Melatonin Receptor Subtypes Research Articles

Industrial and academic approaches to the search for alternative melatonin receptor ligands: An historical survey.

The search for melatonin receptor agonists formed the main part of melatonin medicinal chemistry programs for the last three decades. In this short review, we summarize the two main aspects of these programs: the development of all the necessary tools to characterize the newly synthesized ligands at the two melatonin receptors MT1 and MT2, and the medicinal chemist's approaches to find chemically diverse ligands at these receptors. Both strategies are described. It turns out that the main source of tools were industrial laboratories, while the medicinal chemistry was mainly carried out in academia. Such complete accounts are interesting, as they delineate the spirits in which the teams were working demonstrating their strength and innovative character. Most of the programs were focused on nonselective agonists and few of them reached the market. In contrast, discovery of MT1-selective agonists and melatonergic antagonists with proven in vivo activity and MT1 or MT2-selectivity is still in its infancy, despite the considerable interest that subtype selective compounds may bring in the domain, as the physiological respective roles of the two subtypes of melatonin receptors, is still poorly understood. Poly-pharmacology applications and multitarget ligands have also been considered.

Real-Time Determination of Intracellular cAMP Reveals Functional Coupling of Gs Protein to the Melatonin MT1 Receptor.

Melatonin is a neuroendocrine hormone that regulates the circadian rhythm and many other physiological processes. Its functions are primarily exerted through two subtypes of human melatonin receptors, termed melatonin type-1 (MT1) and type-2 (MT2) receptors. Both MT1 and MT2 receptors are generally classified as Gi-coupled receptors owing to their well-recognized ability to inhibit cAMP accumulation in cells. However, it remains an enigma as to why melatonin stimulates cAMP production in a number of cell types that express the MT1 receptor. To address if MT1 can dually couple to Gs and Gi proteins, we employed a highly sensitive luminescent biosensor (GloSensorTM) to monitor the real-time changes in the intracellular cAMP level in intact live HEK293 cells that express MT1 and/or MT2. Our results demonstrate that the activation of MT1, but not MT2, leads to a robust enhancement on the forskolin-stimulated cAMP formation. In contrast, the activation of either MT1 or MT2 inhibited cAMP synthesis driven by the activation of the Gs-coupled β2-adrenergic receptor, which is consistent with a typical Gi-mediated response. The co-expression of MT1 with Gs enabled melatonin itself to stimulate cAMP production, indicating a productive coupling between MT1 and Gs. The possible existence of a MT1-Gs complex was supported through molecular modeling as the predicted complex exhibited structural and thermodynamic characteristics that are comparable to that of MT1-Gi. Taken together, our data reveal that MT1, but not MT2, can dually couple to Gs and Gi proteins, thereby enabling the bi-directional regulation of adenylyl cyclase to differentially modulate cAMP levels in cells that express different complements of MT1, MT2, and G proteins.

Synthesis and Fluorescence Properties of 4-Cyano and 4-Formyl Melatonin as Putative Melatoninergic Ligands.

Fluorescent ligands are imperative to many facets of chemical biology and medicinal chemistry. Herein, we report the syntheses of two fluorescent melatonin-based derivatives as potential ligands of melatonin receptors. The two compounds, namely, 4-cyano and 4-formyl melatonin (4CN-MLT and 4CHO-MLT, respectively), which differ from melatonin by only two/three atoms that are very compact in size, were prepared using the selective C3-alkylation of indoles with N-acetyl ethanolamines involving the "borrowing hydrogen" strategy. These compounds exhibit absorption/emission spectra that are red-shifted from those of melatonin. Binding studies on two melatonin receptor subtypes showed that these derivatives have a modest affinity and selectivity ratio.

Modeling the Heterodimer Interfaces of Melatonin Receptors.

