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1.
Biomolecules ; 11(5)2021 04 24.
Artigo em Inglês | MEDLINE | ID: mdl-33923147

RESUMO

The blood-brain barrier (BBB) plays an important protective role in the central nervous system and maintains its homeostasis. It regulates transport into brain tissue and protects neurons against the toxic effects of substances circulating in the blood. However, in the case of neurological diseases or primary brain tumors, i.e., gliomas, the higher permeability of the blood-derived substances in the brain tissue is necessary. Currently applied methods of treatment for the primary brain neoplasms include surgical removal of the tumor, radiation therapy, and chemotherapy. Despite the abovementioned treatment methods, the prognosis of primary brain tumors remains bad. Moreover, chemotherapy options seem to be limited due to low drug penetration into the cancerous tissue. Modulation of the blood-brain barrier permeability may contribute to an increase in the concentration of the drug in the CNS and thus increase the effectiveness of therapy. Interestingly, endothelial cells in cerebral vessels are characterized by the presence of adenosine 2A receptors (A2AR). It has been shown that substances affecting these receptors regulate the permeability of the BBB. The mechanism of increasing the BBB permeability by A2AR agonists is the actin-cytoskeletal reorganization and acting on the tight junctions. In this case, the A2AR seems to be a promising therapy target. This article aims to assess the possibility of increasing the BBB permeability through A2AR agonists to increase the effectiveness of chemotherapy and to improve the results of cancer therapy.


Assuntos
Barreira Hematoencefálica/metabolismo , Neoplasias/metabolismo , Receptores A2 de Adenosina/metabolismo , Animais , Transporte Biológico , Barreira Hematoencefálica/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Células Endoteliais/metabolismo , Humanos , Neoplasias/terapia , Neurônios/metabolismo , Permeabilidade , Receptores A2 de Adenosina/fisiologia , Transdução de Sinais/efeitos dos fármacos , Junções Íntimas/efeitos dos fármacos , Junções Íntimas/metabolismo
2.
Molecules ; 26(4)2021 Feb 23.
Artigo em Inglês | MEDLINE | ID: mdl-33672225

RESUMO

Alzheimer's, Parkinson's, and multiple sclerosis are neurodegenerative diseases related by neuronal degeneration and death in specific areas of the central nervous system. These pathologies are associated with neuroinflammation, which is involved in disease progression, and halting this process represents a potential therapeutic strategy. Evidence suggests that microglia function is regulated by A1 and A2A adenosine receptors (AR), which are considered as neuroprotective and neurodegenerative receptors, respectively. The manuscript's aim is to elucidate the role of these receptors in neuroinflammation modulation through potent and selective A1AR agonists (N6-cyclopentyl-2'- or 3'-deoxyadenosine substituted or unsubstituted in 2 position) and A2AAR antagonists (9-ethyl-adenine substituted in 8 and/or in 2 position), synthesized in house, using N13 microglial cells. In addition, the combined therapy of A1AR agonists and A2AAR antagonists to modulate neuroinflammation was evaluated. Results showed that A1AR agonists were able, to varying degrees, to prevent the inflammatory effect induced by cytokine cocktail (tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, and interferon (IFN)-γ), while A2AAR antagonists showed a good ability to counteract neuroinflammation. Moreover, the effect achieved by combining the two most effective compounds (1 and 6) in doses previously found to be non-effective was greater than the treatment effect of each of the two compounds used separately at maximal dose.


Assuntos
Agonistas do Receptor A1 de Adenosina/farmacologia , Antagonistas do Receptor A2 de Adenosina/farmacologia , Inflamação/tratamento farmacológico , Receptor A1 de Adenosina/metabolismo , Receptores A2 de Adenosina/metabolismo , Animais , Células Cultivadas , Inflamação/metabolismo , Camundongos
3.
J Chem Theory Comput ; 16(11): 7207-7218, 2020 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-33112150

RESUMO

Alchemical free energy simulations have long been utilized to predict free energy changes for binding affinity and solubility of small molecules. However, while the theoretical foundation of these methods is well established, seamlessly handling many of the practical aspects regarding the preparation of the different thermodynamic end states of complex molecular systems and the numerous processing scripts often remains a burden for successful applications. In this work, we present CHARMM-GUI Free Energy Calculator (http://www.charmm-gui.org/input/fec) that provides various alchemical free energy perturbation molecular dynamics (FEP/MD) systems with input and post-processing scripts for NAMD and GENESIS. Four submodules are available: Absolute Ligand Binder (for absolute ligand binding FEP/MD), Relative Ligand Binder (for relative ligand binding FEP/MD), Absolute Ligand Solvator (for absolute ligand solvation FEP/MD), and Relative Ligand Solvator (for relative ligand solvation FEP/MD). Each module is designed to build multiple systems of a set of selected ligands at once for high-throughput FEP/MD simulations. The capability of Free Energy Calculator is illustrated by absolute and relative solvation FEP/MD of a set of ligands and absolute and relative binding FEP/MD of a set of ligands for T4-lysozyme in solution and the adenosine A2A receptor in a membrane. The calculated free energy values are overall consistent with the experimental and published free energy results (within ∼1 kcal/mol). We hope that Free Energy Calculator is useful to carry out high-throughput FEP/MD simulations in the field of biomolecular sciences and drug discovery.


