RESUMEN
Human immunodeficiency viruses (HIV) have a high level of genetic diversity. The outlier variants of HIV type 1 (HIV-1) group O are distantly related to HIV-1 group M. Their divergence has an impact on serological diagnosis, with a risk of false-negative results. In this study, we report 20 failure cases, involving patients with primary or chronic infection, in France and Cameroon between 2001 and 2008. Our results indicate that some assays detected group O infection much less efficiently than others. Two major reasons for these false-negative results were identified: the presence or absence of a group O-specific antigen (and the designed sequence) for the detection of antibodies and the greater envelope variability of group O than of group M strains. This study highlights the complexity of screening for these divergent variants and the need to evaluate test performance with a large panel of strains, due to the extensive diversity of group O variants.
Asunto(s)
Reacciones Falso Negativas , Anticuerpos Anti-VIH/sangre , Antígenos VIH/sangre , Infecciones por VIH/diagnóstico , VIH-1/clasificación , VIH-1/inmunología , Pruebas Serológicas/métodos , Camerún , Femenino , Francia , Variación Genética , Humanos , MasculinoRESUMEN
Hemophilia B is a congenital recessive disorder caused by deficiency of coagulation factor IX (FIX). Surgical procedures can be performed in patients with hemophilia using high-purity and/or recombinant FIX, which has been shown to be safe and effective in surgical hemostasis. Liver transplantation is the only potentially curative treatment available for these patients, providing a long-term phenotypic cure for hemophilia. End-stage liver disease together with hemophilia exposes patients to greater risks of bleeding complications during the perioperative period with consequent difficulties in managing coagulopathy. The limited experiences reported by different investigators and the various strategies for clotting factor replacement make it difficult to define a single approach with respect to the optimal dose and method of administering FIX to achieve perioperative hemostasis. The limits of plasma-based coagulation tests--prothrombin time, activated partial thromboplastin time--have made thromboelastography a valid alternative in this kind of surgery. It has been demonstrated to be a useful tool for real-time analysis of clot formation using a whole-blood assay format. Further, it accurately illustrates the clinical effects of procoagulant or anticoagulant interventions. In this article, we have described the usefulness of thromboelastography to monitor the ability of high-purity FIX supplementation to restore a normal coagulation state and to guide the perioperative administration of blood products in a successful orthotopic liver transplantation in a hemophilic patient with deficiencies of factors IX and X, presenting with hepatitis C virus-related cirrhosis and hepatocellular carcinoma.
Asunto(s)
Factor IX/uso terapéutico , Hemofilia B/cirugía , Trasplante de Hígado/métodos , Tromboelastografía , Hemofilia B/complicaciones , Hepatitis C/complicaciones , Hepatitis C/cirugía , Humanos , Fallo Hepático/cirugía , Fallo Hepático/virología , Masculino , Persona de Mediana Edad , Proteínas Recombinantes/uso terapéuticoRESUMEN
Antipsychotic drugs are widely used in people with dementia to treat neuropsychiatic symptoms such as aggression, agitation and psychosis. Using antipsychotic agents in older patients is difficult, because it depends on co-morbid conditions, side effects, dosing strategies, duration of treatments and combinations of various medications. This paper discusses the use of atypical antipsychotics in a 1-year-observation on a group of patients followed by an expert dementia center.
Asunto(s)
Antipsicóticos/uso terapéutico , Trastornos Psicóticos/tratamiento farmacológico , Trastornos Psicóticos/epidemiología , Anciano , Anciano de 80 o más Años , Antipsicóticos/efectos adversos , Enfermedades de los Ganglios Basales/inducido químicamente , Trastornos Cerebrovasculares/inducido químicamente , Trastornos del Conocimiento/inducido químicamente , Femenino , Humanos , Masculino , PolifarmaciaRESUMEN
Target and lead discovery constitute the main components of today's early pharmaceutical research. The aim of target discovery is the identification and validation of suitable drug targets for therapeutic intervention, whereas lead discovery identifies novel chemical molecules that act on those targets. With the near completion of the human genome sequencing, bioinformatics has established itself as an essential tool in target discovery and the in silico analysis of gene expression and gene function are now an integral part of it, facilitating the selection of the most relevant targets for a disease under study. In lead discovery, advances in chemoinformatics have led to the design of compound libraries in silico that can be screened virtually. Moreover, computational methods are being developed to predict the drug-likeness of compounds. Thus, drug discovery is already on the road towards electronic R&D.
