RESUMO
Melatonin is an ancient multi-tasking molecule produced by the pineal gland and by several extrapineal tissues. A variety of activities has been ascribed to this hormone in different physiological and pathological contexts, but little is known about its role in peripheral neuroregeneration. Here, we have exploited two different types of injury to test the capability of melatonin to stimulate regeneration of motor axons: (a) the acute and reversible presynaptic degeneration induced by the spider neurotoxin α-Latrotoxin and (b) the compression/transection of the sciatic nerve. We found that in both cases melatonin administration accelerates the process of nerve repair. This pro-regenerative action is MT1 -mediated, and at least in part due to a sustained activation of the ERK1/2 pathway. These findings reveal a receptor-mediated, pro-regenerative action of melatonin in vivo that holds important clinical implications, as it posits melatonin as a safe candidate molecule for the treatment of a number of peripheral neurodegenerative conditions.
Assuntos
Axônios/efeitos dos fármacos , Melatonina/farmacologia , Neurônios Motores/efeitos dos fármacos , Regeneração Nervosa/efeitos dos fármacos , Traumatismos dos Nervos Periféricos/tratamento farmacológico , Receptor MT1 de Melatonina/agonistas , Nervo Isquiático/efeitos dos fármacos , Animais , Axônios/metabolismo , Axônios/patologia , Células Cultivadas , Modelos Animais de Doenças , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neurônios Motores/metabolismo , Neurônios Motores/patologia , Traumatismos dos Nervos Periféricos/metabolismo , Traumatismos dos Nervos Periféricos/patologia , Fosforilação , Ratos Wistar , Receptor MT1 de Melatonina/metabolismo , Nervo Isquiático/lesões , Nervo Isquiático/metabolismo , Nervo Isquiático/patologia , Transdução de Sinais , Venenos de Aranha/toxicidade , Fatores de TempoRESUMO
Melatonin (MLT) levels fluctuate according to the external light/dark cycle in both diurnal and nocturnal mammals. We previously demonstrated that melatonin MT2 receptor knockout (MT2 -/- ) mice show a decreased nonrapid eye movement sleep over 24 hours and increased wakefulness during the inactive (light) phase. Here, we investigated the role of MT2 receptors in physiological light/dark cycle fluctuations in the activity of dorsal raphe nucleus (DRN) serotonin (5-HT) neurons and anxiety- and depression-like behavior. We found that the 5-HT burst-firing activity was tonically reduced across the whole 24 hours in MT2 -/- mice compared with MT2 +/+ mice. Importantly, the physiological changes in the spontaneous firing activity of DRN 5-HT neurons during the light/dark cycle were nullified in MT2 -/- mice, with a higher DRN 5-HT neural firing activity during the light phase in MT2 -/- than in MT2 +/+ mice. The role of MT2 receptors over DRN 5-HT neurons was confirmed by acute pharmacological studies in which the selective MT2 receptors agonist UCM1014 dose dependently inhibited DRN 5-HT activity, mostly during the dark phase. Compared with MT2 +/+ , MT2 -/- mice displayed an anxiety-like phenotype in the novelty-suppressed feeding and in the light/dark box tests; while anxiety levels in the light/dark box test were lower during the dark than during the light phase in MT2 +/+ mice, the opposite was seen in MT2 -/- mice. No differences between MT2 +/+ and MT2 -/- mice were observed for depression-like behavior in the forced swim and in the sucrose preference tests. These results suggest that MT2 receptor genetic inactivation impacts 5-HT neurotransmission and interferes with anxiety levels by perturbing the physiologic light/dark pattern.