Assuntos
Modelos Moleculares , Solventes/química , Descoberta de Drogas , Ligantes , Receptores A2 de Adenosina/química , Receptores A2 de Adenosina/metabolismo , Termodinâmica
4.
Int J Dev Neurosci ; 80(7): 636-647, 2020 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-32798310

RESUMO

BACKGROUND: This study assessed the effects of an acute stress model upon the long-term hyperalgesia induced by repeated morphine administration in neonatal rats. We also evaluated neurotrophins and cytokines levels; expressions of adenosine and acetylcholine receptors, and acetylcholinesterase enzyme at the spinal cord. MATERIAL AND METHODS: Male Wistar rats were subjected to morphine or saline administration from P8 to P14. Thermal hyperalgesia and mechanical hyperesthesia were assessed using the hot plate (HP) and von Frey (vF) tests, respectively, at postnatal day P30 and P60. After baseline measurements, rats were subjected to a single exercise session, as an acute stress model, at P30 or P60. We measured the levels of BDNF and NGF, interleukin-6, and IL-10 in the cerebral cortex and the brainstem; and the expression levels of adenosine and muscarinic receptors, as well as acetylcholinesterase (AChE) enzyme at the spinal cord. RESULTS: A stress exercise session was not able to revert the morphine-induced hyperalgesia. The morphine and exercise association in rats induced a decrease in the neurotrophins brainstem levels, and A1 , A2A , A2B receptors expression in the spinal cord, and an increase in the IL-6 cortical levels. The exercise reduced M2 receptors expression in the spinal cord of naive rats, while morphine prevented this effect. CONCLUSIONS: Single session of exercise does not revert hyperalgesia induced by morphine in rats; however, morphine plus exercise modulate neurotrophins, IL-6 central levels, and expression of adenosine receptors.


Assuntos
Hiperalgesia/metabolismo , Fatores de Crescimento Neural/metabolismo , Condicionamento Físico Animal/fisiologia , Receptor A1 de Adenosina/metabolismo , Receptores A2 de Adenosina/metabolismo , Acetilcolinesterase/metabolismo , Animais , Citocinas/metabolismo , Hiperalgesia/induzido quimicamente , Masculino , Morfina/efeitos adversos , Ratos , Ratos Wistar , Receptores Colinérgicos/metabolismo
5.
Nitric Oxide ; 104-105: 20-28, 2020 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-32828841

RESUMO

The adenosine (Ado) system may participate in regulation of kidney function in diabetes mellitus (DM), therefore we explored its role and interrelation with NO in the control of renal circulation and excretion in normoglycemic (NG) and streptozotocin-diabetic (DM) rats. Effects of theophylline (Theo), a non-selective Ado receptor antagonist, were examined in anaesthetized NG or in streptozotocin induced diabetic (DM) rats, untreated or after blockade of NO synthesis with l-NAME. We measured arterial blood pressure (MABP), whole kidney blood flow and renal regional flows: cortical and outer- and inner-medullary (IMBF), determined as laser-Doppler fluxes. Renal excretion of water, total solutes and sodium and in situ renal tissue NO signal (selective electrodes) were also determined. Theo experiments disclosed minor baseline vasoconstrictor and vasodilator tone in the kidney of NG and DM rats, respectively. NO blockade increased baseline MABP and decreased renal haemodynamics, similarly in NG and DM rats, indicating comparable vasodilator influence of NO in the two groups. Unexpectedly, in all rats with intact NO synthesis, Ado receptor blockade increased kidney tissue NO. In NO-deficient NG and DM rats, Ado receptor blockade induced comparable renal vasodilatation, suggesting similar vasoconstrictor influence of the Ado system. However, DM rats showed an unexplained association of decreased MABP and IMBF and increased NO signal. Higher baseline renal excretion in DM rats indicated inhibition of renal tubular reabsorption due to the prevalence of natriuretic A2 over antinatriuretic A1 receptors. In conclusion, the experiments provided new insights in functional interrelation of adenosine and NO in normoglycaemia and streptozotocin-diabetes.