Asunto(s)
Biología Computacional , Diseño de Fármacos , Perfilación de la Expresión Génica , Genoma Humano , Humanos , Biblioteca de PéptidosRESUMEN
REV-ERBα and REV-ERBß nuclear receptors regulate several physiological processes, including circadian rhythm and metabolism. A previous study reported the REV-ERBα gene to be co-overexpressed with ERBB2 in breast cancer cell lines. Surprisingly, we found that several tumor types, including a number of breast cancer cell lines, predominantly express the REV-ERBß variant. This pattern was independent of ERBB2 and ER status, and opposite to that of non-cancer mammary epithelial HMEC cells, in which REV-ERBα was the major variant. Consistent with this molecular profile, REV-ERB target genes in both circadian and metabolic pathways were derepressed upon silencing of REV-ERBß, but not REV-ERBα. Strikingly, we found that REV-ERBß is a determinant of sensitivity to chloroquine, a clinically relevant lysosomotropic agent that suppresses autophagy. The cytoprotective function of REV-ERBß appears to operate downstream of autophagy blockade. Through compound screening, we identified ARN5187, a novel lysosomotropic REV-ERBß ligand with a dual inhibitory activity toward REV-ERB-mediated transcriptional regulation and autophagy. Remarkably, although ARN5187 and chloroquine share similar lysosomotropic potency and have a similar effect on autophagy inhibition, ARN5187 is significantly more cytotoxic. Collectively, our results reveal that dual inhibition of REV-ERBß and autophagy is an effective strategy for eliciting cytotoxicity in cancer cells. Furthermore, our discovery of a novel inhibitor compound of both REV-ERB and autophagy may provide a scaffold for the discovery of new multifunctional anticancer agents.
Asunto(s)
Antineoplásicos/farmacología , Autofagia/efectos de los fármacos , Citotoxinas/farmacología , Neoplasias/tratamiento farmacológico , Receptores Citoplasmáticos y Nucleares/antagonistas & inhibidores , Proteínas Represoras/antagonistas & inhibidores , Autofagia/genética , Ensayos de Selección de Medicamentos Antitumorales , Células HEK293 , Células Hep G2 , Humanos , Neoplasias/genética , Neoplasias/metabolismo , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/genética , Miembro 1 del Grupo D de la Subfamilia 1 de Receptores Nucleares/metabolismo , Receptores Citoplasmáticos y Nucleares/genética , Receptores Citoplasmáticos y Nucleares/metabolismo , Proteínas Represoras/genética , Proteínas Represoras/metabolismoRESUMEN
The effects of the metabotropic glutamate receptor antagonist (+)-alpha-methyl-4-carboxyphenylglycine (MCPG) on performance in a water maze and in context-specific associative learning were examined in rats previously implanted with cannulae. MCPG (20.8 micrograms) injected intraventricularly (i.c.v.) before testing impaired the performance of rats in the spatial version of the Morris water maze, but 1/10 of this dose did not. Memory retention, evaluated 24 hr post-training, was also affected by the high dose of MCPG. However, performance in a cued version of the water maze was not impaired by the high dose, excluding effects of the drug on perceptual faculties. The effects of the MCPG were further characterized on performance in another hippocampus-dependent spatial learning task, the context-dependent fear conditioning task. MCPG (20.8 micrograms, i.c.v.) did not interfere with conditioned freezing to context in this task. For comparison, a group of rats was injected with the NMDA receptor blocker MK801. MK801 at a dose that disrupted the performance in the spatial version of the Morris water maze (0.08 mg/kg), significantly reduced freezing compared to controls. These experiments indicate that MCPG-sensitive metabotropic receptors may be required for only a restricted subset of spatial learning tasks, while NMDA receptors may play an integral role in all spatial learning.
