RESUMO
BACKGROUND AND PURPOSE: Statins, a major component of the prevention of cardiovascular disease, aid progenitor cell functions in vivo and in vitro. Statins bearing a NO-releasing moiety were developed for their enhanced anti-inflammatory/anti-thrombotic properties. Here, we investigated if the NO-donating atorvastatin (NCX 547) improved the functions of circulating angiogenic cells (CACs). EXPERIMENTAL APPROACH: Circulating angiogenic cells (CACs) were prepared from peripheral blood monocytes of healthy volunteers and type-2 diabetic patients and were cultured in low (LG) or high glucose (HG) conditions, in presence of atorvastatin or NCX 547 (both at 0.1 µM) or vehicle. Functional assays (outgrowth, proliferation, viability, senescence and apoptosis) were performed in presence of the endothelial NOS inhibitor L-NIO, the NO scavenger c-PTIO or vehicle. KEY RESULTS: Culturing in HG conditions lowered NO in CACs, inhibited outgrowth, proliferation, viability and migration, and induced cell senescence and apoptosis. NCX 547 fully restored NO levels and functions of HG-cultured CACs, while atorvastatin prevented only apoptosis in CACs. The activity of Akt, a pro-survival kinase, was increased by atorvastatin in LG-cultured but not in HG-cultured CACs, whereas NCX 547 increased Akt activity in both conditions. L-NIO partially blunted and c-PTIO prevented NCX 547-induced improvements in CAC functions. Finally, NCX 547 improved outgrowth and migration of CACs prepared from patients with type 2 diabetes. CONCLUSIONS AND IMPLICATIONS: NCX 547 was more effective than atorvastatin in preserving functions of CACs. This property adds to the spectrum of favourable actions that would make NO-releasing statins more effective agents for treating cardiovascular disease.
Assuntos
Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Neovascularização Fisiológica/efeitos dos fármacos , Nitratos/farmacologia , Doadores de Óxido Nítrico/farmacologia , Pirróis/farmacologia , Células-Tronco/efeitos dos fármacos , Idoso , Anticolesterolemiantes/farmacologia , Apoptose/efeitos dos fármacos , Benzoatos/farmacologia , Fármacos Cardiovasculares/farmacologia , Doenças Cardiovasculares/tratamento farmacológico , Doenças Cardiovasculares/metabolismo , Movimento Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Senescência Celular/efeitos dos fármacos , Diabetes Mellitus Tipo 2/sangue , Diabetes Mellitus Tipo 2/tratamento farmacológico , Feminino , Humanos , Imidazóis/farmacologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Masculino , Pessoa de Meia-Idade , Óxido Nítrico Sintase Tipo III/antagonistas & inibidores , Ornitina/análogos & derivados , Ornitina/farmacologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Células-Tronco/citologia , Células-Tronco/metabolismoRESUMO
The IOP lowering effects of NCX 139, a new chemical entity comprising latanoprost amide and a NO-donating moiety, were compared to those of the respective des-nitro analog in in vitro assays and in rabbit and dog models of ocular hypertension. The NO donor, molsidomine as well as the prostamide bimatoprost (Lumigan(®)) and the prostaglandin agonist, latanoprost (Xalatan(®)) were also investigated for comparison. NCX 139 but not its des-nitro analog resulted in NO-mediated vascular relaxant effect in pre-contracted rabbit aortic rings (EC(50)=0.70±0.06 µM; E(max)=80.6±2.9%). Like bimatoprost (IC(50)=3.07±1.3 µM) or latanoprost (IC(50)=0.48±0.15 µM), NCX 139 displaced (3)H-PGF2α binding on recombinant human prostaglandin-F (FP) receptors with an estimated potency of 0.77±0.13 µM. In transient ocular hypertensive rabbits, bimatoprost and latanoprost were not effective while molsidomine elicited a dose-dependent reduction of IOP confirming the responsiveness of rabbits to NO but not to FP receptor agonists. NCX 139 tested at a therapeutically relevant dose, significantly lowered IOP while the des-nitro analog was not effective (0.03% NCX 139, Δ(max)=-12.8±2.0 mmHg). In glaucomatous dogs, 0.03% NCX 139 decreased IOP to a greater extent compared to an equimolar dose of the respective des-nitro derivative (Δ(max)=-4.6±1.0 and -2.7±1.3 mmHg, respectively for NCX 139 and its des-nitro analog). Albeit with low potency, NCX 139 also resulted effective in normotensive dogs while it did not reduce IOP in normotensive rabbits. NCX 139, a compound targeting two different and important mechanisms, is endowed with ocular hypotensive effects more evident in hypertensive conditions which may be of interest in the search of more effective treatments for hypertensive glaucoma.