Assuntos
Adenosina/metabolismo , Diabetes Mellitus Experimental/metabolismo , Rim/metabolismo , Óxido Nítrico/metabolismo , Circulação Renal/efeitos dos fármacos , Eliminação Renal/efeitos dos fármacos , Animais , Pressão Arterial/efeitos dos fármacos , Diabetes Mellitus Experimental/induzido quimicamente , Inibidores Enzimáticos/farmacologia , Masculino , NG-Nitroarginina Metil Éster/farmacologia , Óxido Nítrico/antagonistas & inibidores , Óxido Nítrico Sintase/antagonistas & inibidores , Antagonistas de Receptores Purinérgicos P1/farmacologia , Ratos Sprague-Dawley , Receptor A1 de Adenosina/metabolismo , Receptores A2 de Adenosina/metabolismo , Estreptozocina , Teofilina/farmacologia
6.
Cells ; 9(7)2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32645849

RESUMO

Adenosine (ARs) and metabotropic glutamate receptors (mGluRs) are G-protein coupled receptors (GPCRs) that are modulated in the brain of SAMP8 mice, an animal model of Alzheimer's disease (AD). In the present work, it is shown the presence of ARs and mGluRs in blood serum and derived exosomes from SAMP8 mice as well as its possible modulation by aging and resveratrol (RSV) consumption. In blood serum, adenosine A1 and A2A receptors remained unaltered from 5 to 7 months of age. However, an age-related decrease in adenosine level was observed, while 5'-Nucleotidase activity was not modulated. Regarding the glutamatergic system, it was observed a decrease in mGluR5 density and glutamate levels in older mice. In addition, dietary RSV supplementation caused an age-dependent modulation in both adenosinergic and glutamatergic systems. These GPCRs were also found in blood serum-derived exosomes, which might suggest that these receptors could be released into circulation via exosomes. Interestingly, changes elicited by age and RSV supplementation on mGluR5 density, and adenosine and glutamate levels were similar to that detected in whole-brain. Therefore, we might suggest that the quantification of these receptors, and their corresponding endogenous ligands, in blood serum could have predictive value for early diagnosis in combination with other distinctive hallmarks of AD.


Assuntos
Adenosina/sangue , Adenosina/metabolismo , Exossomos/metabolismo , Receptores de Glutamato Metabotrópico/sangue , Resveratrol/uso terapêutico , Envelhecimento/fisiologia , Doença de Alzheimer/sangue , Doença de Alzheimer/tratamento farmacológico , Animais , Western Blotting , Cromatografia Líquida de Alta Pressão , Masculino , Camundongos , Receptor A1 de Adenosina/sangue , Receptor A1 de Adenosina/metabolismo , Receptores A2 de Adenosina/sangue , Receptores A2 de Adenosina/metabolismo
7.
Cells ; 9(5)2020 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-32443448

RESUMO

Glutamate cytotoxicity is implicated in neuronal death in different neurological disorders including stroke, traumatic brain injury, and neurodegenerative diseases. Adenosine is a nucleoside that plays an important role in modulating neuronal activity and its receptors have been identified as promising therapeutic targets for glutamate cytotoxicity. The purpose of this study is to elucidate the role of adenosine and its receptors on glutamate-induced injury in PC12 cells and to verify the protective effect of the novel A1 adenosine receptor positive allosteric modulator, TRR469. Flow cytometry experiments to detect apoptosis revealed that adenosine has a dual role in glutamate cytotoxicity, with A2A and A2B adenosine receptor (AR) activation exacerbating and A1 AR activation improving glutamate-induced cell injury. The overall effect of endogenous adenosine in PC12 cells resulted in a facilitating action on glutamate cytotoxicity, as demonstrated by the use of adenosine deaminase and selective antagonists. However, enhancing the action of endogenous adenosine on A1ARs by TRR469 completely abrogated glutamate-mediated cell death, caspase 3/7 activation, ROS production, and mitochondrial membrane potential loss. Our results indicate a novel potential therapeutic strategy against glutamate cytotoxicity based on the positive allosteric modulation of A1ARs.


Assuntos
Adenosina/farmacologia , Ácido Glutâmico/toxicidade , Neuroproteção/efeitos dos fármacos , Receptor A1 de Adenosina/metabolismo , Antagonistas do Receptor A1 de Adenosina/farmacologia , Antagonistas do Receptor A2 de Adenosina/farmacologia , Adenosina-5'-(N-etilcarboxamida)/farmacologia , Regulação Alostérica/efeitos dos fármacos , Animais , Carbazóis/farmacologia , Caspases/metabolismo , Morte Celular/efeitos dos fármacos , Colforsina/farmacologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Células PC12 , Piperazinas/farmacologia , Pirróis/farmacologia , Quinazolinas/farmacologia , Ratos , Espécies Reativas de Oxigênio/metabolismo , Receptores A2 de Adenosina/metabolismo , Tiofenos/farmacologia , Triazóis/farmacologia
8.
Drug Res (Stuttg) ; 70(6): 243-256, 2020 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-32349128