Assuntos
Comportamento Animal , Ritmo Circadiano , Emoções , Receptor MT2 de Melatonina/deficiência , Neurônios Serotoninérgicos/metabolismo , Serotonina/metabolismo , Sono REM , Animais , Camundongos , Camundongos Knockout , Receptor MT2 de Melatonina/metabolismo , Serotonina/genéticaRESUMO
Melatonin (MLT) is widely used to treat sleep disorders although the underlying mechanism is still elusive. In mice, using wheel-running detection, we found that exogenous MLT could completely recover the period length prolonged by N-methyl-D-aspartate receptor (NMDAR) impairment due to the injection of the NMDAR antagonist MK-801, a preclinical model of psychosis. The analysis of the possible underlying mechanisms indicated that MLT could regulate the homeostatic state in the ventrolateral preoptic nucleus (VLPO) instead of the circadian process in the suprachiasmatic nucleus (SCN). In addition, our data showed that MK-801 decreased Ca2+ -related CaMKII expression and CREB phosphorylation levels in the VLPO, and MLT could rescue these intracellular impairments but not NMDAR expression levels. Accordingly, Gcamp6 AAV virus was injected in-vivo to further monitor intracellular Ca2+ levels in the VLPO, and MLT demonstrated a unique ability to increase Ca2+ fluorescence compared with MK-801-injected mice. Additionally, using the selective melatonin MT2 receptor antagonist 4-phenyl-2-propionamidotetralin (4P-PDOT), we discovered that the pharmacological effects of MLT upon NMDAR impairments were mediated by melatonin MT2 receptors. Using electroencephalography/electromyography (EEG/EMG) recordings, we observed that the latency to the first nonrapid eye movement (NREM) sleep episode was delayed by MK-801, and MLT was able to recover this delay. In conclusion, exogenous MLT by acting upon melatonin MT2 receptors rescues sleep phase delayed by NMDAR impairment via increasing intracellular Ca2+ signaling in the VLPO, suggesting a regulatory role of the neurohormone on the homeostatic system.
Assuntos
Sinalização do Cálcio/efeitos dos fármacos , Proteína Quinase Tipo 2 Dependente de Cálcio-Calmodulina/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Maleato de Dizocilpina/farmacologia , Melatonina/farmacologia , Área Pré-Óptica/metabolismo , Receptor MT2 de Melatonina/metabolismo , Fases do Sono/efeitos dos fármacos , Animais , Eletroencefalografia , Eletromiografia , Masculino , Melatonina/metabolismo , CamundongosRESUMO
A series of dimeric melatonin analogues 3a-e obtained by connecting two melatonin molecules through the methoxy oxygen atoms with spacers spanning 16-24 atoms and the agomelatine dimer 7 were synthesized and characterized in 2-[125-I]-iodomelatonin binding assays, bioluminescence resonance energy transfer (BRET) experiments, and in functional cAMP and ß-arrestin recruitment assays at MT1 and MT2 receptors. The binding affinity of 3a-e generally increased with increasing linker length. Bivalent ligands 3a-e increased BRET signals of MT1 dimers up to 3-fold compared to the monomeric control ligand indicating the simultaneous binding of the two pharmacophores to dimeric receptors. Bivalent ligands 3c and 7 exhibited important changes in functional properties on the Gi/cAMP pathway but not on the ß-arrestin pathway compared to their monomeric counterparts. Interestingly, 3c (20 atoms spacer) shows inverse agonistic properties at MT2 on the Gi/cAMP pathway. In conclusion, these findings indicate that O-linked melatonin dimers are promising tools to develop signaling pathway-based bivalent melatonin receptor ligands.
Assuntos
Melatonina/análogos & derivados , Melatonina/farmacologia , Receptor MT1 de Melatonina/agonistas , Receptor MT2 de Melatonina/agonistas , Técnicas de Transferência de Energia por Ressonância de Bioluminescência , AMP Cíclico/metabolismo , Agonismo Inverso de Drogas , Células HEK293 , Humanos , Ligantes , Melatonina/metabolismo , Estrutura Molecular , Multimerização Proteica/efeitos dos fármacos , Receptor MT1 de Melatonina/metabolismo , Receptor MT2 de Melatonina/metabolismo , beta-Arrestinas/metabolismoRESUMO
Melatonin (MLT) is a neurohormone that regulates many physiological functions including sleep, pain, thermoregulation, and circadian rhythms. MLT acts mainly through two G-protein-coupled receptors named MT1 and MT2, but also through an MLT type-3 receptor (MT3). However, the role of MLT receptor subtypes in thermoregulation is still unknown. We have thus investigated the effects of selective and non-selective MLT receptor agonists/antagonists on body temperature (Tb) in rats across the 12/12-h light-dark cycle. Rectal temperature was measured every 15 min from 4:00 a.m. to 9:30 a.m. and from 4:00 p.m. to 9:30 p.m., following subcutaneous injection of each compound at either 5:00 a.m. or 5:00 p.m. MLT (40 mg/kg) had no effect when injected at 5 a.m., whereas it decreased Tb during the light phase only when injected at 5:00 p.m. This effect was blocked by the selective MT2 receptor antagonist 4P-PDOT and the non-selective MT1/MT2 receptor antagonist, luzindole, but not by the α1/MT3 receptors antagonist prazosin. However, unlike MLT, neither the selective MT1 receptor partial agonist UCM871 (14 mg/kg) nor the selective MT2 partial agonist UCM924 (40 mg/kg) altered Tb during the light phase. In contrast, UCM871 injected at 5:00 p.m. increased Tb at the beginning of the dark phase, whereas UCM924 injected at 5:00 a.m. decreased Tb at the end of the dark phase. These effects were blocked by luzindole and 4P-PDOT, respectively. The MT3 receptor agonist GR135531 (10 mg/kg) did not affect Tb. These data suggest that the simultaneous activation of both MT1 and MT2 receptors is necessary to regulate Tb during the light phase, whereas in a complex but yet unknown manner, they regulate Tb differently during the dark phase. Overall, MT1 and MT2 receptors display complementary but also distinct roles in modulating circadian fluctuations of Tb.