Assuntos
Anti-Hipertensivos/farmacologia , Glaucoma/tratamento farmacológico , Pressão Intraocular/efeitos dos fármacos , Nitratos/farmacologia , Doadores de Óxido Nítrico/farmacologia , Óxido Nítrico/metabolismo , Prostaglandinas F Sintéticas/metabolismo , Prostaglandinas F Sintéticas/farmacologia , Amidas/farmacologia , Animais , Anti-Hipertensivos/química , Aorta/efeitos dos fármacos , Bimatoprost , Cromatografia Líquida de Alta Pressão , Cloprostenol/análogos & derivados , Cloprostenol/farmacologia , Dinoprosta/metabolismo , Modelos Animais de Doenças , Cães , Glaucoma/metabolismo , Latanoprosta , Masculino , Molsidomina/farmacologia , Nitratos/química , Doadores de Óxido Nítrico/química , Hipertensão Ocular/tratamento farmacológico , Hipertensão Ocular/metabolismo , Prostaglandinas F Sintéticas/química , Coelhos , Espectrometria de Massas em Tandem , Tonometria Ocular , Vasodilatação/fisiologia , Vasodilatadores/farmacologiaRESUMO
BACKGROUND AND PURPOSE: Mature endothelial cells and their progenitors are dysfunctional in diabetes, resulting in deficient neovascularisation following arterial occlusion. This study aimed to evaluate the therapeutic activity of a nitric oxide (NO) releasing statin in the setting of experimental diabetes and peripheral ischaemia. EXPERIMENTAL APPROACH: The effects of NCX 6550, an NO-releasing pravastatin derivative, on angiogenesis in ischaemic limbs was studied in normoglycaemic mice or mice made diabetic by treatment with streptozotocin (STZ). Control mice received an equimolar dosage of the parent statin compound, pravastatin. The therapeutic action of NCX 6550 was also tested in mice lacking the gene for endothelial nitric oxide synthase (eNOS). KEY RESULTS: In normoglycaemic or STZ-diabetic CD1 mice, only NCX 6550 stimulated skeletal muscle revascularisation. In addition, NCX 6550 induced greater improvement in limb reperfusion and salvage, than pravastatin. The number of circulating endothelial progenitor cells was decreased in STZ-diabetic mice, this defect being prevented by NCX 6550 and, to a lesser extent by pravastatin. In vitro, high glucose concentrations reduced the migratory capacity of endothelial progenitor EPCs, which was partly reversed by preincubation with pravastatin and completely reversed by NCX 6550. The postischaemic recovery of eNOS knockout mice was severely impaired as a consequence of depressed angiogenesis and this recovery was improved by treatment with NCX 6550, but not with pravastatin. CONCLUSIONS AND IMPLICATIONS: These findings indicate that incorporation of a bioactive NO moiety improves the therapeutic profile of statins for the treatment of peripheral vascular disease.
Assuntos
Diabetes Mellitus Experimental/tratamento farmacológico , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Isquemia/tratamento farmacológico , Neovascularização Fisiológica/efeitos dos fármacos , Nitratos/uso terapêutico , Pravastatina/análogos & derivados , Pravastatina/uso terapêutico , Animais , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Diabetes Mellitus Tipo 1/tratamento farmacológico , Diabetes Mellitus Tipo 1/metabolismo , Diabetes Mellitus Tipo 1/patologia , Membro Posterior/irrigação sanguínea , Hipertensão/tratamento farmacológico , Hipertensão/metabolismo , Hipertensão/patologia , Isquemia/metabolismo , Isquemia/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/deficiência , Óxido Nítrico Sintase Tipo III/genéticaRESUMO
BACKGROUND: Topical nitric oxide-releasing dexamethasone (NCX1021) may avoid the negative effects of dexamethasone phosphate. AIMS: To obtain more information on the role of nitric oxide in glaucoma and to compare a nitric oxide-releasing dexamethasone with dexamethasone phosphate with regard to intraocular pressure (IOP) and ocular haemodynamics in an experimental rabbit model. METHODS: Six rabbits were treated with dexamethasone phosphate 0.1% in the right eye and with NCX1021 in the left eye for 5 weeks. The parameters considered were IOP, nitric oxide marker levels in aqueous humour, ocular haemodynamics of ophthalmic artery (by means of colour Doppler imaging), expression of endothelial nitric oxide synthase (eNOS)in ciliary processes and histology of ciliary bodies. RESULTS: Dexamethasone increased IOP levels, NCX1021 did not. Nitrite and cyclic guanosine monophosphate levels in aqueous humour were lowered by dexamethasone and increased by NCX1021. Resistivity index of the ophthalmic artery was increased, eNOS expression was reduced and ciliary bodies showed histological lesions in dexamethasone-treated eyes, not in NCX1021-treated ones. CONCLUSIONS: NCX1021 may avoid the IOP increase, impairment of ocular blood flow and the morphological changes in the ciliary bodies possibly induced by corticosteroid treatment.
