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The role of stress in the etiology of depression has been largely reported. In this line, exogenous glucocorticoids are employed to mimic the influence of stress on the development of depression. The N/OFQ-NOP receptor system has been implicated in the modulation of stress and emotional behaviors. In fact, the blockade of NOP receptors induces antidepressant effects and increases resilience to acute stress. This study investigated the effects of the NOP receptor blockade on dexamethasone-treated mice exposed to acute and prolonged swimming stress. Swiss and NOP(+/+) and NOP(−/−) mice were treated with dexamethasone, and the protective effects of the NOP antagonist SB-612111 (10 mg/kg, ip) or imipramine (20 mg/kg, ip) were investigated in three swimming sessions. The re-exposure to swim stress increased immobility time in Swiss and NOP(+/+), but not in NOP(−/−) mice. Acute and repeated dexamethasone administration induced a further increase in the immobility time, and facilitated body weight loss in Swiss mice. Single administration of SB-612111, but not imipramine, prevented swimming stress- and dexamethasone-induced increase in the immobility time. Repeated administrations of SB-612111 prevented the deleterious effects of 5 days of dexamethasone treatment. Imipramine also partially prevented the effects of repeated glucocorticoid administration on the immobility time, but did not affect the body weight loss. NOP(−/−) mice were more resistant than NOP(+/+) mice to inescapable swimming stress, but not dexamethasone-induced increase in the immobility time and body weight loss. In conclusion, the blockade of the NOP receptor facilitates an active stress copying response and attenuates body weight loss due to repeated stress.

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES A depressão é um distúrbio sério e comum em todo mundo e sua recorrência está intimamente ligada a prejuízos na vida dos indivíduos que a portam. As suas principais características, tais como humor deprimido, anedonia, irritabilidade, dificuldades de concentração, fadiga, aumento ou diminuição do apetite, insônia ou hipersonia e prejuízos cognitivos e locomotores, justificam a sua gravidade. Os tratamentos atuais causam muitos efeitos colaterais e demoram para iniciar a produzir o efeito terapêutico, o que apoia a necessidade para o desenvolvimento de novos fármacos. Nas últimas duas décadas vários estudos têm mostrado que o receptor NOP pode ser um alvo terapêutico valioso no campo da terapia farmacológica e seu antagonismo tem se mostrado uma estratégia promissora devido aos efeitos antidepressivos relatados em estudos pré-clínicos e clínicos. Algumas ferramentas farmacológicas endereçadas ao NOP têm sido desenvolvidas, tais como SB–612111 e o composto 35 (C–35). Nesse sentido, o presente trabalho tem o objetivo avaliar, por meio de técnicas de simulação computacional, utilizando a Teoria do Funcional da Densidade (DFT) e do Método de Fracionamento Molecular com Capas Conjugadas (MFCC), as particularidades energéticas presentes na interação entre o receptor NOP e o ligante SB-612111 e o ligante C-35, que são antagonistas. A partir da obtenção das estruturas cristalinas no PDB e da utilização das ferramentas inerentes ao campo da simulação computacional, foi possível analisar as contribuições energéticas presentes em cada complexo formado por esses ligantes e o receptor em questão. No total foram observados 102 (101) fragmentos de aminoácidos para SB (C-35), sendo que a maioria dos resíduos energeticamente mais relevantes fazem parte do bolsão hidrofóbico de ligação e os fragmentos com maior energia atrativa estão situados a uma distância de até 3 Å, em ambos os complexos: NOP/SB (ASP130 > GLN107 > TYR309 > TYR131); NOP/C-35 (ASP130 > GLN107 > ASP110 > ILE127 > TYR131 > GLN280 > TYR309). Os resultados aqui apresentados e discutidos possuem uma relevância substancial, dado