Melatonin receptors are Class A G protein-coupled receptors (GPCRs) that regulate a plethora of physiological activities in response to the rhythmic secretion of melatonin from the pineal gland. Melatonin is a key regulator in the control of circadian rhythm and has multiple functional roles in retinal physiology, memory, immunomodulation and tumorigenesis. The two subtypes of human melatonin receptors, termed MT1 and MT2, utilize overlapping signaling pathways although biased signaling properties have been reported in some cellular systems. With the emerging concept of GPCR dimerization, melatonin receptor heterodimers have been proposed to participate in system-biased signaling. Here, we used computational approaches to map the dimerization interfaces of known heterodimers of melatonin receptors, including MT1/MT2, MT1/GPR50, MT2/GPR50, and MT2/5-HT2C. By homology modeling and membrane protein docking analyses, we have identified putative preferred interface interactions within the different pairs of melatonin receptor dimers and provided plausible structural explanations for some of the unique pharmacological features of specific heterodimers previously reported. A thorough understanding of the molecular basis of melatonin receptor heterodimers may enable the development of new therapeutic approaches against aliments involving these heterodimeric receptors.

Primary role for melatonin MT2 receptors in the regulation of anhedonia and circadian temperature rhythm

Circadian rhythms disturbance is widely observable in patients with major depression (MD) and is also associated with depression vulnerability. Of them, disturbed melatonin secretion rhythm is particularly relevant to MD and is strongly phase-locked to core body temperature (CBT) rhythm.Here we aim to study the specific role of each melatonin receptor (MT1 and MT2) subtype in melatonin regulation of circadian CBT and its possible relationship with depressive-like behaviors.MT1-/- , MT2-/- and WT (C57BL/6) mice were used. Anhedonia, using the sucrose intake test, circadian CBT, environmental place preference (EPP) conditioning and vulnerability to chronic social defeat stress (CSDS) procedure were studied. Moreover, the antidepressant effects of reboxetine (15 mg/kg/day, i.p.) for three weeks or ketamine (15 mg/kg i.p. every four days, 4 doses in total) were studied. Further, exposure to ultra-mild stress induced by individual housing for several weeks was also studied in these mice.MT2-/- mice showed anhedonia and lower CBT compared to WT and MT1-/-. In addition, while reward exposure raised nocturnal CBT in WT this increase did not take place in MT2-/- mice. Further, MT2-/- mice showed an enhanced vulnerability to stress-induced anhedonia and social avoidance as well as an impaired acquisition of novelty seeking behavior. Both reboxetine and ketamine reverted anhedonia and induced a clear anti-helpless behavior in the tail suspension test (TST). Reboxetine raised CBT in mice and reverted ultra-mild stress-induced anhedonia.Our findings show a primary role for MT2 receptors in the regulation of circadian CBT as well as anhedonia and suggest that these receptors could be involved in depressive disorders associated to disturbed melatonin function.

Modulation of human ovarian function by melatonin.

Melatonin, a hormone which is primarily released by the pineal gland, has a wide range of actions in the female reproductive tract. While the melatonin receptor subtype, MT3, has been identified in amphibian animals and birds, in humans and other mammals, melatonin acts through, MT1 and MT2 receptor subtypes which are expressed in human ovaries. The rhythmic release of melatonin starts at puberty and continues throughout fertile female life, affecting and regulating diverse ovarian functions. Here, we discuss the importance of melatonin in regulating folliculogenesis, oocyte quality, ovulation and luteal function, sex steroid receptor gene expression, ovarian steroidogenesis including the production and steroidogenic enzyme activities in the egg and thecal cells. Melatonin improves the egg quality and increases the chance of success of in vitro fertilization (IVF). In view of such extensive actions, melatonin is central to the fertility in females. The objective of this review is to recapitulate the current understanding of the role of melatonin and its receptors.

The law of the synergy and hygiene lighting (literature review)