Assuntos
Dexametasona/análogos & derivados , Glaucoma/tratamento farmacológico , Glucocorticoides/farmacologia , Pressão Intraocular/efeitos dos fármacos , Doadores de Óxido Nítrico/farmacologia , Animais , Humor Aquoso/química , Western Blotting/métodos , Corpo Ciliar/química , Corpo Ciliar/efeitos dos fármacos , Dexametasona/química , Dexametasona/farmacologia , Glaucoma/metabolismo , Masculino , Modelos Animais , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo III/análise , Nitritos/análise , Artéria Oftálmica/efeitos dos fármacos , Coelhos , Resistência Vascular/efeitos dos fármacosRESUMO
BACKGROUND: NO-releasing statins are new chemical entities, combining HMG-CoA reductase inhibition and slow NO release, that possess stronger anti-inflammatory and antiproliferative activities than the native statins. OBJECTIVE: We evaluated the antithrombotic effects of nitropravastatin (NCX-6550) by assessing its activity on platelet activation and tissue factor (TF) expression by mononuclear cells in vitro and in vivo. METHODS AND RESULTS: In vitro, NCX-6550 inhibited (1) U46619- and collagen-induced platelet aggregation in buffer and plasma; (2) collagen-induced P-selectin expression in whole blood and (3) platelet adhesion to collagen-coated coverslips under high shear stress. These effects were displayed at concentrations of NCX-6550 ranging from 25 to 100 mum, and were totally reverted by the guanylylcyclase inhibitor ODQ (10 microm). Equimolar concentrations of pravastatin had no influence on these parameters of platelet function. LPS- and PMA-induced TF expression by blood mononuclear cells was also inhibited by NCX-6550 (IC50 13 microm), but not by pravastatin, as assessed by functional and immunological assays and by real-time PCR. In a mouse model of platelet pulmonary thromboembolism, induced by the i.v. injection of collagen plus epinephrine, pretreatment with NCX-6550 (24-48 mg kg(-1)) significantly reduced platelet consumption, lung vessel occlusion and mortality. Moreover, nitropravastatin markedly inhibited the generation of procoagulant activity by spleen mononuclear cells and peritoneal macrophages in mice treated with LPS. In these in vivo models too, pravastatin failed to affect platelet activation and monocyte/macrophage procoagulant activity. CONCLUSIONS: Our results show that nitropravastatin exerts strong antithrombotic effects in vitro and in vivo, and may represent an interesting antiatherothrombotic agent for testing in acute coronary syndromes.
Assuntos
Plaquetas/efeitos dos fármacos , Doadores de Óxido Nítrico/farmacologia , Nitrocompostos/farmacologia , Pravastatina/farmacologia , RNA Mensageiro/metabolismo , Tromboplastina/metabolismo , Animais , Plaquetas/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Humanos , Técnicas In Vitro , Concentração Inibidora 50 , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Macrófagos Peritoneais/efeitos dos fármacos , Macrófagos Peritoneais/metabolismo , Masculino , Camundongos , Nitratos/sangue , Óxido Nítrico/metabolismo , Doadores de Óxido Nítrico/administração & dosagem , Nitritos/sangue , Nitrocompostos/administração & dosagem , Selectina-P/metabolismo , Adesividade Plaquetária/efeitos dos fármacos , Agregação Plaquetária/efeitos dos fármacos , Pravastatina/administração & dosagem , Pravastatina/análogos & derivados , Embolia Pulmonar/mortalidade , Embolia Pulmonar/patologia , Embolia Pulmonar/prevenção & controle , Baço/citologia , Baço/efeitos dos fármacos , Baço/metabolismo , Tromboplastina/genéticaRESUMO
Here, we report the antimycobacterial activity of NCX 976, a new molecule obtained adding a NO moiety to the fluoroquinolone ciprofloxacin, on Mycobacterium tuberculosis H37Rv strain, both in a cell-free model and in infected human macrophages. Unlike unaltered ciprofloxacin, NCX976 displayed a marked activity also at low-nanomolar concentrations.
Assuntos
Anti-Infecciosos/farmacologia , Ciprofloxacina/farmacologia , Mycobacterium tuberculosis/efeitos dos fármacos , Óxido Nítrico/química , Anti-Infecciosos/química , Sistema Livre de Células , Ciprofloxacina/química , Desenho de Fármacos , Humanos , Macrófagos/microbiologia , Mycobacterium tuberculosis/crescimento & desenvolvimentoRESUMO
The selective A2A receptor antagonist [3H]SCH 58261 was injected intravenously in mice and the radioactivity accumulating in various brain regions was determined by tissue sampling. Radioactivity levels in regions of interest such as the striatum were highest 15 min after injection and quickly declined thereafter (30 min and 1 h postinjection) in a time-dependent manner. The amount of labelling was ranked as follows: striatum (4.6 +/- 0.3 fmol/mg protein) >> cortex > hippocampus > pons = hypothalamus > cerebellum (0.5 +/- 0.05 fmol/mg protein). Specific labelling of the A2A receptor occurred in striatum and cortex because significantly less radioactivity accumulated in these areas from adenosine A2A receptor knockout mice as compared to wild-type littermates. In control outbred CD1 mice, a striatum-to-cerebellum ratio of 7.6 +/- 0.6 was found. At 30 min postinjection, the nonselective adenosine receptor antagonist caffeine reduced the radioactivity due to [3H]SCH 58261 in the striatum by 32% at 1 mg/kg i.p. and by 66% at the stimulant dose of 6.25 mg/kg i.p. Radioactivity in the striatum was lowered, respectively, by 66 and 86% 30 min after injection of 3 or 10 mg/kg i.p. doses of unlabelled SCH 58261. The present results indicate that [3H]SCH 58261 directly labels striatal A2A receptors in vivo. Thus [3H]SCH 58261 is an excellent tool for studying brain distribution and A2A receptor occupancy of various compounds ranging from xanthines, such as caffeine, to other A2A antagonists.
