis. The clinical significance of the seasonally changing immunomodulatory role of melatonin is discussed in this review.     

Specific binding of melatonin by purified cell nuclei from spleen and thymus of the rat

In the present paper, we show that pineal hormone melatonin interacts with purified cell nuclei from rat spleen and thymus. Binding of 2-[]iodomelatonin ([]melatonin) by cell nuclei fulfills all criteria for binding to a receptor site. Binding exhibited properties such as dependence on time and temperature as well as reversibility, saturability, high affinity, and specificity. Results suggested binding to single classes of binding sites. The dissociation constants (K_d) for the binding sites in the spleen and thymus nuclei were 68 and 102 pM, respectively. These data are in close agreement with data obtained from kinetic studies, in which the kinetically derived values of the dissociation constant in the spleen and thymus nuclei were 166 and 537 pM, respectively. The affinities for melatonin of these nuclear binding sites suggest that they may recognize the physiological concentrations of melatonin in the tissues. Finally, we have demonstrated that binding of []melatonin by the nuclei is displaced by CGP 52608, a specific ligand of the putative nuclear melatonin receptor RZR/ROR. Results strongly suggest that in addition to membrane receptor-related mechanisms, nuclear receptors may be involved in the regulation of immune system by melatonin.     

A novel biochemical model linking dysfunctions in brain melatonin, proopiomelanocortin peptides, and serotonin in autism

A novel biochemical model for autism is presented, which proposes that a subgroup of autistic individuals may have a hypersecretion of pineal melatonin that produces a cascade of biochemical effects including a corresponding hyposecretion of pituitary proopiomelanocortin (POMC) peptides and a hypersecretion of hypothalamic opioid peptides and serotonin (5-HT). The model is reviewed, and supporting animal and clinical research, is summarized. The first arm of the model suggests that increases in pineal melatonin results in hypersecretion of 5-HT in hypothalamus and blood. The second arm of the model indicates that hypersecretion of melatonin also inhibits the release of hypothalamic corticotrophin-releasing hormone (CRH). Hyposecretion of CRH may result in decreased release of both pituitary B-endorphin (B-E) and adrenocorticotrophin hormone (ACTH); this, in turn, may result in decreased plasma concentrations of B-E, ACTH, and cortisol. In autism, a genetically determined hypersecretion of hypothalamic B-E may further contribute to an inhibition of pituitary B-E because of negative feedback inhibition. Therefore, autism may reflect a dysfunction in the pineal-hypothalamic-pituitary-adrenal axis which, modulates POMC and 5-HT systems of the brain. This model is consistent with numerous clinical investigations implicating hypersecretion of brain 5-HT and opioid peptides in autism. The model may have heuristic importance in guiding future research in the biochemistry of autism.     

Influence of maternal melatonin on melatonin receptors in rat offspring

Summary Using quantitative autoradiography, we have studied the influence of maternal plasma melatonin on the expression and density of melatonin receptors in the brain and pituitary of rat offspring. At birth, the same structures displayed melatonin receptors whether the rats were born to and reared by intact or pinealectomized dams. The receptor density was, however, about 20% lower in the group born to pinealectomized dams. At postnatal day 9, when the pups of both groups synthetize rhythmically their own melatonin, this difference was suppressed. These results indicate that melatonin does not appear to be a requirement for the expression of its receptors, but seems to play a stimulatory role in their synthesis.     

The influence of phototherapy on serotonin and melatonin in non-seasonal depression

Circadian profiles of melatonin in serum and serotonin in blood were assessed before and after 7 days of artificial light treatment in 30 patients with non-seasonal depression and 12 healthy subjects. Patients and volunteers were allocated at random to either dim (50 lux) or bright light (2,500 lux) for 2 hours daily. The study has not been completed yet. Preliminary findings are presented here. Light treatment modifies marginally the circadian melatonin profiles of depressed patients and healthy subjects; however, it augments blood serotonin throughout the day. This increase is seen in all patients and healthy subjects after bright as well as dim light. These results suggest that the influence of light is more pronounced on serotonin than melatonin metabolism.     

Blood levels of melatonin, serotonin, cortisol, and prolactin in relation to the circadian rhythm of platelet serotonin uptake.

Blood levels of melatonin, serotonin, cortisol, prolactin, and serotonin uptake by platelets were measured at 08:00, 14:00, 20:00, 02:00, and 08:00 hours in 10 healthy men who ranged in age from 27 to 35 years. The Km values of serotonin active transport by platelets were significantly correlated with melatonin blood levels. There were no other significant correlations. The secretion of steroid hormones and prolactin showed an increase at 02:00 hours; levels of prolactin decreased at 08:00 hours, but steroid levels continued to rise. This finding suggests either a direct effect of melatonin on serotonin active transport or the influence of the suprachiasmatic nucleus on serotonin uptake by platelets. It is also possible that there is a simultaneous decrease in serotonin uptake and decline from peak levels of melatonin due to the rise in steroid secretion.