RESUMO

Adenosine A1 and/or A2A receptor antagonists hold promise for the potential treatment of neurological conditions, such as Parkinson's disease. Herein, a total of seventeen benzocycloalkanone derivatives were synthesised and evaluated for affinity towards adenosine receptors (A1 and A2A AR). The obtained results allowed for the conclusion that affinity and/or selectivity of the 2-benzylidene-1-indanone and -tetralone derivatives toward A1 and/or A2A ARs may be modulated by the nature of the substituents (either -OH, -OCH3 or morpholine) attached at position C4 of the 1-indanone core and C5 of the 1-tetralone core as well as the meta (C3') and/or para (C4') position(s) on ring B. Several compounds (2A: -B: , 3B: -C: and 4A: -B: ) possessed affinity for the A1 and/or A2A AR below 10 µM. Additionally, compounds 2A: , 3B: and 4A: were A1 AR antagonists. These results, once again, confirmed the importance of C4 methoxy-group substitution on ring A in combination with meta (C3') and/or para (C4') hydroxyl-group substitution ring B of the 2-benzylidene-1-indanone scaffold leading to drug-like compounds 1H: and 1J: with affinity in the nanomolar-range.


Assuntos
Antagonistas do Receptor A1 de Adenosina/farmacologia , Antagonistas do Receptor A2 de Adenosina/farmacologia , Técnicas de Química Sintética/métodos , Doença de Parkinson/tratamento farmacológico , Antagonistas do Receptor A1 de Adenosina/síntese química , Antagonistas do Receptor A2 de Adenosina/síntese química , Antagonistas do Receptor A2 de Adenosina/uso terapêutico , Chalconas/química , Química Farmacêutica/métodos , Simulação por Computador , Humanos , Estrutura Molecular , Receptor A1 de Adenosina/metabolismo , Receptores A2 de Adenosina/metabolismo , Relação Estrutura-Atividade
9.
Molecules ; 25(5)2020 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-32164183

RESUMO

Adenosine Receptor Type 2A (A2AAR) plays a role in important processes, such as anti-inflammatory ones. In this way, the present work aimed to search for compounds by pharmacophore-based virtual screening. The pharmacokinetic/toxicological profiles of the compounds, as well as a robust QSAR, predicted the binding modes via molecular docking. Finally, we used molecular dynamics to investigate the stability of interactions from ligand-A2AAR. For the search for A2AAR agonists, the UK-432097 and a set of 20 compounds available in the BindingDB database were studied. These compounds were used to generate pharmacophore models. Molecular properties were used for construction of the QSAR model by multiple linear regression for the prediction of biological activity. The best pharmacophore model was used by searching for commercial compounds in databases and the resulting compounds from the pharmacophore-based virtual screening were applied to the QSAR. Two compounds had promising activity due to their satisfactory pharmacokinetic/toxicological profiles and predictions via QSAR (Diverset 10002403 pEC50 = 7.54407; ZINC04257548 pEC50 = 7.38310). Moreover, they had satisfactory docking and molecular dynamics results compared to those obtained for Regadenoson (Lexiscan®), used as the positive control. These compounds can be used in biological assays (in vitro and in vivo) in order to confirm the potential activity agonist to A2AAR.


Assuntos
Receptores A2 de Adenosina/metabolismo , Adenosina/análogos & derivados , Adenosina/farmacologia , Agonistas do Receptor A2 de Adenosina/farmacologia , Humanos , Ligantes , Simulação de Acoplamento Molecular/métodos , Simulação de Dinâmica Molecular , Relação Quantitativa Estrutura-Atividade
10.
Clin Cancer Res ; 26(9): 2176-2187, 2020 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-31953314

RESUMO

PURPOSE: There are several agents in early clinical trials targeting components of the adenosine pathway including A2AR and CD73. The identification of cancers with a significant adenosine drive is critical to understand the potential for these molecules. However, it is challenging to measure tumor adenosine levels at scale, thus novel, clinically tractable biomarkers are needed. EXPERIMENTAL DESIGN: We generated a gene expression signature for the adenosine signaling using regulatory networks derived from the literature and validated this in patients. We applied the signature to large cohorts of disease from The Cancer Genome Atlas (TCGA) and cohorts of immune checkpoint inhibitor-treated patients. RESULTS: The signature captures baseline adenosine levels in vivo (r 2 = 0.92, P = 0.018), is reduced after small-molecule inhibition of A2AR in mice (r 2 = -0.62, P = 0.001) and humans (reduction in 5 of 7 patients, 70%), and is abrogated after A2AR knockout. Analysis of TCGA confirms a negative association between adenosine and overall survival (OS, HR = 0.6, P < 2.2e-16) as well as progression-free survival (PFS, HR = 0.77, P = 0.0000006). Further, adenosine signaling is associated with reduced OS (HR = 0.47, P < 2.2e-16) and PFS (HR = 0.65, P = 0.0000002) in CD8+ T-cell-infiltrated tumors. Mutation of TGFß superfamily members is associated with enhanced adenosine signaling and worse OS (HR = 0.43, P < 2.2e-16). Finally, adenosine signaling is associated with reduced efficacy of anti-PD1 therapy in published cohorts (HR = 0.29, P = 0.00012). CONCLUSIONS: These data support the adenosine pathway as a mediator of a successful antitumor immune response, demonstrate the prognostic potential of the signature for immunotherapy, and inform patient selection strategies for adenosine pathway modulators currently in development.