Assuntos
Adenosina/metabolismo , Encéfalo/efeitos dos fármacos , Neurônios/efeitos dos fármacos , Fármacos Neuroprotetores/metabolismo , Antagonistas de Receptores Purinérgicos P1 , Pirimidinas/metabolismo , Triazóis/metabolismo , Animais , Sítios de Ligação/efeitos dos fármacos , Sítios de Ligação/fisiologia , Ligação Competitiva/efeitos dos fármacos , Ligação Competitiva/fisiologia , Barreira Hematoencefálica/efeitos dos fármacos , Barreira Hematoencefálica/fisiologia , Encéfalo/citologia , Encéfalo/metabolismo , Cafeína/farmacologia , Relação Dose-Resposta a Droga , Interações Medicamentosas/fisiologia , Masculino , Camundongos , Camundongos Knockout , Neurônios/metabolismo , Inibidores de Fosfodiesterase/farmacologia , Ensaio Radioligante , Receptor A2A de Adenosina , Receptores Purinérgicos P1/deficiência , Receptores Purinérgicos P1/genética , Trítio/metabolismoRESUMO
1. Adenosine, an ubiquitous neuromodulator, and its analogues have been shown to produce 'depressant' effects in animal models believed to be relevant to depressive disorders, while adenosine receptor antagonists have been found to reverse adenosine-mediated 'depressant' effect. 2. We have designed studies to assess whether adenosine A2A receptor antagonists, or genetic inactivation of the receptor would be effective in established screening procedures, such as tail suspension and forced swim tests, which are predictive of clinical antidepressant activity. 3. Adenosine A2A receptor knockout mice were found to be less sensitive to 'depressant' challenges than their wildtype littermates. Consistently, the adenosine A2A receptor blockers SCH 58261 (1 - 10 mg kg(-1), i.p.) and KW 6002 (0.1 - 10 mg kg(-1), p.o.) reduced the total immobility time in the tail suspension test. 4. The efficacy of adenosine A2A receptor antagonists in reducing immobility time in the tail suspension test was confirmed and extended in two groups of mice. Specifically, SCH 58261 (1 - 10 mg kg(-1)) and ZM 241385 (15 - 60 mg kg(-1)) were effective in mice previously screened for having high immobility time, while SCH 58261 at 10 mg kg(-1) reduced immobility of mice that were selectively bred for their spontaneous 'helplessness' in this assay. 5. Additional experiments were carried out using the forced swim test. SCH 58261 at 10 mg kg(-1) reduced the immobility time by 61%, while KW 6002 decreased the total immobility time at the doses of 1 and 10 mg kg(-1) by 75 and 79%, respectively. 6. Administration of the dopamine D2 receptor antagonist haloperidol (50 - 200 microg kg(-1) i.p.) prevented the antidepressant-like effects elicited by SCH 58261 (10 mg kg(-1) i.p.) in forced swim test whereas it left unaltered its stimulant motor effects. 7. In conclusion, these data support the hypothesis that A2A receptor antagonists prolong escape-directed behaviour in two screening tests for antidepressants. Altogether the results support the hypothesis that blockade of the adenosine A2A receptor might be an interesting target for the development of effective antidepressant agents.
Assuntos
Antidepressivos/farmacologia , Antagonistas de Receptores Purinérgicos P1 , Animais , Comportamento Animal/efeitos dos fármacos , Blefaroptose/induzido quimicamente , Blefaroptose/prevenção & controle , Modelos Animais de Doenças , Antagonistas de Dopamina/farmacologia , Antagonistas dos Receptores de Dopamina D2 , Relação Dose-Resposta a Droga , Feminino , Haloperidol/farmacologia , Imobilização , Masculino , Camundongos , Camundongos Knockout , Atividade Motora/efeitos dos fármacos , Purinas/farmacologia , Pirimidinas/farmacologia , Receptor A2A de Adenosina , Receptores de Dopamina D2/fisiologia , Receptores Purinérgicos P1/genética , Reserpina/administração & dosagem , Natação , Fatores de Tempo , Triazóis/farmacologiaRESUMO
Nociceptin or orphanin FQ (N/OFQ) is the natural ligand of the opioid receptor-like 1 receptor (ORL-1), which has been also classified as the fourth member of the opioid family of receptors and named OP(4). Elucidation of the biological role of N/OFQ has been hampered by the lack of compounds that selectively block the OP(4) receptor. Recently, a N/OFQ derivative, [Nphe(1)]N/OFQ(1-13)NH(2), has been found to possess OP(4) antagonistic properties both in vitro and in vivo models. We investigated its spinal effect in the chronic constriction injury of the sciatic nerve in the rat, a model relevant to neuropathic pain in humans. Intrathecal (i.t.) administration of N/OFQ (0.2--20 nmoles) dose-dependently reversed mechanical allodynic-like behavior, while [Nphe(1)]N/OFQ(1-13)NH(2) (20--120 nmoles, i.t.) was ineffective on its own. [Nphe(1)]N/OFQ(1-13)NH(2) (60--120 nmoles, i.t.) antagonized N/OFQ (about 80% of reduction) but did not modify the activity of morphine (20 nmoles, i.t.). These results further support, for the first time in a chronic model of pain, the specific antagonistic profile of [Nphe(1)]N/OFQ(1-13)NH(2)vs the OP(4) receptor. This pseudopeptide is an interesting pharmacological tool to better clarify the role of N/OFQ in pathophysiology.