Assuntos
Temperatura Corporal/efeitos dos fármacos , Melatonina/administração & dosagem , Receptor MT1 de Melatonina/agonistas , Receptor MT1 de Melatonina/metabolismo , Receptor MT2 de Melatonina/agonistas , Acetamidas/administração & dosagem , Acetamidas/farmacologia , Compostos de Anilina/administração & dosagem , Compostos de Anilina/farmacologia , Animais , Injeções Subcutâneas , Masculino , Melatonina/farmacologia , Fotoperíodo , Ratos , Ratos Wistar , Receptor MT1 de Melatonina/antagonistas & inibidores , Receptor MT2 de Melatonina/antagonistas & inibidores , Receptor MT2 de Melatonina/metabolismo , Tetra-Hidronaftalenos/administração & dosagem , Tetra-Hidronaftalenos/farmacologia , Triptaminas/administração & dosagem , Triptaminas/farmacologiaRESUMO
A series of new pyrimidine thioethers, recognized as the key intermediates in the synthesis of S-DABO antivirals, were prepared and evaluated both in vivo and in silico. The purpose of this evaluation was to find novel structural analogues of the known antihypoxic drug Isothiobarbamine endowed with improved pharmacological profile. The in vivo studies led to the identification of compounds 5c, 5e, and 5f endowed with antidepressant/anxiolytic, performance enhancing, and nootropic properties. Compounds 5c and 5f were further tested in mice affected by social depression and were able to increase motor and tentative search activity compared to control groups, along with higher interaction frequency and better results in a sucrose preference test. Overall, these data suggested a better psychoemotional state of the animals, treated with compounds 5c, and 5f. Moreover, 5c and 5f exhibited minimal acute toxicity, lower than Fluoxetine hydrochloride. Molecular modelling studies finally indicated the plausible biomolecular mechanism of action of compounds 5c, 5e, and 5f, which seem to bind GABA-A, melatonin, and sigma-1 receptors. Moreover, three-dimensional structure-activity relationships enabled to define a SAR model that will be of great utility for the design of further structurally optimized compounds of the above mentioned chemotype.
Assuntos
Ansiolíticos , Nootrópicos , Animais , Camundongos , Ansiolíticos/farmacologia , Sulfetos , Antidepressivos/farmacologia , PirimidinasRESUMO
In crystal structures of melatonin MT1 and MT2 receptors, a lipophilic subpocket has been characterized which accommodates the phenyl ring of the potent agonist 2-phenylmelatonin. This subpocket appears a key structural element to achieve high binding affinity and selectivity for the MT2 receptor. A series of 2-arylindole ligands was synthesized to probe the requirements for the optimal occupation and interaction with the 2-phenyl binding pocket. Thermodynamic integration simulations applied to MT1 and MT2 receptors in complex with the α-naphthyl derivative provided a rationale for the MT2-selectivity and investigation on the binding mode of a couple of atropisomers allowed to define the available space and arrangement of substituents inside the subpocket. Interestingly, more hydrophilic 2-aza-substituted compounds displayed high binding affinity and molecular dynamics simulations highlighted polar interaction with residues from the subpocket that could be responsible for their potency.