Assuntos
Antagonistas do Receptor A2 de Adenosina/uso terapêutico , Adenosina/metabolismo , Imunoterapia/métodos , Neoplasias/terapia , Animais , Biomarcadores Tumorais/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular Tumoral , Bases de Dados Genéticas , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patologia , Prognóstico , Distribuição Aleatória , Receptores A2 de Adenosina/metabolismo , Transdução de Sinais/genética , Taxa de Sobrevida , Transcriptoma
11.
Chem Phys Lipids ; 227: 104852, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31866438

RESUMO

G protein-coupled receptors (GPCRs) are transmembrane receptors that mediate a large number of cellular responses. The organization of GPCRs into dimers and higher-order oligomers is known to allow a larger repertoire of downstream signaling events. In this context, a crosstalk between the adenosine and dopamine receptors has been reported, indicating the presence of heterodimers that are functionally relevant. In this paper, we explored the effect of membrane cholesterol on the adenosine2A (A2A) and dopamine D3 (D3) receptors using coarse-grain molecular dynamics simulations. We analyzed cholesterol interaction sites on the A2A receptor and were able to reproduce the sites indicated by crystallography and previous atomistic simulations. We predict novel cholesterol interaction sites on the D3 receptor that could be important in the reported cholesterol sensitivity in receptor function. Further, we analyzed the formation of heterodimers between the two receptors. Our results suggest that membrane cholesterol modulates the relative population of several co-existing heterodimer conformations. Both direct receptor-cholesterol interaction and indirect membrane effects contribute toward the modulation of heterodimer conformations. These results constitute one of the first examples of modulation of GPCR hetero-dimerization by membrane cholesterol, and could prove to be useful in designing better therapeutic strategies.


Assuntos
Colesterol/metabolismo , Receptores A2 de Adenosina/metabolismo , Receptores de Dopamina D3/metabolismo , Sítios de Ligação , Colesterol/química , Dimerização , Humanos , Bicamadas Lipídicas/química , Bicamadas Lipídicas/metabolismo , Simulação de Dinâmica Molecular , Ligação Proteica , Receptores A2 de Adenosina/química , Receptores de Dopamina D3/química
13.
J Neural Transm (Vienna) ; 126(12): 1577-1588, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31522257

RESUMO

Ketamine (KET), an anesthetic, analgesic, and a sedative N-methyl-D-aspartate (NMDA) receptor antagonist agent, exposure during neonatal period may lead to learning impairment, behavioral abnormalities, and cognitive decline in the later years of life. In recent studies, it has been reported that sedative-acting α2 agonist dexmedetomidine (DEX), which is commonly used in clinical practice with KET, has neuroprotective effects and prevents the undesirable effects of anesthesia. To elucidate the underlying mechanisms of these actions, we investigated the interaction between NMDA receptors α2 adrenoceptor and adulthood behaviors in neonatally KET and/or DEX administrated mice. Balb/c male mice were administrated with saline, KET (75 mg/kg), DEX (10 µg/kg), or KET + DEX (75 mg/kg + 10 µg/kg) on postnatal day 7. During adulthood (8-10 weeks old) mice were subjected to elevated plus maze, open field, and Morris water maze tests. After behavioral tests, hippocampus samples were extracted for mRNA expression studies of NMDAR subunits (GluN1, GluN2A, and GluN2B) and α2 adrenoceptor subunits (α2A, α2B, and α2C) by real-time PCR. Ketamine increased horizontal and vertical locomotor activity (p < 0.01) and impaired spatial learning-memory (p < 0.05). DEX increased anxiety-like behavior (p < 0.01), but did not affect spatial learning-memory and locomotor activity. KET + DEX impaired spatial learning-memory (p < 0.01), increased horizontal locomotor activity (p < 0.01), and anxiety-like behavior (p < 0.05). Our study implies that DEX cannot prevent the adverse effects of KET, on spatial learning-memory, and locomotor activity. In addition to this, it can be thought that during brain development, there is an interaction between NMDAR and α2 adrenoceptor systems.


Assuntos
Anestésicos Dissociativos/farmacologia , Comportamento Animal/efeitos dos fármacos , Dexmedetomidina/farmacologia , Hipnóticos e Sedativos/farmacologia , Ketamina/farmacologia , Animais , Animais Recém-Nascidos , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Masculino , Camundongos Endogâmicos BALB C , Receptores A2 de Adenosina/efeitos dos fármacos , Receptores A2 de Adenosina/metabolismo , Receptores de N-Metil-D-Aspartato/efeitos dos fármacos , Receptores de N-Metil-D-Aspartato/metabolismo
14.
Neurochem Int ; 131: 104550, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31563462