The previously formulated laws of light hygiene, which allow analyzing the results of the complex impact of stimuli of different nature on the biological object, should be supplemented by the Q-law and the law of synergy. The analysis of the action of the law of synergy in the melatonin-glutamate system is carried out and the General relations characterizing the effect of the synergy of influences are considered. In violation of the Krebs cycle, glutamate can accumulate in the intercellular space. This is a manifestation of the Q-law effect in the glutamate synthesis system in the “neuron-astrocyte” system, which can not only be synthesized in the presence of glucose but also come from food. Astrocytes are important for the whole system, because they provide nutrition to the nervous tissue, contribute to the emergence and formation of the blood-brain barrier. The blood-brain barrier acts as a kind of vacuum cleaner, absorbing excess glutamate and withdrawing it into the systemic circulation, where it does not have a damaging effect. With an excess of dietary glutamate in the blood, there is a decrease in the efficiency of the blood-brain barrier to maintain the optimal level of synthesized glutamate. The effectiveness of glutamate is influenced by melatonin, which provides protection against neurotoxicity caused by glutamate. A system of subtypes of melatonin receptors in functional structures of the eye, the neurotransmitter of which is glutamate and GABA (gamma-aminobutyric acid), is considered. The effects of the Q-law and the law of synergy are demonstrated on a system in which the stimuli acting on neurons include a dose of light, a dose of melatonin, and a dose of glutamate. The cycle of glutamate in neurons obeys the Q-law, and its total dose is formed from both food and synthesized from glucose in the body of glutamate. The complex effect of the total dose of glutamate and melatonin on the sensitivity of ganglion cells obeys the law of synergy. To maintain the synergy effect in the “spectrum of light-melatonin-glutamate” system, it is necessary to comply with many hygienic requirements for maintaining the optimal glutamate cycle (minimizing the dose of dietary glutamate and sugar consumption), the spectral composition of light and the level of cellular melatonin in mitochondria.

Melatonin reduces neuropathic pain behavior and glial activation through MT2 melatonin receptor modulation in a rat model of lysophosphatidylcholine-induced demyelination neuropathy

In this study, we investigated whether melatonin treatment prevents development of neuropathic pain via suppression of glial mitogen-activated protein kinases (MAPKs) activation in the cuneate nucleus (CN) in a lysophosphatidylcholine (LPC)-induced median nerve demyelination neuropathy model. Rats were fed orally with melatonin once a day at a dose of 37.5, 75, or 150 mg/kg 30 min before until 3 days after LPC treatment. Subsequently, behavioral tests were conducted on these animals, and immunohistochemistry and immunoblotting were used for qualitative and quantitative analysis of glia and MAPKs, including ERK, JNK, and p38, activation. Enzyme-linked immunosorbent assays were applied to measure pro-inflammatory cytokine responses. Furthermore, intra-CN microinjection of S26131 (MT1 receptor antagonist), 4P-PDOT (MT2 receptor antagonist), or prazosin (MT3 receptor antagonist) were performed to investigate the association between melatonin receptor subtypes and effects of melatonin on demyelination neuropathy. LPC treatment of the median nerve induced a significant increase in glial fibrillary acidic protein (GFAP; an astrocyte marker) and ED1 (an activated microglia marker) immunoreactivity in the ipsilateral CN and led to development of neuropathic pain behavior. Inspection of GFAP-immunoreactive astrocytes revealed that astrocytic hypertrophy, but not proliferation, contributed to increased GFAP immunoreactivity. Double immunofluorescence showed that both GFAP-immunoreactive astrocytes and ED1-immunoreactive microglia co-expressed p-ERK, p-JNK, and p-p38 immunoreactivity. Melatonin administration dose-dependently reduced neuropathic pain behavior, decreased glial and MAPKs activation, and diminished the release of pro-inflammatory cytokines in the ipsilateral CN after LPC treatment. Furthermore, 4P-PDOT, but not S26131 or prazosin, antagonized the therapeutic effects of melatonin. In conclusion, administration of melatonin, via its cognate MT2 receptor, inhibited activation of glial MAPKs, production of pro-inflammatory cytokines, and development of demyelination-induced neuropathic pain behavior.

Melatonin receptor ligands: A pharmaco-chemical perspective.

Melatonin MT1 and MT2 receptor ligands have been vigorously explored for the last 4 decades. Inspection of approximately 80 publications in the field revealed that most melatonergic ligands were structural analogues of melatonin combining three essential features of the parent compound: an aromatic ring bearing a methoxy group and an amide side chain in a relative arrangement similar to that present in melatonin. While several series of MT2 -selective agents-agonists, antagonists, or partial agonists-were reported, the field was lacking MT1 -selective agents. Herein, we describe various approaches toward the development of melatonergic ligands, keeping in mind that most of the molecules/pharmacophores obtained were essentially melatonin copies, even though diverse tri- or tetra-cyclic compounds were explored. In addition to lack of structural diversity, only few studies examined the activity of the reported melatonergic ligands in vivo. Moreover, an extensive pharmacological characterization including biopharmaceutical stability, pharmacokinetic properties, specificity toward other major receptors to name a few remained scarce. For example, many of the antagonists described were not stable in vivo, were not selective for the melatonin receptor subtype of interest, and were not fully characterized from a pharmacological standpoint. Indeed, virtual screening of large compound libraries has led to the recent discovery of potent and selective melatonin receptor agonists and partial agonists of new chemotypes. Having said this, the melatonergic field is still lacking subtype-selective melatonin receptor antagonists "active" in vivo, which are critical to our understanding of melatonin and melatonin receptors' role in basic physiology and disease.