que nas últimas décadas, principalmente nos últimos anos, tem ocorrido uma intensa investigação e investimento na busca do desenvolvimento de antidepressivos mais eficazes. Portanto, análises como aqui descritas podem auxiliar no avanço do cenário farmacológico direcionado aos antidepressivos, tendo como alvo o receptor NOP. Depression is a serious and common disorder throughout the world and its recurrence is closely linked to harm in the lives of the individuals who carry it. Its main characteristics, such as depressed mood, anhedonia, irritability, concentration difficulties, fatigue, increased or decreased appetite, insomnia or hypersomnia, and cognitive and locomotor losses, justify its gravity. Current treatments cause many side effects and take a long time to start producing the therapeutic effect, which supports the need for the development of new drugs. In the last two decades, several studies have shown that the NOP receptor can be a valuable therapeutic target in the field of pharmacological therapy and its antagonism has shown to be a promising strategy due to the antidepressant effects reported in preclinical and clinical studies. Some pharmacological tools addressing to NOP have been developed, such as SB-612111 and compound 35 (C-35). In this sense, the present work aims to evaluate, by means of computer simulation techniques, using the Functional Density Theory (DFT) and the Molecular Fractionation Method with Conjugate Covers (MFCC), the energetic particularities present in the interaction between the NOP receptor and the SB-612111 ligand and the C-35 ligand, which are antagonists. From obtaining the crystalline structures in the PDB and using the tools inherent to the field of computational simulation, it was possible to analyze the energetic contributions present in each complex formed by these binders and the receptor in question. In total 102 (101) amino acid fragments for SB (C-35) were observed, with most of the most energetically relevant residues being part of the hydrophobic bonding pocket and the fragments with the most attractive energy are located at a distance of up to 3 Å, in both complexes: NOPSB (ASP130 > GLN107 > TYR309 > TYR131); NOP-C-35 (ASP130 > GLN107 > ASP110 > ILE127 > TYR131 > GLN280 > TYR309). The results presented and discussed here are of substantial relevance, given that in recent decades, especially in recent years, there has been intense research and investment in the search for the development of more effective antidepressants. Therefore, analyses as described here can help in the advancement of the pharmacological scenario directed to antidepressants, targeting the NOP receptor.

A fibromialgia ? caracterizada por dor generalizada, sendo acompanhada por dist?rbios funcionais e afetivos. Este estudo avaliou a implica??o do receptor do pept?deo nociceptina/orfanina FQ (NOPr) em um modelo murino de fibromialgia. Os protocolos experimentais foram aprovados pela Comiss?o de ?tica no Uso de Animais (CEUA/PUCRS 15/00487). A fibromialgia foi induzida em camundongos f?meas CF-1 (20-24 g, 4 semanas de idade) pela administra??o de reserpina (0,25 mg/kg; via subcut?nea), uma vez ao dia, durante 3 dias consecutivos. Grupos controle receberam ve?culo. No quarto dia, os camundongos foram tratados com uma dose ?nica de nociceptina (N/OFQ) ou, do antagonista pept?dico seletivo, UFP-101, administrados por via intraperitoneal (i.p., 0,3-5 nmol/kg), intracerebroventricular (i.c.v., 0,3 -1 nmol/s?tio) ou intratecal (i.t., 0,3-5 nmol/s?tio), 30 min antes das sess?es experimentais. Em outra s?rie de experimentos, os animais foram tratados com o antagonista UFP-101 (1 nmol/kg) ou, com o antagonista n?o pept?dico, SB-612111 (6,6 ?mol/kg), administrados por via intraperitoneal, durante tr?s dias consecutivos, 30 min ap?s a inje??o di?ria de reserpina. No 4? dia, os animais tamb?m receberam um dos antagonistas, 30 min antes das avalia??es comportamentais. Os animais foram submetidos aos testes de Von Frey, placa quente, nado for?ado, labirinto em cruz elevado, rotarod e de preens?o palmar. A express?o da pr?