Assuntos
Aminas , Ácidos Cicloexanocarboxílicos , Antagonistas de Entorpecentes , Neuralgia/tratamento farmacológico , Peptídeos Opioides/antagonistas & inibidores , Peptídeos Opioides/farmacologia , Fragmentos de Peptídeos/farmacologia , Doenças do Sistema Nervoso Periférico/tratamento farmacológico , Ácido gama-Aminobutírico , Acetatos/farmacologia , Analgésicos/farmacologia , Animais , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Interações Medicamentosas/fisiologia , Gabapentina , Hiperalgesia/tratamento farmacológico , Hiperalgesia/metabolismo , Hiperalgesia/fisiopatologia , Masculino , Morfina/farmacologia , Entorpecentes/farmacologia , Neuralgia/metabolismo , Neuralgia/fisiopatologia , Limiar da Dor/efeitos dos fármacos , Doenças do Sistema Nervoso Periférico/metabolismo , Doenças do Sistema Nervoso Periférico/fisiopatologia , Ratos , Ratos Sprague-Dawley , Receptores Opioides/metabolismo , Nervo Isquiático/cirurgia , Medula Espinal/efeitos dos fármacos , Medula Espinal/metabolismo , Medula Espinal/fisiopatologia , Receptor de Nociceptina , NociceptinaRESUMO
In the early 1990s it became clear that the A2A adenosine receptor had characteristics that made it distinct from the other A1, A2B and A3 adenosine receptors. Great progress has been made with the discovery of selective A2A receptor antagonists. A variety of synthetic substitutions on the xanthine moiety led the chemists of Kyowa-Hakko to discover that introduction of the styryl group in the 8 position of xanthines was critical in achieving compounds endowed with selective A2A receptor antagonistic properties. One compound, KW 6002, (E)1,3-diethyl-8-(3,4-dimethoxystyryl)-7-methylxanthine, is currently being developed for treatment of Parkinson's disease. A number of non-xanthine heterocycles have also been synthesized starting from the non-selective adenosine antagonist CGS 15943, a triazoloquinazoline. Thus, replacement of the phenyl ring of CGS 15943 with a heterocyclic ring such as pyrazole or imidazole, led to a series of interesting compounds whose prototype, SCH 58261, 7-(2-phenylethyl)-5-amino-2-(2-furyl)-pyrazolo[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine, has become a reference A2A receptor antagonist. Modification of N7 substituents has progressed to optimize A2A receptor selectivity and pharmacokinetic characteristics. A related class of compounds having a bicyclic instead of the tricyclic ring structure is also of interest. The prototype of these triazintriazolo derivatives, ZM 241385, is a potent A2A receptor antagonist; however, it also shows interactions with A2B receptors. The relevance of the A2A receptors in specific disease states, especially in the central nervous system, makes this class of adenosine receptor blockers of interest for treatment of neurodegenerative disorders such as Parkinson's disease.
Assuntos
Antagonistas de Receptores Purinérgicos P1 , Animais , Humanos , Doenças Neurodegenerativas/tratamento farmacológico , Purinas/farmacologia , Pirimidinas/farmacologia , Receptor A2A de Adenosina , Triazóis/farmacologiaRESUMO
The A(2A)R is largely coexpressed with D(2)Rs and enkephalin mRNA in the striatum where it modulates dopaminergic activity. Activation of the A(2A)R antagonizes D(2)R-mediated behavioral and neurochemical effects in the basal ganglia through a mechanism that may involve direct A(2A)R-D(2)R interaction. However, whether the D(2)R is required for the A(2A)R to exert its neural function is an open question. In this study, we examined the role of D(2)Rs in A(2A)R-induced behavioral and cellular responses, by using genetic knockout (KO) models (mice deficient in A(2A)Rs or D(2)Rs or both). Behavioral analysis shows that the A(2A)R agonist 2-4-(2-carboxyethyl)phenethylamino-5'-N-ethylcarboxamidoadenosine reduced spontaneous as well as amphetamine-induced locomotion in both D(2) KO and wild-type mice. Conversely, the nonselective adenosine antagonist caffeine and the A(2A)R antagonist 8-(3-chlorostyryl)caffeine produced motor stimulation in mice lacking the D(2)R, although the stimulation was significantly attenuated. At the cellular level, A(2A)R inactivation counteracted the increase in enkephalin expression in striatopallidal neurons caused by D(2)R deficiency. Consistent with the D(2) KO phenotype, A(2A)R inactivation partially reversed both acute D(2)R antagonist (haloperidol)-induced catalepsy and chronic haloperidol-induced enkephalin mRNA expression. Together, these results demonstrate that A(2A)Rs elicit behavioral and cellular responses despite either the genetic deficiency or pharmacological blockade of D(2)Rs. Thus, A(2A)R-mediated neural functions are partially independent of D(2)Rs. Moreover, endogenous adenosine acting at striatal A(2A)Rs may be most accurately viewed as a facilitative modulator of striatal neuronal activity rather than simply as an inhibitory modulator of D(2)R neurotransmission.