Melatonin mediates monochromatic light–induced proliferation of T/B lymphocytes in the spleen via the membrane receptor or nuclear receptor

Our studies found that melatonin mediates the monochromatic light–induced lymphocyte proliferation in chickens. However, melatonin receptor subtypes contain membrane receptor (Mel1a/Mel1b/Mel1c) and nuclear receptor (Retinoic acid receptor–related orphan receptor [ROR] α/RORβ/RORγ) and are characteristic with cell specificity. This study compared receptor pathway of melatonin, which mediated the monochromatic light–induced T/B lymphocyte proliferations in chickens. Newly hatched chicks were randomly divided into white light, red light, green light (GL), and blue light groups. Green light promoted the membrane receptor expression in the spleen but decreased the nuclear receptor level compared with that of red light. These changes were accompanied by increase of T/B lymphocyte proliferation and plasma melatonin level under GL. Pinealectomy reversed aforementioned changes and resulted in no differences among the light-treated groups. Supplementation of exogenous melatonin enhanced GL-induced T/B lymphocyte proliferation in the spleen but was reversed by Mel1c antagonist prazosin and RORα agonist SR1078 and enhanced by RORα antagonist SR3335. However, Mel1b antagonist 4P-PDOT and RORγ antagonist GSK increased the stimulation effect of melatonin on GL-induced T lymphocyte proliferation but no effect on the B-lymphocyte proliferation. These results indicate that melatonin promotes the GL-induced T lymphocyte proliferation through Mel1b, Mel1c, and RORα/RORγ; however, the Mel1a, Mel1c, and RORα may be involved in the B lymphocyte proliferation.

Trade-off expression of melatonin receptor subtypes (Mel1a and Mel1b) and androgen receptor in lung of a tropical bird, Perdicula asiatica.

The role of circulatory steroid hormone along with melatonin in lung of any seasonally breeding bird has never been explored so far. This could be interesting because steroid hormones are immunosuppressive while melatonin is immunostimulatory in nature. In our present study, we report the effect of exogenous melatonin and testosterone on expression of melatonin receptor subtypes (Mel1a and Mel1b ) and androgen receptor in lung of a tropical bird Perdicula asiatica. Birds were collected from vicinity of Varanasi and acclimatized in laboratory with sufficient food and water. The birds were treated with melatonin and testosterone at dose of 25µg/100g B.wt./day and 1mg/100g B.wt./day, respectively, for 28days. At the end of the experiment, the birds were sacrificed and lung tissue and blood sample were collected for immunohistochemistry, Western blot analysis and hormonal assay. Testosterone treatment increased circulatory testosterone and upregulated expression of androgen receptors whereas downregulated expression of melatonin receptor subtypes Mel1a and Mel1b . Melatonin administration increased peripheral melatonin and upregulated expression of melatonin receptor subtypes Mel1a and Mel1b while downregulated androgen receptor. Thus, our results suggest that a trade-off relationship between melatonin and testosterone exists in regulation of their receptors in lung of Perdicula asiatica.

Tuning melatonin receptor subtype selectivity in oxadiazolone-based analogues: Discovery of QR2 ligands and NRF2 activators with neurogenic properties

New multi-target indole and naphthalene derivatives containing the oxadiazolone scaffold as a bioisostere of the melatonin acetamido group have been developed. The novel compounds were characterized at melatonin receptors MT1R and MT2R, quinone reductase 2 (QR2), lipoxygenase-5 (LOX-5), and monoamine oxidases (MAO-A and MAO-B), and also as radical scavengers. We found that selectivity within the oxadiazolone series can be modulated by modifying the side chain functionality and co-planarity with the indole or naphthalene ring. In phenotypic assays, several oxadiazolone-based derivatives induced signalling mediated by the transcription factor NRF2 and promoted the maturation of neural stem-cells into a neuronal phenotype. Activation of NRF2 could be due to the binding of indole derivatives to KEAP1, as deduced from surface plasmon resonance (SPR) experiments. Molecular modelling studies using the crystal structures of QR2 and the KEAP1 Kelch-domain, as well as the recently described X-ray free-electron laser (XFEL) structures of chimeric MT1R and MT2R, provided a rationale for the experimental data and afforded valuable insights for future drug design endeavours.