-pr?-nociceptina ppN/OFQ e do NOPr foi determinada por RT-qPCR e imunoistoqu?mica. O microPET [18F]-FDG foi utilizado para avaliar os padr?es de ativa??o cerebral em camundongos tratados com reserpina. A distribui??o do tamanho das fibras musculares do masseter e do gastrocn?mio foi avaliada atrav?s de an?lise histol?gica. A ?rea e densidade das mitoc?ndrias no m?sculo esquel?tico foram analisadas por microscopia eletr?nica de transmiss?o. Nos protocolos de tratamentos agudos, a administra??o i.t. ou i.p. de N/OFQ (1 nmol/s?tio ou 1 nmol/kg, respectivamente) reduziu significativamente a alod?nia mec?nica induzida pela reserpina. Contudo, a administra??o i.p. de N/OFQ, na dose de 5 nmol/kg, teve efeito oposto, induzindo hipernocicep??o. Em rela??o aos efeitos agudos do UFP-101, este antagonista pept?dico, administrado pelas vias i.c.v. (1 nmol/s?tio), i.t. (3 e 5 nmol/s?tio) ou i.p. (1, 3 e 5 nmol/kg), reduziu significativamente a hipersensibilidade mec?nica em camundongos tratados com reserpina. O tratamento agudo com N/OFQ ou UFP 101 n?o alterou significativamente a hipersensibilidade t?rmica, pelas vias i.c.v. ou i.p. A administra??o i.t. de N/OFQ (3 nmol/s?tio) ou de UFP-101 (5 nmol/s?tio) teve um efeito inibit?rio significativo na nocicep??o t?rmica. No teste da nata??o for?ada, a reserpina elevou o tempo de imobilidade, e este foi inibido de forma significativa pela N/OFQ, administrada pelas vias i.c.v (1 nmol/s?tio) ou i.t. (3 nmol/s?tio). N/OFQ e UFP-101 n?o modificaram nenhum par?metro relacionado ? ansiedade. Os tratamentos repetidos com UFP-101 e com SB-612111 reduziram a alod?nia mec?nica (37 ? 8% e 43 ? 15,2%), a hipernocicep??o t?rmica (32,2 ? 5% e 45 ? 17,5%), melhoraram a coordena??o motora no rotarod (aumento de 7 e 2 vezes no tempo de perman?ncia) e a for?a de preens?o palmar (15 ? 16% e 9 ? 5.5%), respectivamente. A administra??o de ambos antagonistas n?o foi capaz de alterar par?metros de ansiedade ou depress?o. A fibromialgia induzida por reserpina foi associada ao aumento na express?o de RNAm para a ppN/OFQ na medula espinhal lombar (dia 3) e no masseter (dias 1 e 2), enquanto a express?o do RNAm do NOPr foi aumentada no m?sculo masseter (dia 1). Alternativamente, a express?o de RNAm do NOPr foi reduzida no t?lamo/hipot?lamo (dia 3). A an?lise por imunoistoqu?mica revelou express?o aumentada do NOPr no g?nglio da raiz dorsal (dia 4). O UFP-101 causou uma diminui??o no metabolismo de [18F]-FDG no giro do cingulado, no col?culo superior, no mesenc?falo esquerdo, no col?culo inferior esquerdo e no col?culo inferior direito de camundongos tratados com reserpina. Al?m disso, o UFP-101 preveniu as altera??es induzidas pela reserpina na distribui??o do tamanho das fibras musculares, de acordo com a avalia??o dos cortes histol?gicos do masseter e do gastrocn?mio. Tanto a indu??o da fibromialgia pela reserpina, quanto o tratamento cr?nico com UFP-101, n?o alteraram a ?rea mitocondrial. Em resumos, os dados do presente estudo indicam que o bloqueio farmacol?gico do NOPr reduziu os sintomas de dor, fadiga e adinamia, recuperando tamb?m os padr?es de ativa??o cerebral e as altera??es musculares esquel?ticas no modelo de fibromialgia experimental induzido pela reserpina. Al?m disso, express?o da ppN/OFQ e do NOPr foi alterada pela indu??o de fibromialgia, tanto em s?tios centrais, quanto perif?ricos, refor?ando a relev?ncia do sistema N/OFQ-NOPr na patofisiologia da fibromialgia. Fibromyalgia is characterized by widespread pain, being accompanied by functional and affective disorders. This study evaluated the implication of nociceptin/orphanin FQ peptide receptor (NOPr) in a mouse model of fibromyalgia. The local Animal Ethics Committee approved the experimental protocols (15/00487). Fibromyalgia was induced in female CF-1 mice (20-24 g, 4 