Asunto(s)
Benzoatos/farmacología , Antagonistas de Aminoácidos Excitadores/farmacología , Glicina/análogos & derivados , Aprendizaje por Laberinto/efectos de los fármacos , Receptores de Glutamato Metabotrópico/antagonistas & inhibidores , Animales , Condicionamiento Psicológico/efectos de los fármacos , Señales (Psicología) , Maleato de Dizocilpina/farmacología , Relación Dosis-Respuesta a Droga , Miedo/efectos de los fármacos , Glicina/farmacología , Inyecciones Intraventriculares , Masculino , Ratas , Percepción Espacial/efectos de los fármacosRESUMEN
The effect of the inhibition of aminopeptidase and enkephalinase A on the pain threshold of mice and rats was investigated, using bestatin and thiorphan as selective peptidase inhibitors. The results indicate that both enzymes are relevant to the catabolism of enkephalins in vivo; however, their simultaneous activation requires particular conditions. These conclusions are based on the following observations: (1) Only concomitant intracerebral treatment with both inhibitors led to an increase in the threshold of animal pain, whereas, in the presence of exogenous peptides, the concomitant injection of both inhibitors in mice elicited an analgesic response greater than the sum of the effects of each single inhibitor. (2) This response could be seen only after acute trauma; in fact, when the drugs were injected through a plastic cannula, only enkephalinase A inhibition was effective in increasing analgesia induced by exogenous peptides.
Asunto(s)
Aminopeptidasas/metabolismo , Endopeptidasas/metabolismo , Dolor/fisiopatología , Aminopeptidasas/antagonistas & inhibidores , Animales , Encefalina Leucina/farmacología , Leucina/análogos & derivados , Leucina/farmacología , Masculino , Ratones , Neprilisina , Inhibidores de Proteasas , Ratas , Ratas Endogámicas , Umbral Sensorial/efectos de los fármacos , Umbral Sensorial/fisiología , Especificidad de la Especie , Tiorfan , Tiopronina/análogos & derivados , Tiopronina/farmacologíaRESUMEN
The synthesis and preliminary biological evaluation of novel (E)-3-(2-(N-phenylcarbamoyl)-vinyl)pyrrole-2-carboxylic acids bearing alkyl, acyl, alkoxy, phenyl, and halo substituents at the 4- and 5-positions of the pyrrole ring are reported. These compounds were studied for their in vitro affinity at the strychnine-insensitive glycine-binding site of the N-methyl-D-aspartate (NMDA) receptor complex. In the [3H]glycine binding assay (E)-4,5-dibromo-3-(2-(N-phenylcarbamoyl)vinyl)pyrrole-2-carboxylic acid 6w (pKi = 7.95 +/- 0.01) and the 4-bromo-5-methyl 6j (pKi = 7.24 +/- 0.01) and 4,5-dimethyl 6g (pKi = 6.70 +/- 0.03) analogues were the most active compounds of the series. Qualitative structure-activity analysis points to a negative correlation between bulk of the C-4 and C-5 substituents and affinity which is enhanced by halo-substituents. QSAR analysis by the Hansch descriptors F, R, pi, and MR, on a subset of compounds with pKi > or = 4, indicates that electron-withdrawing groups at C-4 and C-5 enhance the affinity. Bulk and lipophilicity are also relevant for the substituents at these positions. 6g was found to be a full antagonist (alpha = 0; enhancement of the [3H]TCP binding). The in vivo potency of 6g, 6j, and 6w was evaluated by the inhibition of NMDA-induced convulsions in mice by both the i.v. and po routes; 6w was the most active compound (ED50 = 3 x 10(-3) (0.8-10) g/kg, i.v. and 30 x 10(-3) (4.5-61) g/kg, p.o.). The results of this study indicate that the 3,4-disubstitutedpyrrole-2-carboxylate represents a novel template for the design of new glycine antagonists.