P.351 Omega-3 fatty acids for major depressive disorder with high inflammation: A personalized approach

Epilepsy is a chronic neurologic disorder which often induces numerous adverse long-term neurologic effects, such as behavioral and cognitive deficits, increased predisposition to additional seizures, and cell injury or death. Cognitive dysfunction, depression, anxiety and sleep disorders are some of the highly prevalent and most disabling complications of epilepsy. The mechanisms that lead to the generation of epileptic comorbidities are poorly understood. Treatment for epileptic complications still remains a challenge because of the poor adherence and drug interactions associated with multi drug prescriptions and also for the fear of worsening seizures by the individual medications for complications. Melatonin, an endogenous hormone secreted by pineal gland has a prominent role in epilepsy. Agomelatine is a novel antidepressant which acts as melatonin MT1 and MT2 receptor agonist and serotonin 5Ht2C receptor antagonist. The combined action at MT1/2 and 5HT2C receptors, reduction in the depolarization-evoked release of glutamate, strong neuroprotective action and possible antioxidant properties of agomelatine could make it a potential agent in the treatment of epilepsy. The effect of agomelatine on hippocampal neuronal cell survival and neurogenesis, neuroprotective effect in hippocampus and frontal cortex and the antioxidant potential may contribute to the protective action of agomelatine against epilepsy induced memory decline. Agomelatine is proven to be an antidepressant and it has relieved anxiety symptoms and improved the quality of sleep in patients with depressive disorder. The action of agomelatine as a melatonin agonist and the consequent circadian resynchronizing property as well as its action as 5-HT2C receptor antagonist, could possibly suggest an antidepressant and anxiolytic action of agomelatine in epilepsy induced depressive behavior and anxiety. Since one of the many causes of sleep disruption in epilepsy is circadian rhythm disturbances and sleep promoting and circadian effects of melatonin is attributed to the MT1and MT2 subtypes of human melatonin receptors, agomelatine may also have a promising effect on epilepsy induced sleep disruptions. Thus with all these potential pharmacological actions, agomelatine could be recommended as a potential drug to treat epilepsy and its complications.

Melatonin suppresses milk fat synthesis by inhibiting the mTOR signaling pathway via the MT1 receptor in bovine mammary epithelial cells.

Milk fat content is an important criterion for assessing milk quality and is one of the main target traits of dairy cattle breeding. Recent studies have shown the importance of melatonin in regulating lipid metabolism, but the potential effects of melatonin on milk fat synthesis in bovine mammary epithelial cells (BMECs) remain unclear. Here, we showed that melatonin supplementation at 10μmol/L significantly downregulated the mRNA expression of lipid metabolism-related genes and resulted in lower lipid droplet formation and triglyceride accumulation. Moreover, melatonin significantly upregulated melatonin receptor subtype melatonin receptor 1a (MT1) gene expression, and the negative effects of melatonin on milk fat synthesis were reversed by treatment with the nonselective MT1/melatonin receptor subtype melatonin receptor 1b (MT2) antagonist. However, a selective MT2 antagonist did not modify the negative effects of melatonin on milk fat synthesis. In addition, KEGG analysis revealed that melatonin inhibition of milk fat synthesis may occur via the mTOR signaling pathway. Further analysis revealed that melatonin significantly suppressed the activation of the mTOR pathway by restricting the phosphorylation of mTOR, 4E-BP1, and p70S6K, and the inhibition of melatonin on milk fat synthesis was reversed by mTOR activator MHY1485 in BMECs. Furthermore, in vivo experiments in Holstein dairy cows showed that exogenous melatonin significantly decreased milk fat concentration. Our data from in vitro and in vivo studies revealed that melatonin suppresses milk fat synthesis by inhibiting the mTOR signaling pathway via the MT1 receptor in BMECs. These findings lay a foundation to identify a new potential means for melatonin to modulate the fat content of raw milk in Holstein dairy cows.