RESUMO

Caffeine is the most consumed psychostimulant drug in the world, acting as a non-selective antagonist of adenosine receptors A1R and A2AR, which are widely expressed in retinal layers. We have previously shown that caffeine, when administered acutely, acts on A1R to potentiate the NMDA receptor-induced GABA release. Now we asked if long-term caffeine exposure also modifies GABA uptake in the avian retina and which mechanisms are involved in this process. Chicken embryos aged E11 were injected with a single dose of caffeine (30 mg/kg) in the air chamber. Retinas were dissected on E15 for ex vivo neurochemical assays. Our results showed that [3H]-GABA uptake was dependent on Na+ and blocked at 4 °C or by NO-711 and caffeine. This decrease was observed after 60 min of [3H]-GABA uptake assay at E15, which is accompanied by an increase in [3H]-GABA release. Caffeine increased the protein levels of A1R without altering ADORA1 mRNA and was devoid of effects on A2AR density or ADORA2A mRNA levels. The decrease of GABA uptake promoted by caffeine was reverted by A1R activation with N6-cyclohexyl adenosine (CHA) but not by A2AR activation with CGS 21680. Caffeine exposure increased cAMP levels and GAT-1 protein levels, which was evenly expressed between E11-E15. As expected, we observed an increase of GABA containing amacrine cells and processes in the IPL, also, cAMP pathway blockage by H-89 decreased caffeine mediated [3H]-GABA uptake. Our data support the idea that chronic injection of caffeine alters GABA transport via A1R during retinal development and that the cAMP/PKA pathway plays an important role in the regulation of GAT-1 function.


Assuntos
Bloqueadores do Receptor Tipo 1 de Angiotensina II/farmacologia , Antagonistas de Receptores de Angiotensina/farmacologia , Cafeína/farmacologia , Estimulantes do Sistema Nervoso Central/farmacologia , AMP Cíclico/fisiologia , Ácido gama-Aminobutírico/metabolismo , Adenosina/análogos & derivados , Adenosina/farmacologia , Células Amácrinas/efeitos dos fármacos , Células Amácrinas/metabolismo , Animais , Cafeína/antagonistas & inibidores , Embrião de Galinha , AMP Cíclico/metabolismo , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Proteínas da Membrana Plasmática de Transporte de GABA/metabolismo , Fenetilaminas/farmacologia , Receptor A1 de Adenosina/efeitos dos fármacos , Receptor A1 de Adenosina/metabolismo , Receptor Tipo 1 de Angiotensina/efeitos dos fármacos , Receptores A2 de Adenosina/efeitos dos fármacos , Receptores A2 de Adenosina/metabolismo , Retina/efeitos dos fármacos , Retina/embriologia , Retina/crescimento & desenvolvimento , Transdução de Sinais/efeitos dos fármacos
15.
Drugs R D ; 19(4): 319-328, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31452117

RESUMO

BACKGROUND: Pulmonary arterial hypertension is a hemodynamic disorder. Signs and symptoms are generally difficult to recognize because they are non-specific. The current treatment for pulmonary arterial hypertension offers no cure or prevention; therefore, it is important to explore treatment avenues for novel pulmonary arterial hypertension treatments. In this study, we tested the hypothesis: pulmonary vasodilator responses of adenosine are dependent on the activation of L-type calcium channels, independent of the synthesis of nitric oxide from L-arginine, activation of adenosine triphosphate-sensitive potassium channels, and the release of cyclooxygenase products. METHODS: We performed an isolated lobar lung preparation in mongrel cats. The thromboxane A2 analog U-46619 was used to increase lobar arterial pressure to a high steady level. We recorded responses to adenosine and other vasodepressor agents in the pulmonary vascular bed of a cat under conditions of controlled pulmonary blood flow and constant left atrial pressure. RESULTS: These data show that adenosine has significant vasodepressor activity in the pulmonary vascular bed of the cat. The data suggest that pulmonary vasodilator responses to adenosine are partially dependent on the activation of adenosine 1 and 2 receptor pathways, and independent of the activation of cyclooxygenase activation, adenosine triphosphate-sensitive K + channels, or synthesis of nitric oxide in the pulmonary vascular bed of the cat. CONCLUSIONS: Vasodepressor effects of adenosine are species specific, and this species specificity will impact the development of future testing and treatments for pulmonary arterial hypertension. Clinical studies are warranted to see if adenosine moieties could play a therapeutic role in patients with pulmonary arterial hypertension and/or other pulmonary pathogeneses.