week-old) by reserpine administration (0.25 mg/kg; subcutaneous route), once a day, during 3 consecutive days. Control groups received vehicle. On the fourth day, mice were acutely treated with the selective NOP agonist nociceptin (N/OFQ), or with the selective peptide antagonist UFP-101, given by intraperitoneal (i.p., 0.3 5 nmol/kg), intracerebroventricular (i.c.v., 0.3-1 nmol/site), or intrathecal (i.t., 0.3-5 nmol/site) routes, 30 min before the experimental sessions. In a separate set of experiments, the animals were treated with the peptide UFP-101 (1 nmol/kg) or the non-peptide SB-612111 (6.6 ?mol/kg) antagonists, given by intraperitoneal route, during three consecutive days, 30 min after daily reserpine injection. At the 4th day, mice also received the antagonist, dosed 30 min before evaluations. The animals were subjected to Von Frey, hot-plate, forced swimming, elevated plus-maze, rotarod and grasping tests. Pre-pro-nociceptin (ppN/OFQ) and NOPr expression was determined by RT-qPCR and immunohistochemistry. The [18F]-FDG microPET imaging was used to assess the brain activation patterns in reserpine-treated mice. The fiber size distribution of masseter and gastrocnemius muscles was evaluated by histological analysis. The mitochondria area and density in the skeletal muscle were analysed by transmission electron microscopy (TEM). In the acute protocols of treatment, the i.t. or i.p. administration of N/OFQ (1 nmol/site or 1 nmol/kg, respectively) significantly reduced the mechanical allodynia. However, i.p. treatment with N/OFQ at the dose of 5 nmol/kg had an opposite effect, leading to hypernociception. Concerning the UFP-101 effects, this peptide antagonist given i.c.v. (1 nmol/site), i.t. (3 and 5 nmol/site) or i.p. (1, 3 and 5 nmol/kg) significantly reduced the mechanical hypersensitivity in mice treated with reserpine. The acute treatment with N/OFQ or UFP-101 did not significantly alter the thermal hypersensivity, when given by i.c.v. or i.p. routes. The i.t. administration of N/OFQ (3 nmol/site) and UFP-101 (5 nmol/site) had a significant inhibitory effect on the thermal nociception. The immobility time was significantly inhibited by N/OFQ, given by i.c.v (1 nmol/site) or i.t. (3 nmol/site) routes. N/OFQ and UFP-101 did not modify any anxiety-related parameter. The chronic treatment with UFP-101 and SB-612111 reduced the mechanical allodynia (37 ? 8% and 43 ? 15.2%) and the thermal hypernociception (32.2 ? 5% and 45 ? 17.5%), besides improving the motor coordination in the rotational apparatus (7 and 2-fold increase in permanence time) and the grasping strength (15 ? 16% and 9 ? 5.5%), respectively. None of the antagonists altered the parameters of anxiety or depression. Reserpine-induced fibromyalgia was associated with an increase in ppN/OFQ mRNA expression in the lumbar spinal cord (day 3) and masseter (days 1 and 2), whereas NOPr mRNA expression was increased in the masseter muscle (day 1). Alternatively, NOPr m RNA expression was reduced in the thalamus/hypothalamus (day 3). The immunohistochemistry analysis revealed an increased expression of NOPr in the dorsal root ganglion (DRG; on day 4). UFP-101 led to a decrease in the [18F]-FDG metabolism in cingulate gyrus, superior colliculus, left midbrain, left inferior colliculus and right inferior colliculus of reserpine treated mice. Additionally, UFP-101 prevented reserpine-induced changes in fiber size distribution, according to the assessment of masseter and gastrocnemius histological sections. TEM analysis revealed that either the induction of fibromyalgia by reserpine, or the chronic treatment with UFP-101, did not alter the mitochondrial area or density. The expression of nociceptin and NOPr was altered in the mouse model of fibromyalgia induced by reserpine. Remarkably, UFP-101 improved the symptoms of pain, fatigue and adinamia, also recovering the brain activation patterns and the muscle fiber changes in this experimental paradigm. Our data shed new lights on the mechanisms underlying the fibromyalgia pathogenesis, supporting a role for NOPr in this syndrome. Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior - CAPES