Melatonin Receptor Mel1b- and Mel1c-mediated Green Light Induced the Secretion of Growth Hormone in Anterior Pituitary of Chicks.

Previous studies have found that melatonin was related to the growth and development in avian. Therefore, the newly hatched broilers were exposed to colors of light to establish a model of pinealectomy and explored the mechanism of pineal melatonin promoting growth hormone (GH) secretion. The results showed that green light (GL) promoted the levelsof GH, pituitary-specific transcription factor-1 (Pit-1) genes and proteins in pituitary. Moreover, the mRNA and proteinlevels of melatonin receptor subtypes Mel1b and Mel1c in the pituitary in GL were higher than other monochromatic light groups. After pinealectomy, the expression of Pit-1, GH, Mel1b and Mel1c in pituitary decreased. In vitro, exogenous melatonin promoted the level of Pit-1 mRNA and the secretion of GH in anterior pituitary cells. However, when melatonin was added with exogenous selective Mel1b antagonist (4-phenyl-2-propionamideotetralin) and selective Mel1c antagonist (prazosin), the levelof Pit-1 mRNA and the GH secretion decreased. When selective Mel1b and Mel1c antagonists were added simultaneously, the decrease in Pit-1 mRNA and GH secretion was more significant. These results indicated that pineal melatonin promotes the expression of Pit-1 under GL by binding to melatonin receptor subtypes Mel1b and Mel1c in the pituitary, thereby increasing GH secretion and promoting the growth.

XFEL structures of the human MT2 melatonin receptor reveal the basis of subtype selectivity.

The human MT11 and MT22 melatonin receptors are G protein-coupled receptors (GPCRs) involved in the regulation of circadian rhythm and sleep patterns3. Drug development efforts target both receptors for treatment of insomnia, circadian rhythm and mood disorders, and cancer3, while MT2 has also been implicated in type 2 diabetes (T2D)4,5. Here we report the X-ray Free Electron Laser (XFEL) structures of the human MT2 receptor in complex with agonists 2-phenylmelatonin (2-pmt) and ramelteon6 at resolutions of 2.8 Å and 3.3 Å, respectively, along with two structures of function-related mutants, H2085.46A (superscripts represent the Ballesteros-Weinstein residue numbering nomenclature7) and N862.50D, obtained in complex with 2-pmt. Comparison of the MT2 structures with MT18 reveals that, despite the fact that the orthosteric ligand-binding site residues are conserved, there are notable conformational variations as well as differences in [3H]-melatonin dissociation kinetics that provide new insights into the selectivity between melatonin receptor subtypes. In addition to the membrane-buried lateral ligand entry channel that is also observed in MT1, the MT2 structures reveal a narrow opening towards the solvent in the extracellular part of the receptor. We provide functional and kinetic data supporting a prominent role for the intramembrane ligand entry in both receptors, while simultaneously suggesting the possibility of an extracellular entry path in MT2. Our findings contribute to a molecular understanding of melatonin receptor subtype selectivity and ligand access modes, which are essential for the design of highly selective melatonin tool compounds and therapeutic agents.

Melatonin receptor expression in vocal, auditory, and neuroendocrine centers of a highly vocal fish, the plainfin midshipman (Porichthys notatus).

Melatonin plays a central role in entraining activity to the day-night cycle in vertebrates. Here, we investigate neuroanatomical substrates of melatonin-dependent vocal-acoustic behavior in the nocturnal and highly vocal teleost fish, the plainfin midshipman (Porichthys notatus). Using in situ hybridization (ISH) and quantitative real-time PCR (qPCR), we assess the mRNA distribution and transcript abundance of melatonin receptor subtype 1B (mel1b), shown to be important for vocalization in midshipman fish and songbirds. ISH shows robust mel1b expression in major nodes of the central vocal and auditory networks in the subpallium, preoptic area (POA), anterior hypothalamus, dorsal thalamus, posterior tuberculum, midbrain torus semicircularis and periaqueductal gray, and hindbrain. Mel1b label is also abundant in secondary targets of the olfactory, visual, and lateral line systems, as well as telencephalic regions that have been compared to the amygdala, extended amygdala, striatum, septum, and hippocampus of tetrapods. Q-PCR corroborates mel1b abundance throughout the brain and shows significant increases in the morning compared with nighttime in tissue samples inclusive of the telencephalon and POA, but remains stable in other brain regions. Plasma melatonin levels show expected increase at night. Our findings support the hypothesis that melatonin's stimulatory effects on vocal-acoustic mechanisms in midshipman is mediated, in part, by melatonin binding in vocal, auditory, and neuroendocrine centers. Together with robust mel1b expression in multiple telencephalic nuclei and sensory systems, the results further indicate an expression pattern comparable to that in birds and mammals that is indicative of melatonin's broad involvement in the modulation of physiology and behavior.