Asunto(s)
Acrilamidas , Anticonvulsivantes , Glicinérgicos , Pirroles , Receptores de Glicina/antagonistas & inhibidores , Receptores de N-Metil-D-Aspartato/efectos de los fármacos , Acrilamidas/síntesis química , Acrilamidas/química , Acrilamidas/farmacología , Animales , Anticonvulsivantes/síntesis química , Anticonvulsivantes/química , Anticonvulsivantes/farmacología , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/metabolismo , Corteza Cerebral/ultraestructura , Glicinérgicos/síntesis química , Glicinérgicos/química , Glicinérgicos/farmacología , Indoles/química , Indoles/farmacología , Ratones , Modelos Moleculares , N-Metilaspartato/toxicidad , Pirroles/síntesis química , Pirroles/química , Pirroles/farmacología , Ratas , Receptores de N-Metil-D-Aspartato/metabolismo , Convulsiones/inducido químicamente , Relación Estructura-Actividad , Sinapsis/efectos de los fármacos , Sinapsis/metabolismoRESUMEN
A series of analogues of the indole-2-carboxylate GV150526, currently in clinical trials as a potential neuroprotective agent for the control of the cerebral damage after stroke onset, was designed based on previous studies dealing with the electronic features of the north-east region of the glycine binding site associated with the NMDA receptor. In particular, the substitution of the para position of the terminal phenyl ring of GV150526 with suitable hydrophilic groups resulted in the identification of a new class of glycine antagonists. These compounds exhibited nanomolar in vitro affinity to the glycine binding site, high receptor selectivity, and outstanding in vivo potency. In particular, 3-[(E)-2-[(4-ureidomethylphenyl)aminocarbonyl]ethenyl]-4, 6-dichloroindole-2-carboxylic acid was found to be highly effective in the middle cerebral artery occlusion (MCAo) model in the rat, an animal model of focal ischemia, when given both prior to and after the occlusion of the middle cerebral artery. Notably, a significant neuroprotective effect was seen in this model postischaemia, when the administration of this compound was delayed up to 6 h from the occlusion of the middle cerebral artery, further confirming the wide therapeutic window seen for GV150526A.
Asunto(s)
Isquemia Encefálica/tratamiento farmacológico , Glicinérgicos/síntesis química , Indoles/química , Indoles/síntesis química , Fármacos Neuroprotectores/síntesis química , Receptores de Glicina/antagonistas & inhibidores , Urea/análogos & derivados , Animales , Anticonvulsivantes/síntesis química , Anticonvulsivantes/farmacología , Sitios de Unión/efectos de los fármacos , Ácidos Carboxílicos , Infarto Cerebral/tratamiento farmacológico , Modelos Animales de Enfermedad , Glicina/metabolismo , Glicinérgicos/farmacología , Indoles/farmacología , Ratones , Estructura Molecular , Fármacos Neuroprotectores/farmacología , Unión Proteica , Ratas , Ratas Sprague-Dawley , Receptores de N-Metil-D-Aspartato/metabolismo , Urea/síntesis química , Urea/farmacologíaRESUMEN
A series of 5-phenyl-3-ureidobenzodiazepine-2,4-diones was synthesized and evaluated as cholecystokinin-B (CCK-B) receptor antagonists. Structure-activity relationship (SAR) studies revealed the importance of the N-1 substituent for potent and selective CCK-B affinity. Addition of substituents at the urea side chain provided in some cases more potent compounds. Moreover the introduction of bulky substituents such as adamantylmethyl at N-1 and resolution of the racemic ureas resulted in our lead compound GV150013.
Asunto(s)
Ansiolíticos/síntesis química , Benzodiazepinas/síntesis química , Receptores de Colecistoquinina/antagonistas & inhibidores , Animales , Ansiolíticos/química , Ansiolíticos/farmacología , Benzodiazepinas/química , Benzodiazepinas/farmacología , Callithrix , Corteza Cerebral/metabolismo , Cristalografía por Rayos X , Cobayas , Células HeLa , Humanos , Técnicas In Vitro , Membranas , Ratones , Modelos Moleculares , Páncreas/metabolismo , Ensayo de Unión Radioligante , Ratas , Receptor de Colecistoquinina A , Receptor de Colecistoquinina B , Receptores de Colecistoquinina/metabolismo , Estereoisomerismo , Relación Estructura-ActividadRESUMEN
A series of indole-2-carboxylates bearing suitable chains at the C-3 position of the indole nucleus was synthesized and evaluated in terms of in vitro affinity using [3H]glycine binding assay and in vivo potency by inhibition of convulsions induced by N-methyl-D-aspartate (NMDA) in mice. 3-[2-[(Phenylamino)carbonyl]ethenyl]-4,6-dichloroindole-2-carboxyl ic acid (8) was an antagonist at the strychnine-insensitive glycine binding site (noncompetitive inhibition of the binding of [3H]TCP, pA2 = 8.1) displaying nanomolar affinity for the glycine binding site (pKi = 8.5), coupled with high glutamate receptor selectivity (> 1000-fold relative to the affinity at the NMDA, AMPA, and kainate binding sites). This indole derivative inhibited convulsions induced by NMDA in mice, when administered by both iv and po routes (ED50 = 0.06 and 6 mg/kg, respectively). The effect of the substituents on the terminal phenyl ring of the C-3 side chain was investigated. QSAR analysis suggested that the pKi value decreases with lipophilicity and steric bulk of substituents and increases with the electron donor resonance effect of the groups present in the para position of the terminal phenyl ring. According to these results the terminal phenyl ring of the C-3 side chain should lie in a nonhydrophobic pocket of limited size, refining the proposed pharmacophore model of the glycine binding site associated with the NMDA receptor.