Assuntos
Adenosina/análogos & derivados , Atividade Motora/fisiologia , Receptores de Dopamina D2/fisiologia , Receptores Purinérgicos P1/fisiologia , Adenosina/farmacologia , Anfetaminas/farmacologia , Animais , Cafeína/análogos & derivados , Cafeína/farmacologia , Catalepsia/patologia , Corpo Estriado/metabolismo , Corpo Estriado/patologia , Antagonistas de Dopamina/farmacologia , Encefalinas/biossíntese , Encefalinas/genética , Expressão Gênica , Haloperidol/farmacologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fenetilaminas/farmacologia , Agonistas do Receptor Purinérgico P1 , Antagonistas de Receptores Purinérgicos P1 , RNA Mensageiro , Receptor A2A de Adenosina , Receptores de Dopamina D1/biossíntese , Receptores de Dopamina D2/biossíntese , Receptores Purinérgicos P1/biossínteseRESUMO
A direct pathophysiological role of Familial Alzheimer's Disease (FAD)-associated Presenilin 1 (PS1) mutations in neuronal vulnerability remains a controversial matter. We evaluated the relationship between PS1 and excitotoxicity in four different experimental models of neurotoxicity by using primary neurons from (i) transgenic (tg) mice overexpressing a human FAD-linked PS1 variant (L286V mutation), (ii) tg mice overexpressing human wild-type (wt) PS1, (iii) PS1 knockout mice, and (iv) wt mice in which PS1 gene expression was knocked down by antisense treatment. We found that primary neurons overexpressing mutated PS1 showed an increased vulnerability to both excitotoxic and hypoxic-hypoglycemic damage when compared with neurons obtained from either mice overexpressing human wt PS1 or in wt mice. In addition, reduced excitotoxic damage was obtained in neurons in which PS1 expression was absent or diminished. Data obtained in in vivo experimental models of excitotoxicity partially supported the in vitro observations. Accelerated neuronal death was demonstrated in the hippocampus of mice overexpressing mutated PS1 after peripheral administration of kainic acid in comparison with wt animals. However, measurement of the infarct volume after middle cerebral artery occlusion did not show significant difference between the two animal groups. The results altogether suggest that expression of FAD-linked PS1 variants increases the vulnerability of neurons to a specific type of damage in which excitotoxicity plays a relevant role. In addition, they support the view that reduction of endogenous PS1 expression results in neuroprotection.
Assuntos
Doença de Alzheimer/metabolismo , Agonistas de Aminoácidos Excitatórios/farmacologia , Ácido Caínico/farmacologia , Proteínas de Membrana/metabolismo , N-Metilaspartato/farmacologia , Neurônios/efeitos dos fármacos , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/patologia , Células Cultivadas , Modelos Animais de Doenças , Expressão Gênica/efeitos dos fármacos , Humanos , Infarto da Artéria Cerebral Média/fisiopatologia , Proteínas de Membrana/genética , Proteínas de Membrana/fisiologia , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Degeneração Neural , Neurônios/citologia , Neurônios/patologia , Neurônios/fisiologia , Presenilina-1Assuntos
Peptídeos Opioides , Receptores Opioides , Animais , Vias Autônomas/efeitos dos fármacos , Comportamento Aditivo/tratamento farmacológico , Humanos , Antagonistas de Entorpecentes , Peptídeos Opioides/farmacologia , Peptídeos Opioides/uso terapêutico , Dor/tratamento farmacológico , Receptores Opioides/agonistas , Receptor de Nociceptina , NociceptinaRESUMO
Interleukin-10 (IL-10) is a powerful suppressor of cellular immune responses, with a postulated role in brain inflammation. First, we have evaluated the role of this cytokine in ischaemic brain damage using IL-10 knockout (IL-10-/-) mice. The middle cerebral artery (MCA) was occluded in either IL-10-/- or wild-type animals of corresponding strain (C57Bl/6) and age. Infarct volume was assessed 24 h later in serial brain sections. Brain infarct produced by MCA occlusion was 30% larger in the IL-10-/- than in wild-type mice (21. 8 +/- 1.2 vs. 16.9 +/- 1.0 mm3, respectively; P < 0.01; Student's t-test). To further characterize these findings, studies were extended to in vitro models. Primary neuronal cortical cultures derived from IL-10-/- animals were more susceptible to both excitotoxicity and combined oxygen-glucose deprivation compared with cell cultures from wild-type mice. Moreover, when added to the culture medium, recombinant murine IL-10 (0.1-100 ng/mL) exerted a concentration-dependent prevention of neuronal damage induced by excitotoxicity in both cortical and cerebellar granule cell cultures taken from either strain. The accordance of in vivo and in vitro data allows us to suggest a potential neuroprotective role of IL-10 against cerebral ischaemia when administered exogenously or made available from endogenous sources.