Assuntos
Melatonina , Receptor MT1 de Melatonina , Receptor MT2 de Melatonina , Ligantes , Melatonina/análogos & derivados , Melatonina/química , Melatonina/metabolismo , Simulação de Dinâmica Molecular , Receptor MT1 de Melatonina/química , Receptor MT1 de Melatonina/metabolismo , Receptor MT2 de Melatonina/química , Receptor MT2 de Melatonina/metabolismoRESUMO
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.
Assuntos
Transtorno Depressivo Maior , Ketamina , Melatonina , Anedonia , Animais , Ritmo Circadiano , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Reboxetina , Receptor MT1 de Melatonina , Receptor MT2 de Melatonina , TemperaturaRESUMO
BACKGROUND: Experiments in the late nineties showed an inverse relationship in the eye levels of melatonin and dopamine, thereby constituting an example of eye parameters that are prone to circadian variations. The underlying mechanisms are not known but these relevant molecules act via specific cell surface dopamine and melatonin receptors. This study investigated whether these receptors formed heteromers whose function impact on eye physiology. We performed biophysical assays to identify interactions in heterologous systems. Particular heteromer functionality was detected using Gi coupling, MAPK activation, and label-free assays. The expression of the heteroreceptor complexes was assessed using proximity ligation assays in cells producing the aqueous humor and human eye samples. Dopamine D3 receptors (D3Rs) were identified in eye ciliary body epithelial cells. We discovered heteromers formed by D3R and either MT1 (MT1R) or MT2 (MT2R) melatonin receptors. Heteromerization led to the blockade of D3R-Gi coupling and regulation of signaling to the MAPK pathway. Heteromer expression was negatively correlated with intraocular hypertension. CONCLUSIONS: Heteromers likely mediate melatonin and dopamine actions in structures regulating intraocular pressure. Significant expression of D3R-MT1R and D3R-MT1R was associated with normotensive conditions, whereas expression diminished in a cell model of hypertension. A clear trend of expression reduction was observed in samples from glaucoma cases. The trend was marked but no statistical analysis was possible as the number of available eyes was 2.
Assuntos
Corpo Ciliar/metabolismo , Células Epiteliais/metabolismo , Glaucoma/patologia , Hipertensão Ocular/patologia , Receptor MT1 de Melatonina/metabolismo , Receptor MT2 de Melatonina/metabolismo , Receptores de Dopamina D3/metabolismo , Estudos de Casos e Controles , Glaucoma/metabolismo , Células HEK293 , Humanos , Hipertensão Ocular/metabolismo , Multimerização ProteicaRESUMO
The main objective of this study was to investigate potential effectiveness of agomelatine pretreatment in the prevention of diabetes itself and encephalopathy, with a focus on brain tissue oxidative stress and inflammatory processes in streptozotocin (STZ)-induced diabetic mice. Interleukine-1ß (IL-1ß) and TACR1 (NK1), which is a tachykinine receptor, were used for the investigation of inflammation in the brain regions including raphe nucleus, periaqueductal gyrus (PAG), amygdala, and nucleus accumbens. The effects of agomelatine on total antioxidant capacity were also evaluated. In the in vitro part of the study, the effects of agomelatine on cell viability were investigated in dorsal root ganglion (DRG) neurons. Fasting blood glucose levels were measured 72 h after STZ injection to determine the diabetic condition. Agomelatine pretreatment prevented both hyperglycemia and hypoinsulinemia in STZ-treated mice. When STZ was injected to induce diabetes in mice, neither hyperglycemia nor hypoinsulinemia was developed in agomelatine pretreated mice and 6 weeks after development of diabetes, agomelatine treatment significantly decreased levels of IL-1ß mRNA in raphe nucleus and nucleus accumbens. TACR1 mRNA levels were lower in raphe nucleus, PAG, and amygdala of agomelatine-treated diabetic mice. The increase in total antioxidant capacity after agomelatine administration may responsible for its beneficial effect in the prevention of diabetes. We showed that agomelatine reversed high glucose-induced cell viability decreases in DRG neurons. Both the antihyperglycemic and antioxidant effects of agomelatine might have contributed to the DRG neuron viability improvement. In conclusion, agomelatine seems to both prevent development of diabetes and reverse the encephalopathic changes caused by diabetes.