Asunto(s)
Antagonistas de Aminoácidos Excitadores/farmacología , Glicinérgicos/farmacología , Glicina/antagonistas & inhibidores , Indoles/farmacología , Animales , Sitios de Unión , Unión Competitiva , Ácidos Carboxílicos , Corteza Cerebral/efectos de los fármacos , Antagonistas de Aminoácidos Excitadores/síntesis química , Antagonistas de Aminoácidos Excitadores/química , Antagonistas de Aminoácidos Excitadores/metabolismo , Glicina/metabolismo , Glicinérgicos/síntesis química , Glicinérgicos/química , Glicinérgicos/metabolismo , Indoles/síntesis química , Indoles/química , Indoles/metabolismo , Espectroscopía de Resonancia Magnética , Ratones , Estructura Molecular , N-Metilaspartato/farmacología , Ratas , Receptores de Glutamato/metabolismo , Relación Estructura-Actividad , Estricnina/farmacologíaRESUMEN
A cohort mortality study of 5000 vinyl chloride manufacturers is ongoing in 9 Italian plants. They represent the entire workforce of those ever employed in the production of the monomer and its polymerization. The objectives of the study are to investigate the mortality of the exposed population and to clear up the carcinogenic spectrum of vinyl chloride. This article gives the results for 3 out of 9 plants, Rosignano, Ferrara and Ravenna, which represent about 25% of the total cohort. The expected deaths have been calculated using the mortality rates of the Italian population. For the deceased persons information from the death certificates were used in the analysis of mortality; additional clinical and pathological data were collected (best pathological evidence, b.p.e.). In Ferrara a statistically significant excess for all malignant tumors and lung cancer was detected. In Rosignano and Ravenna the number of observed deaths were small and therefore no comments can be made on cancer mortality. The cohort study is ongoing in the 6 remaining cohorts and the future analysis will consider duration and level of exposure and latency.
Asunto(s)
Neoplasias/inducido químicamente , Enfermedades Profesionales/inducido químicamente , Cloruro de Vinilo/efectos adversos , Compuestos de Vinilo/efectos adversos , Adulto , Métodos Epidemiológicos , Humanos , Italia , Masculino , Neoplasias/mortalidad , Enfermedades Profesionales/mortalidad , Estadística como AsuntoRESUMEN
In the brain, cholecystokinin (CCK) has been described to act as a central neurotransmitter or neuromodulator involved in functions such as food consumption, stress and anxiety. Recently, the CCK system has been involved in drug dependence phenomena and proposed to be correlated to a putative state of 'drug preferring' phenotype within free choice tests. CCK exerts its action in the CNS through at least two different G-protein coupled high affinity receptors, CCK1 and CCK2. Various selective CCK receptor agonists and antagonists have been synthesised. In particular, L-364,718 has been demonstrated to be a potent and selective CCK1 receptor antagonist, whereas L-365,260 is a potent and selective CCK2 receptor antagonist. More recently, GV150013 has been reported to be a highly selective CCK2 receptor antagonist. This paper reviews the putative role of the CCK system within drug dependence phenomena. In particular, it analyses the relationship between central CCK activity and the exhibition of spontaneous preference for drugs of abuse, such as cocaine or alcohol. The potential therapeutic role for CCK receptor antagonists is also discussed.