Assuntos
Isquemia Encefálica/imunologia , Encéfalo/imunologia , Interleucina-10/genética , Interleucina-10/imunologia , Animais , Encéfalo/irrigação sanguínea , Encéfalo/citologia , Isquemia Encefálica/patologia , Morte Celular/efeitos dos fármacos , Morte Celular/fisiologia , Células Cultivadas , Agonistas de Aminoácidos Excitatórios/farmacologia , Glucose/farmacologia , Infarto da Artéria Cerebral Média/imunologia , Infarto da Artéria Cerebral Média/patologia , Interleucina-10/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , N-Metilaspartato/farmacologia , Necrose , Neurônios/citologia , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Neurotoxinas/farmacologia , Oxigênio/farmacologia , Proteínas Recombinantes/farmacologia , Fator de Necrose Tumoral alfa/biossíntese , Fator de Necrose Tumoral alfa/imunologiaRESUMO
BACKGROUND-We determined whether repeated caffeine administration at different dosages and for different periods of time (400 or 600 mg/d for 1 week or 400 mg/d for 2 weeks) upregulates human platelet adenosine A(2A) receptors and is accompanied by increases in cAMP accumulation and decreases in aggregation and calcium levels after stimulation of A(2A) receptors by the selective agonist 2-hexynyl-5'-N-ethylcarboxamidoadenosine (HE-NECA). METHODS AND RESULTS-Platelets were obtained from peripheral venous blood of 45 healthy human volunteers at the end of 2 weeks of caffeine abstinence and at 12, 60, and 108 hours after the last dose of caffeine. The lowest dose of caffeine, when given for only 7 days, had no effect. Increasing the total dose, either by giving 400 mg/d for 14 days or giving 600 mg/d, resulted in binding assays performed with the adenosine A(2A) receptor radioligand [(3)H]SCH 58261 [5-amino-7(phenylethyl)-2-(2-furyl)-pyrazolo[4,3-e]-1,2,4-triazolo[1, 5-c]pyrimidine], in the upregulation of A(2A) receptors. Moreover, the potency of HE-NECA to produce antiaggregatory effects, a rise in cAMP accumulation, and a decrease in calcium levels was significantly increased. CONCLUSIONS-Chronic caffeine intake can lead to upregulation of adenosine A(2A) receptors, which is accompanied by sensitization, in a time- and dose-dependent manner, to the actions of the agonist HE-NECA.
Assuntos
Plaquetas/metabolismo , Cafeína/farmacologia , Receptores Purinérgicos P1/sangue , Adenosina-5'-(N-etilcarboxamida)/análogos & derivados , Adenosina-5'-(N-etilcarboxamida)/farmacologia , Adulto , Cálcio/metabolismo , AMP Cíclico/metabolismo , Relação Dose-Resposta a Droga , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Miocárdio/metabolismo , Agregação Plaquetária/efeitos dos fármacos , Inibidores da Agregação Plaquetária/farmacologia , Agonistas do Receptor Purinérgico P1 , Pirimidinas/metabolismo , Receptor A2A de Adenosina , Receptores Purinérgicos P1/fisiologia , Fatores de Tempo , Triazóis/metabolismo , Regulação para CimaRESUMO
Immunophilin ligands such as rapamycin, FK506 and GPI-1046 have been reported to increase neurite outgrowth in vitro and to have neuroprotective activity in vitro and in vivo. In this study, however, FK506 and GPI-1046 (0.1-1000 nM) had little effect on neurite outgrowth in PC12 cells in either the presence or absence of nerve growth factor. In contrast, rapamycin markedly increased neurite outgrowth in PC12 cells in the presence of a low concentration of nerve growth factor (EC(50)=10 nM). Unlike FK506 and GPI-1046, rapamycin is an inhibitor of cell cycle progression. Other cell cycle inhibitors such as ciclopirox and flavopiridol also increased neurite outgrowth in PC12 cells in the presence of a low concentration of nerve growth factor (EC(50)=250 nM and 100 nM, respectively). The neuroprotective effects of FK506, rapamycin and GPI-1046 were also tested in a rodent model of permanent focal cerebral ischemia. FK506 and rapamycin decreased infarct volume by 40% and 37%, respectively, whereas GPI-1046 was ineffective. These data do not support the previous suggestion that FK506 and GPI-1046 increase neurite outgrowth of PC12 cells in vitro. Rapamycin increases neurite outgrowth of PC12 cells, an effect that can be ascribed to its ability to inhibit cell cycle progression. The neuroprotective effect of FK506 and rapamycin against cerebral ischemia is probably not due to differentiation of neuronal precursors or stimulation of neuronal regeneration.
Assuntos
Ciclo Celular/efeitos dos fármacos , Neuritos/efeitos dos fármacos , Pirrolidinas/farmacologia , Sirolimo/farmacologia , Tacrolimo/farmacologia , Animais , Isquemia Encefálica/patologia , Isquemia Encefálica/prevenção & controle , Ciclo Celular/fisiologia , Ciclopirox , Relação Dose-Resposta a Droga , Flavonoides/farmacologia , Inibidores do Crescimento/farmacologia , Masculino , Fator de Crescimento Neural/farmacologia , Neuritos/fisiologia , Células PC12 , Piperidinas/farmacologia , Piridonas/farmacologia , Ratos , Ratos Sprague-DawleyRESUMO
1 Stimulation of the opioid receptor-like1 (ORL-1) receptor by nociceptin (NC) produces hyperalgesia and reverses the antinociceptive effects induced by opioids. Most studies concerning the central effects of NC were conducted using acute pain models. The role NC may play in chronic inflammation remains unelucidated. 2 The present study was undertaken to assess the action of NC in the Freund's adjuvant-induced monoarthritic rat model. The effects of drugs known to act as analgesics in this model were evaluated. The effects of NC, NCNH2, and the ORL-1 ligand, [Phe1psi(CH2-NH)Gly2]NC(1-13)NH2 ([F/G]NC(1-13)NH2), were also studied alone or in association with morphine. 3 NC (1 - 30 nmol, i. c.v.) was inactive, whilst NCNH2 (10 nmol, i.c.v.) exerted hyperalgesic effects (-4.5+/-0.9 vs -0.7+/-0.8 s of vehicle-treated animals). [F/G]NC(1-13)NH2 (0.01 - 10 nmol, i.c.v.) induced hyperalgesia in the arthritic paw (-3.3+/-0.6 vs -0.3+/-0.5 s of vehicle-treated animals; 10 nmol). 4 Both NC (0.01 - 10 nmol, i.c.v. ) and [F/G]NC(1-13)NH2 (0.01 - 1 nmol, i.c.v), 30 min after morphine (3 mg kg-1, s.c.) induced an immediate and short-lived reversal of morphine effects (2.6+/-0.3 vs 10.4+/-1.0 and 1.2+/-1.5 vs 9.3+/-1.1 s of morphine alone, respectively), therefore displaying anti-opioid activity. 5 In the Freund's adjuvant-induced rat model of arthritis, both NC and [F/G]NC(1-13)NH2 act as anti-opioid peptides. Furthermore, NCNH2 and [F/G]NC(1-13)NH2 induce hyperalgesia when given alone. Further investigations and the identification of a centrally acting ORL-1 antagonist are necessary to better understand the role of NC in pain mechanisms.