INTRODUÇÃO: A agressividade é um comportamento comum a diversas espécies animais, incluindo humanos. Entretanto, a violência e a impulsividade, associadas à agressividade, são um problema social e podem ser consideradas patológicas, pois estão presentes em vários transtornos psiquiátricos. Diversas áreas encefálicas estão associadas à expressão do comportamento agressivo, como a amígdala, hipotálamo e córtex pré-frontal. Vários sistemas de neurotransmissão estão mediando o comportamento agressivo, dentre eles: serotonina, dopamina, noradrenalina e GABA. De maneira geral, os alvos terapêuticos disponíveis para controle da agressividade modulam a função dos sistemas de neurotransmissores acima. A nociceptina/orfanina FQ (N/OFQ) é um heptadecapeptídeo que atua como ligante do receptor NOP. Evidências clínicas e pré-clínicas mostram o envolvimento do sistema N/OFQ – receptor NOP com transtornos psiquiátricos, incluindo aqueles nos quais a agressividade está associada. OBJETIVO: Este trabalho investigou o efeito de fármacos clássicos e ligantes do receptor NOP no comportamento agressivo de camundongos machos, por meio do teste do residente-intruso. MÉTODOS: Foram utilizados camundongos Swiss machos. Valproato 300 mg/kg, Lítio 50 mg/kg, Carbamazepina 20 mg/kg e Diazepam 1 mg/kg foram os fármacos clássicos utilizados nesse estudo. Dentre os ligantes NOP utilizados destacam-se: Ro 65-6570 (0,01 – 1 mg/kg), agonista pleno, AT-090 (0,01-0,1 mg/kg), agonista parcial, SB-612111 (1- 10 mg/kg), antagonista NOP. Para o teste do residente-intruso, camundongos machos foram isolados por 7 dias (residentes). Nos 8° e 11° dia, foram realizadas sessões de avaliação da agressividade, por meio da inserção de um camundongo intruso na caixa do residente por 10 min. No dia 8, a agressividade basal foi avaliada sem qualquer tratamento prévio; no dia 11, o mesmo camundongo residente foi novamente avaliado, após ter recebido o tratamento relativo ao seu grupo experimental. O campo aberto foi utilizado para avaliar o efeito dos fármacos na atividade locomotora. RESULTADOS: Valproato, Lítio, Carbamazepina reduziram o comportamento agressivo no teste do residente-intruso, enquanto que o tratamento com Diazepam não afetou a agressividade dos residentes. A administração de Ro 65-6570 (em todas as doses testadas) e AT-090 (na dose mais alta), aumentou o comportamento agressivo. Já o agonista parcial, AT-090, nas menores doses, reduziu discretamente a agressividade dos residentes. O tratamento com SB-612111 não modificou o comportamento agressivo dos animais. Nenhum dos tratamentos alterou a atividade locomotora dos animais. CONCLUSÃO: Os fármacos clássicos utilizados na clínica para tratamento de transtornos psiquiátricos, os quais incluem sintomas de agressividade, foram eficazes em controlar a agressividade nos camunodngos residentes. Por outro lado, a ativação do receptor NOP tende a aumentar o comportamento agressivo, enquanto que o bloqueio deste sinal não foi modifica este comportamento. Em última análise, com estes dados sugere-se que os agonistas NOP poderiam promover como efeito adverso aumento da agressividade. INTRODUCTION: Several species including humans display aggressive behavior. However, violence and impulsivity related to aggressiveness represent a social problem. Indeed, aggressive behavior can be considered symptoms of many psychiatric disorders. Some of the brain areas involved in aggression include amygdala, hypothalamus, and prefrontal cortex. Aggressiveness is modulated by different neurotransmitters, such as serotonin, dopamine, noradrenaline and GABA. These systems represent the therapeutic targets available to treat aggressiveness. The nociceptin/orphanin FQ (N/OFQ) is a heptadecapeptide acting as