Asunto(s)
Conducta Adictiva/metabolismo , Colecistoquinina/metabolismo , Trastornos Relacionados con Cocaína/metabolismo , Núcleo Accumbens/metabolismo , Receptores de Colecistoquinina/antagonistas & inhibidores , Animales , Conducta Adictiva/tratamiento farmacológico , Colecistoquinina/química , Trastornos Relacionados con Cocaína/tratamiento farmacológico , Humanos , Fenotipo , Receptores de Colecistoquinina/uso terapéuticoRESUMEN
Intracerebroventricular (ICV) injection of N-methyl-D-aspartate (NMDA) was shown to induce generalized seizures in mice. The competitive NMDA antagonists DL-2-amino-5-phosphonovaleroate (DL-AP7) and 3-(2-carboxypiperazin-4-yl)propyl-1-phosphonate (CPP), the NMDA "channel blocker" antagonist (+)-5-methyl-10,11-dihydro 5H-dibenzo-[a,d] cycloheptan-5,10-imine maleate (MK-801) and the strychnine-insensitive glycine antagonists kynurenic acid (KYNA) and 7-chloro-kynurenic acid (7-Cl-KYNA), when co-administered (ICV) with NMDA, antagonized NMDA-induced generalized seizures. Administration (ICV) of DL-AP7, CPP and MK-801 resulted in impared learning performance in a passive avoidance task in mice, with ED50 close to the anticonvulsant dose. The glycine antagonists KYNA and 7-Cl-KYNA at high doses significantly failed to affect performance in the same model of learning. The results indicate that compounds acting at the strychnine-insensitive glycine site may have a larger "therapeutic window" as anticonvulsants than antagonists of the NMDA receptor and channel.
Asunto(s)
Aprendizaje/efectos de los fármacos , N-Metilaspartato/farmacología , Receptores de Neurotransmisores/efectos de los fármacos , Convulsiones/inducido químicamente , Estricnina/farmacología , 2-Amino-5-fosfonovalerato/análogos & derivados , 2-Amino-5-fosfonovalerato/farmacología , Animales , Maleato de Dizocilpina/farmacología , Inyecciones Intraventriculares , Ácido Quinurénico/análogos & derivados , Ácido Quinurénico/farmacología , Masculino , Ratones , N-Metilaspartato/antagonistas & inhibidores , Piperazinas/farmacología , Receptores de GlicinaRESUMEN
Acute ethanol treatment produces a significant decrease of specific 3H-GABA binding in cerebellum while no changes were detectable in other brain areas. Scatchard analysis shows a decrease in receptor affinity but not in the number of GABA binding sites. On the other hand, chronic administration of ethanol selectively increases specific 3H-GABA binding in the striatum. Kinetic analysis of these data shows that ethanol chronic administration produces a significant increase in the number of GABA binding sites. These data may be useful for the understanding of clinical pictures following acute and chronic ethanol intoxication.
Asunto(s)
Encéfalo/metabolismo , Etanol/farmacología , Ácido gamma-Aminobutírico/metabolismo , Animales , Encéfalo/efectos de los fármacos , Cinética , Masculino , Ratas , Factores de TiempoRESUMEN
The effects of ethanol have been studied in three strains of mice (DBA 2J, albino, and C57 BL/6J) having different populations of opiate receptors. Acute ethanol treatment induces a significant increase in striatal dopamine metabolism only in the mouse strains (C57 and albino) that are rich in enkephalinergic receptors upon nigrostriatal dopaminergic fibers. After chronic ethanol, the same strains develop tolerance to striatal dihydroxyphenylacetic acid increase, while the striatal dopaminergic recognition sites become supersensitive. DBA mice, which have lower numbers of enkephalinergic receptors and higher levels of enkephalins in the striatum, fail to show changes in central dopaminergic function after acute or chronic ethanol treatment. Our results indicate the importance of an interaction between ethanol and opiate receptors in determining the neurochemical and behavioral effects of ethanol.
Asunto(s)
Dopamina/fisiología , Etanol/farmacología , Receptores Opioides/efectos de los fármacos , Animales , Unión Competitiva , Cuerpo Estriado/metabolismo , Encefalina Metionina , Encefalinas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Endogámicos DBA , Ratones Endogámicos , Especificidad de la Especie , Espiperona/metabolismoRESUMEN
Glutamate-induced changes in intracellular free Ca++ concentration ([Ca++]i) were recorded in resting and electrically-stimulated primary cultures of rat cerebral cortical cells, employing the Ca++ indicator Fura 2. A brief (10 min) exposure to glutamate led to a concentration-dependent basal [Ca++]i increase, measured 30 min after glutamate removal. In order to unmask more subtle modifications in [Ca++]i movements associated with neurosecretion, the glutamate effect was also studied in electrically-stimulated cells. The application of trains (10 s) of electrical pulses (intensity 30 mA, duration 1 ms) induced frequency-related Na+- and Ca++-dependent [Ca++]i transients. A 5 min treatment with 50 microM glutamate reduced to 48% the electrically-evoked [Ca++]i transients, evaluated 30 min after glutamate challenge. The neuroprotective effect of sodium 4,6-dichloro-3-[(E)-3-(N-phenyl)propenamide]indole-2-carboxylate (GV150526A), a new indole derivative with high affinity and selectivity for the glycine site of the NMDA receptor-channel complex, was compared with that of DL-2-amino-5-phosphonopentanoic acid (AP5), ifenprodil, 7-chlorokynurenic acid and 2,3-dihydroxy-6-nitro-7-sulfamoylbenzo(f)-quinoxaline (NBQX) on glutamate-induced [Ca++]i changes in resting and electrically-stimulated cells. In both experimental conditions, GV150526A showed to be the most potent compound. Moreover, GV150526A and 7-chlorokynurenic acid were 2-3 times more active in stimulated neurons than in resting neurons, indicating a major involvement of the glycine site in the protection of the cells kept in an active state.