Sleep well. Untangling the role of melatonin MT1 and MT2 receptors in sleep.

The pharmacological potential of targeting selectively melatonin MT1 or MT2 receptors has not yet been exploited in medicine. Research using selective MT1/MT2 receptor ligands and MT1/MT2 receptor knockout mice has indicated that the activation of MT2 receptors selectively increases non-rapid eye movement (NREM) sleep whereas MT1 receptors seem mostly implicated in the regulation of REM sleep. Moreover, MT1 knockout mice show an increase in NREM sleep, while MT2 knockout a decrease, suggesting an opposite role of these two receptors. A recent paper in mice by Sharma et al (J Pineal Res, 2018, e12498) found that MT1 but not MT2 receptors are expressed on orexin neurons in the perifornical lateral hypothalamus (PFH). Moreover, after injecting melatonin or luzindole into the mouse PFH, the authors suggest that melatonin promotes NREM sleep because activates PFH MT1 receptors, which in turn inhibit orexin neurons that are important in promoting arousal and maintaining wakefulness. In this commentary, we have critically commented on some of these findings on the bases of previous literature. In addition, we highlighted the fact that no conclusions could be drawn on the melatonin receptor subtype mediating the effects of melatonin on sleep because the authors used the non-selective MT1/MT2 receptors antagonist luzindole. More solid research should further characterize the pharmacological function of these two melatonin receptors in sleep.

Differential Regulation of Cell Proliferation and Apoptosis by Melatonin Receptor Subtype-Signaling in the Adult Murine Brain

Background/Aims: Zeitgeber time (ZT)-dependent changes in cell proliferation and apoptosis are regulated by melatonin receptor (MT)-mediated signaling in the adult hippocampus and hypothalamic-hypophyseal system. There are two G-protein-coupled MT subtypes, MT1 and MT2. Therefore, the present study examined which MT subtype is required for the regulation of ZT-dependent changes in cell proliferation and/or apoptosis in the adult murine brain and pituitary. Methods: Adult melatonin-proficient (C3H) mice with targeted deletion of MT1 (MT1 KO) or MT2 (MT2 KO) were adapted to a 12-h light/12-h dark photoperiod and sacrificed at ZT00, ZT06, ZT12, and ZT18. Immunohistochemistry for Ki67 or activated caspase-3 served to quantify proliferating and apoptotic cells in the hippocampal subgranular zone (SGZ) and granule cell layer, the hypothalamic median eminence (ME), and the hypophyseal pars tuberalis. Results: ZT-dependent changes in cell proliferation were found exclusively in the SGZ and ME of MT1 KO mice, while apoptosis showed no ZT-dependent changes in the regions analyzed, neither in MT1 nor in MT2 KO mice. Comparison with our previous studies in C3H mice with functional MTs and MT1/2 KO mice revealed that MT2-mediated signaling is required and sufficient for ZT-dependent changes in cell proliferation in the SGZ and ME, while ZT-dependent changes in apoptosis require signaling from both MT subtypes. Conclusions: Our results indicate that generation and timing of ZT-dependent changes in cell proliferation and apoptosis by melatonin require different MT subtype constellations and emphasize the importance to shed light on the specific function of each receptor subtype in different tissues and physiological conditions.

Seasonal Reproduction in Vertebrates: Melatonin Synthesis, Binding, and Functionality Using Tinbergen's Four Questions.

One of the many functions of melatonin in vertebrates is seasonal reproductive timing. Longer nights in winter correspond to an extended duration of melatonin secretion. The purpose of this review is to discuss melatonin synthesis, receptor subtypes, and function in the context of seasonality across vertebrates. We conclude with Tinbergen’s Four Questions to create a comparative framework for future melatonin research in the context of seasonal reproduction.