Assuntos
Adenosina/uso terapêutico , Canais de Cálcio Tipo L/metabolismo , Hipertensão Pulmonar/tratamento farmacológico , Vasodilatadores/uso terapêutico , Animais , Gatos , Modelos Animais de Doenças , Feminino , Hipertensão Pulmonar/metabolismo , Masculino , Óxido Nítrico/metabolismo , Canais de Potássio/metabolismo , Prostaglandina-Endoperóxido Sintases/metabolismo , Receptor A1 de Adenosina/metabolismo , Receptores A2 de Adenosina/metabolismo , Transdução de Sinais
16.
J Neurosci ; 39(36): 7206-7217, 2019 09 04.
Artigo em Inglês | MEDLINE | ID: mdl-31315945

RESUMO

Adenosine 2A receptor (A2AR)-containing indirect medium spiny neurons (iMSNs) in the dorsomedial striatum (DMS) contribute to reward-seeking behaviors. However, those roles for ethanol-seeking behaviors remain unknown. To investigate ethanol-seeking behaviors, we used an ethanol-containing reward (10% ethanol and 10% sucrose solution; 10E10S). Upon conditioning with 10E10S, mice that initially only preferred 10% sucrose, not 10E10S, showed a stronger preference for 10E10S. Then, we investigated whether the manipulation of the DMS-external globus pallidus (GPe) iMSNs circuit alters the ethanol-containing reward (10E10S) seeking behaviors using the combination of pharmacologic and optogenetic approaches. DMS A2AR activation dampened operant conditioning-induced ethanol-containing reward, whereas A2AR antagonist abolished the effects of the A2AR agonist and restored ethanol-containing reward-seeking. Moreover, pre-ethanol exposure potentiated the A2AR-dependent reward-seeking. Interestingly, mice exhibiting ethanol-containing reward-seeking showed the reduction of the DMS iMSNs activity, suggesting that disinhibiting iMSNs decreases reward-seeking behaviors. In addition, we found that A2AR activation reversed iMSNs neural activity in the DMS. Similarly, optogenetic stimulation of the DMS-GPe iMSNs reduced ethanol-containing reward-seeking, whereas optogenetic inhibition of the DMS-GPe iMSNs reversed this change. Together, our study demonstrates that DMS A2AR and iMSNs regulate ethanol-containing reward-seeking behaviors.SIGNIFICANCE STATEMENT Our findings highlight the mechanisms of how operant conditioning develops the preference of ethanol-containing conditioned reward. Mice exhibiting ethanol-containing reward-seeking showed a reduction of the indirect medium spiny neuronal activity in the dorsomedial striatum. Pharmacological activation of adenosine A2A receptor (A2AR) or optogenetic activation of indirect medium spiny neurons dampened operant conditioned ethanol-containing reward-seeking, whereas inhibiting this neuronal activity restored ethanol-containing reward-seeking. Furthermore, repeated intermittent ethanol exposure potentiated A2AR-dependent reward-seeking. Therefore, our finding suggests that A2AR-containing indirect medium spiny neuronal activation reduces ethanol-containing reward-seeking, which may provide a potential therapeutic target for alcohol use disorder.


Assuntos
Alcoolismo/fisiopatologia , Globo Pálido/fisiopatologia , Neurônios/fisiologia , Recompensa , Agonistas do Receptor A3 de Adenosina/farmacologia , Antagonistas do Receptor A3 de Adenosina/farmacologia , Animais , Condicionamento Operante , Etanol/farmacologia , Globo Pálido/citologia , Globo Pálido/fisiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Receptores A2 de Adenosina/metabolismo
17.
PLoS One ; 14(6): e0218619, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31237895

RESUMO

(-)-Incarvillateine (INCA) is a natural product that has garnered attention due to its purported analgesic effects and historical use as a pain reliever in China. α-Truxillic acid monoesters (TAMEs) constitute a class of inhibitors targeting fatty acid binding protein 5 (FABP5), whose inhibition produces analgesia in animal models. The structural similarity between INCA and TAMEs motivated us to assess whether INCA exerts its antinociceptive effects via FABP inhibition. We found that, in contrast to TAMEs, INCA did not exhibit meaningful binding affinities toward four human FABP isoforms (FABP3, FABP4, FABP5 and FABP7) in vitro. INCA-TAME, a putative monoester metabolite of INCA that closely resembles TAMEs also lacked affinity for FABPs. Administration of INCA to mice produced potent antinociceptive effects while INCA-TAME was without effect. Surprisingly, INCA also potently suppressed locomotor activity at the same dose that produces antinociception. The motor suppressive effects of INCA were reversed by the adenosine A2 receptor antagonist 3,7-dimethyl-1-propargylxanthine. Collectively, our results indicate that INCA and INCA-TAME do not inhibit FABPs and that INCA exerts potent antinociceptive and motor suppressive effects at equivalent doses. Therefore, the observed antinociceptive effects of INCA should be interpreted with caution.


Assuntos
Alcaloides/farmacologia , Analgésicos/farmacologia , Locomoção/efeitos dos fármacos , Monoterpenos/farmacologia , Nociceptividade/efeitos dos fármacos , Receptores A2 de Adenosina/metabolismo , Agonistas do Receptor A2 de Adenosina/farmacologia , Antagonistas do Receptor A2 de Adenosina/farmacologia , Animais , Proteínas de Ligação a Ácido Graxo/metabolismo , Humanos , Masculino , Camundongos , Ligação Proteica , Teobromina/análogos & derivados , Teobromina/farmacologia
18.
Chem Commun (Camb) ; 55(40): 5724-5727, 2019 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-31038495

RESUMO

Giant unilamellar protein vesicles (GUPs) were formed with the adenosine A2A receptor (A2AR) incorporated in the lipid bilayer and protein partitioning into the liquid ordered and liquid disordered phases was observed. When no ligand is bound, A2AR partitions preferentially into the liquid disordered phase of GUPs, while ligand-bound A2AR partitions into the liquid ordered phase.