Assuntos
Artrite/tratamento farmacológico , Adjuvante de Freund/efeitos adversos , Entorpecentes/farmacologia , Peptídeos Opioides/farmacologia , Dor/tratamento farmacológico , Fragmentos de Peptídeos/farmacologia , Animais , Artrite/induzido quimicamente , Artrite/complicações , Doença Crônica , Modelos Animais de Doenças , Interações Medicamentosas , Ligantes , Masculino , Morfina/farmacologia , Atividade Motora/efeitos dos fármacos , Dor/etiologia , Ratos , Ratos Endogâmicos Lew , Receptores Opioides/metabolismo , Receptor de Nociceptina , NociceptinaRESUMO
Reports of serious cardiac arrhythmia associated with some second-generation antihistamines have prompted concern for their prescription. This article reviews the nature of the adverse events reported and concludes that the blockade of potassium channels, particularly the subtype responsible for the rapid component of the delayed rectifier current (IKr), is largely responsible for such adverse cardiac events. Consequently, antihistamines with little or no interaction with these channels are expected to have the greatest safety margin. The main cardiac arrhythmia of concern is that of torsades de pointes, a potentially fatal phenomenon characterized by prolonged ventricular depolarization that manifests as a prolonged QT interval and polymorphic ventricular tachycardia, with twisting of the QRS complexes. Based on pre-clinical and clinical evidence, it appears that loratadine, cetirizine, and fexofenadine are safe from cardiac arrhythmia via the IKr channel, whereas astemizole and terfenadine have a propensity to cause ventricular tachyarrhythmias.
Assuntos
Arritmias Cardíacas/induzido quimicamente , Antagonistas dos Receptores Histamínicos H1/efeitos adversos , Animais , Eletrocardiografia , Feminino , Antagonistas dos Receptores Histamínicos H1/farmacologia , Humanos , Masculino , Canais de Potássio/efeitos dos fármacos , Fatores de Risco , Fatores Sexuais , Torsades de Pointes/induzido quimicamenteRESUMO
BACKGROUND: Caffeine acts mainly via blockade of adenosine receptors, which have been classified into A1, A2A, A2B, and A3 subtypes. We determined whether repeated caffeine administration (750 mg/d for 1 week) upregulates the human platelet A2A adenosine receptor and is accompanied by sensitization of platelet responses (increase in cAMP accumulation and decrease in platelet aggregation) to selective stimulation of the A2A receptors. METHODS AND RESULTS: Platelets were obtained from peripheral venous blood of 9 human volunteers at the end of 1 week of caffeine abstinence (control) and at 12 and 60 hours after the last dose of caffeine (withdrawal). The A2A receptor radioligand [3H]SCH 58261 (5-amino-7(phenylethyl)-2-(2-furyl)-pyrazolo[4,3-e]-1,2,4-triazolo[1, 5-c]-pyrimidine) bound to a single affinity class of sites in platelet membranes from controls with a Bmax of 98+/-2 fmol/mg protein and a KD of 1.29+/-0.05 nmol/L. At 12 and 60 hours after caffeine withdrawal, the radioligand bound with similar affinity (KD=1.36+/-0.06 and 1.21+/-0.05 nmol/L, respectively), but the Bmax was increased (P<0.01) to 128+/-3 and 132+/-2 fmol/mg protein. The A2A receptor agonist 2-hexynyl-5'-N-ethylcarboxamidoadenosine (HE-NECA) increased cAMP accumulation (EC50=59+/-3 nmol/L) and inhibited (IC50=90+/-6 nmol/L) aggregation of control platelets. The EC50 values for HE-NECA to increase cAMP accumulation of platelets were reduced (P<0.01) at 12 and 60 hours after caffeine withdrawal (31+/-3 and 21+/-2 nmol/L, respectively). The IC50 values for HE-NECA to inhibit ADP-induced platelet aggregation were 50+/-5 and 30+/-2 nmol/L at 12 and 60 hours after caffeine withdrawal, respectively. CONCLUSIONS: Chronic caffeine intake leads to upregulation of A2A receptors and is accompanied by sensitization to the actions of the agonist HE-NECA.