Assuntos
Acetamidas/farmacologia , Glicemia/efeitos dos fármacos , Encefalopatias/prevenção & controle , Encéfalo/efeitos dos fármacos , Diabetes Mellitus Experimental/prevenção & controle , Hiperglicemia/prevenção & controle , Hipoglicemiantes/farmacologia , Insulina/sangue , Fármacos Neuroprotetores/farmacologia , Animais , Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Biomarcadores/sangue , Glicemia/metabolismo , Encéfalo/metabolismo , Encéfalo/patologia , Encefalopatias/sangue , Encefalopatias/induzido quimicamente , Encefalopatias/patologia , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/genética , Gânglios Espinais/efeitos dos fármacos , Gânglios Espinais/metabolismo , Gânglios Espinais/patologia , Hiperglicemia/sangue , Hiperglicemia/induzido quimicamente , Hiperglicemia/genética , Mediadores da Inflamação/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/metabolismo , Masculino , Camundongos Endogâmicos BALB C , Ratos Wistar , Receptores da Neurocinina-1/genética , Receptores da Neurocinina-1/metabolismo , EstreptozocinaRESUMO
Melatonin (MLT) has been implicated in several pathophysiological states, including pain. MLT mostly activates two G-protein coupled receptors, MT1 and MT2. In this review, we present the analgesic properties of MLT in preclinical and clinical studies, giving particular emphasis to the effects mediated by MT2 receptors and to recent investigations demonstrating the analgesic effects of MT2 receptor partial agonists in chronic and acute/inflammatory pain conditions. MT2 receptors are localized in specific brain areas, including the reticular and the ventromedial nuclei of the thalamus (part of the ascending nociceptive pathway) and the ventrolateral periaqueductal grey matter (vlPAG) (part of the descending antinociceptive pathway). MLT displays analgesic properties in several animal paradigms of chronic, acute, inflammatory and neuropathic pain; importantly, these effects are mediated by MT2 receptors since they are blocked by selective MT2 antagonists. In different pain paradigms, UCM924 and UCM765, two selective MT2 receptor partial agonists, produce analgesic effects with higher potency than MLT, thus confirming the involvement of MT2 receptors in pain. Notably, these compounds do not induce sedation and motor impairments. Although their analgesic mechanism of action is not yet completely elucidated, they act on antinociceptive descending pathways by stimulating MT2 receptors on glutamatergic neurons of the vlPAG, which in turn activate OFF cells and inhibit ON cells of the rostral ventromedial medulla (RVM). Collectively, there is strong preclinical evidence suggesting the pharmacological potential of MT2 receptor partial agonists, which also have a favorable toxicological profile. These compounds may be further developed as novel analgesic drugs.
Assuntos
Analgésicos/farmacologia , Inflamação/tratamento farmacológico , Melatonina/farmacologia , Neuralgia/tratamento farmacológico , Receptor MT2 de Melatonina/agonistas , Acetamidas/farmacologia , Compostos de Anilina/farmacologia , Animais , Encéfalo/metabolismo , Encéfalo/fisiopatologia , Estudos Clínicos como Assunto , Humanos , Inflamação/fisiopatologia , Melatonina/fisiologia , Neuralgia/fisiopatologia , Receptor MT2 de Melatonina/fisiologia , Medula Espinal/metabolismo , Medula Espinal/fisiopatologiaRESUMO
We have reported that clinically relevant concentrations of valproic acid (VPA) upregulate the G protein-coupled melatonin MT1 receptor in rat C6 glioma cells, and both MT1 and MT2 receptors in the rat hippocampus. The melatonin MT2 receptor is relatively enriched in the hippocampus, where it is thought to be involved in modulating synaptic plasticity and cognitive function. Importantly, a significant decrease in MT2 expression has been observed in the hippocampus of Alzheimer's patients. Therefore, we examined whether the global upregulation of this receptor (and also the MT1) by VPA, observed in earlier RT-PCR and real time PCR studies, could be localized to more discrete hippocampal regions, which are involved in cognitive function. In situ hybridization of rat brain slices, following chronic VPA treatment (3mg/mL or 4mg/mL in drinking water), revealed a significant upregulation of the MT2 receptor mRNA in the CA1, CA2, CA3 and dentate gyrus (DG) regions of the rat hippocampus. In contrast, the MT1 receptor was not detected in the hippocampus by in situ hybridization. The significant induction of melatonin MT2 receptor expression by VPA in hippocampal regions involved in learning, memory and/or neural stem cell proliferation, suggests that a combinatorial therapeutic strategy involving VPA together with melatonin or other MT2 agonists, would be beneficial in neurodegenerative disorders such as Alzheimer's disease.