endogenous ligand of NOP receptor. Clinical and preclinical findings suggest the involvement of N/OFQ – NOP receptor system with psychiatric disorders, including those related to aggressiveness. AIM: This study investigated the effects of standard drugs as well as NOP receptor ligands on aggressiveness in mice submitted to the resident-intruder test. METHODS: Male Swiss mice were used to develop this study. Valproate 300 mg/kg, Lithium 50 mg/kg, Carbamazepine 20 mg/kg, and Diazepam 1 mg/kg were used as standard drugs. The NOP ligands Ro 65-6570 (0.01 – 1 mg/kg), full agonist, AT-090 (0,01 – 0,1 mg/kg), partial agonist, and SB-612111 (1 – 10 mg/kg), antagonist, were used. In the resident-intruder test, male mice were housed individually for 7 days (residents) before the experiment. The aggressiveness of each resident mouse was tested twice, at 8th and 11th days, by inserting an intruder mouse in the resident cage for 10 min. Day 8 of experiment, the basal aggressiveness of resident mice was recorded without pharmacological treatment; Day 11 of experiment, the same mouse was re-tested after being treated. The open field was used to evaluated the spontaneous locomotor activity . RESULTS: Valproate, Lithium, and Carbamazepine reduced the aggressive behavior of resident mice, while Diazepam did not affect the agressiveness. Ro 65-6570 (at all doses) and AT-090 (at the highest dose), increased aggressiveness. The partial agonist, AT-090, at lowest doses, slightly reduced aggressive behavior. The treatment with SB-61211 did not modified the aggressive behavior of mice. None of the treatments affected the locomotor activity. CONCLUSION: Standard drugs used in therapy for psychiatric disorders were effective on aggressiveness control in the resident mice. In contrast, the activation of NOP receptor tends to increase the aggressive behavior, while the blockade of this signal did not modify this behavior. Ultimately, these data suggest that NOP agonists could increase aggressive behavior as an adverse event.

International audience; To investigate whether the endogenous neuropeptide nociceptin/orphanin FQ (N/OFQ) contributes to the death of dopamine neurons in Parkinson's disease, we undertook a genetic and a pharmacological approach using NOP receptor knockout (NOP(-/-)) mice, and the selective and potent small molecule NOP receptor antagonist (-)-cis-1-methyl-7-[[4-(2,6-dichlorophenyl)piperidin-1-yl]methyl]-6,7,8,9-tetrahydro-5H-benzocyclohepten-5-ol (SB-612111). Stereological unbiased methods were used to estimate the total number of dopamine neurons in the substantia nigra of i) NOP(-/-) mice acutely treated with the parkinsonian neurotoxin 1-methyl-4-phenyl-1,2,5,6-tetrahydropyridine (MPTP), ii) naïve mice subacutely treated with MPTP, alone or in combination with SB-612111, iii) rats injected with a recombinant adeno-associated viral (AAV) vector overexpressing human mutant p.A53T α-synuclein, treated with vehicle or SB-612111. NOP(-/-) mice showed a 50% greater amount of nigral dopamine neurons spared in response to acute MPTP compared to controls, which was associated with a milder motor impairment. SB-612111, given 4days after MPTP treatment to mimic the clinical condition, prevented the loss of nigral dopamine neurons and striatal dopaminergic terminals caused by subacute MPTP. SB-612111, administered a week after the AAV injections in a clinically-driven protocol, also increased by 50% both the number of spared nigral dopamine neurons and striatal dopamine terminals, and prevented accompanying motor deficits induced by α-synuclein. We conclude that endogenous N/OFQ contributes to dopamine neuron loss in pathogenic and etiologic models of Parkinson's disease through NOP receptor-mediated mechanisms. NOP receptor antagonists might prove effective as disease-modifying agents in Parkinson's disease, through the rescue of degenerating nigral dopamine neurons and/or the protection of the healthy ones.