Assuntos
Lipídeos/química , Receptores A2 de Adenosina/metabolismo , Animais , Sítios de Ligação , Células HEK293 , Humanos , Ligantes , Bicamadas Lipídicas , Transição de Fase , Ligação Proteica , Células Sf9
19.
Neurosci Lett ; 704: 141-144, 2019 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-30974229

RESUMO

Traumatic brain injury (TBI) represents one of the leading causes of death worldwide. Its pathophysiology involves several neurochemical events including mitochondrial dysfunction. Since mitochondrial respiration plays a key role in cell survival, pharmacological interventions targeting mitochondrial function have been highlighted as a powerful tool against the neurodegenerative process triggered by TBI. Guanosine (GUO), a neuroprotective molecule in different neurological disorders involving neurotoxicity, has shown protective properties after TBI, however its mechanism of action is not well understood in the central nervous system (CNS). Therefore, the aim of this study is to evaluate the possible target receptor involved in the protective GUO effects on TBI-induced mitochondrial dysfunction in the cerebral cortex of rats. Results show that a single dose of GUO (7.5 mg/kg) injected 40 min after a fluid percussion injury (FPI) protects against loss of mitochondrial membrane potential and increase of reactive oxygen species 8 h post-TBI. These effects were specifically blocked by a pretreatment (10 min after TBI) with an A1 adenosine receptor antagonist (DPCPX 1 mg/kg). In contrast, pretreatment with an A2A adenosine receptor antagonist (SCH 58261 0.05 mg/kg) did not alter GUO effects. These findings suggest that acute GUO neuroprotection following TBI involves the modulation of the adenosinergic system, especially A1 adenosine receptor.


Assuntos
Lesões Encefálicas Traumáticas/tratamento farmacológico , Guanosina/farmacologia , Mitocôndrias/efeitos dos fármacos , Fármacos Neuroprotetores/farmacologia , Receptor A1 de Adenosina/metabolismo , Receptores A2 de Adenosina/metabolismo , Animais , Lesões Encefálicas Traumáticas/metabolismo , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/metabolismo , Guanosina/uso terapêutico , Masculino , Potencial da Membrana Mitocondrial , Mitocôndrias/fisiologia , Fármacos Neuroprotetores/uso terapêutico , Ratos Wistar , Espécies Reativas de Oxigênio/metabolismo
20.
Exp Neurol ; 317: 66-77, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-30822423

RESUMO

Apnea of prematurity (AOP) defined as cessation of breathing for 15-20 s, is commonly seen in preterm infants. Caffeine is widely used to treat AOP due to its safety and effectiveness. Caffeine releases respiratory arrest by competing with adenosine for binding to adenosine A1 and A2A receptors (A1R and A2AR). Long before its use in treating AOP, caffeine has been used as a psychostimulant in adult brains. However, the effect of caffeine on developing brains remains unclear. We found that A1R proteins for caffeine binding were expressed in the brains of neonatal rodents and preterm infants (26-27 weeks). Neonatal A1R proteins colocalized with PSD-95, suggesting its synaptic localization. In contrast, our finding on A2R expression in neonatal neurons was restricted to the mRNA level as detected by single cell RT/PCR due to the lack of specific A2AR antibody. Furthermore, caffeine (200 µM) at a dose twice higher than the clinically relevant dose (36-130 µM) had minor or no effects on several basic neuronal functions, such as neurite outgrowth, synapse formation, expression of A1R and transcription of CREB-1 and c-Fos, further supporting the safety of caffeine for clinical use. We found that treatment with CoCl2 (125 µM), a hypoxia mimetic agent, for 24 h triggered neuronal death and nuclear accumulation of HIF-1α in primary neuronal cultures. Subsequent treatment with caffeine at a concentration of 100 µM alleviated CoCl2-induced cell death and prevented nuclear accumulation of HIF-1α. Consistently, caffeine treatment in early postnatal life of neonatal mice (P4-P7) also prevented subsequent hypoxia-induced nuclear increase of HIF-1α. Together, our data support the utility of caffeine in alleviating hypoxia-induced damages in developing neurons.


Assuntos
Cafeína/farmacologia , Núcleo Celular/metabolismo , Estimulantes do Sistema Nervoso Central/farmacologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Hipóxia/metabolismo , Neurônios/efeitos dos fármacos , Animais , Animais Recém-Nascidos , Núcleo Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cobalto , Hipóxia/induzido quimicamente , Ratos , Receptor A1 de Adenosina/metabolismo , Receptores A2 de Adenosina/metabolismo
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