Asunto(s)
Calcio/metabolismo , Corteza Cerebral/metabolismo , Antagonistas de Aminoácidos Excitadores/farmacología , Ácido Glutámico/toxicidad , Indoles/farmacología , Animales , Células Cultivadas , Corteza Cerebral/efectos de los fármacos , Interacciones Farmacológicas , Estimulación Eléctrica , Piperidinas/farmacología , Quinoxalinas/farmacología , Ratas , Ratas Sprague-Dawley , Receptores de N-Metil-D-Aspartato/antagonistas & inhibidoresRESUMEN
Recently, voltammetry with carbon fibre electrodes (CFE) has been implemented for real time measurement of nitrogen monoxide (NO) indicating that it is oxidised at the potential value of nitrites, approximately +700 mV. In contrast, here we show that modified CFE can monitor NO at oxidation potentials different than that of nitrites, i.e. +550 mV. Indeed, at +550 mV a significant increase of amperometric current levels was obtained when NO but not nitrites, were added to a phosphate buffer saline solution (PBS). Differential pulse voltammetry (DPV) supports these findings as two oxidation peaks were obtained when examining air preserved NO; peak 1 at +550 mV and peak 2 at +700 mV, respectively. In contrast, only peak 2 was monitored when nitrites or a solution of NO oxidised in air was added to PBS. Biological support to these in vitro data comes from the observation that the relaxation of an adrenaline-contracted aortic ring produced via addition of NO is concomitant with peak 1 at +550 mV. The relaxation is almost completed before the appearance of peak 2 at +700 mV. Furthermore, in vivo experiments performed in the striatum of rats show that the amperometric signal monitored at +550 mV is responsive to glutamatergic stimulation or inhibition of NO synthase.
Asunto(s)
Carbono , Electrofisiología/métodos , Potenciales de la Membrana/fisiología , Microelectrodos/tendencias , Neuroquímica/métodos , Óxido Nítrico/análisis , Nitritos/análisis , 2-Amino-5-fosfonovalerato/farmacología , Animales , Aorta/efectos de los fármacos , Aorta/metabolismo , Carbono/normas , Fibra de Carbono , Electrofisiología/instrumentación , Inhibidores Enzimáticos/farmacología , Epinefrina/farmacología , Agonistas de Aminoácidos Excitadores/farmacología , Antagonistas de Aminoácidos Excitadores/farmacología , Masculino , Microelectrodos/normas , Contracción Muscular/efectos de los fármacos , Contracción Muscular/fisiología , Músculo Liso Vascular/efectos de los fármacos , Músculo Liso Vascular/metabolismo , N-Metilaspartato/farmacología , NG-Nitroarginina Metil Éster/farmacología , Neostriado/efectos de los fármacos , Neostriado/metabolismo , Neuroquímica/instrumentación , Ratas , Ratas WistarRESUMEN
The role of the metabotropic glutamate receptor 1 (mGluR1) in synaptic plasticity was investigated in vivo in the intact hippocampus of mutant mice lacking this receptor. In a previous study we showed reduced long-term potentiation (LTP) in the dentate gyrus of mGluR1 -/- mice in vivo, but not when LTP was studied in a slice preparation. A possible explanation of this difference is that dentate neurons receive more inhibitory synaptic drive in vivo than in slice preparation where many inhibitory axon collaterals are lost. We report here that another form of synaptic plasticity, paired-pulse depression of the population spike, is also abnormal in the dentate gyrus of mGluR1-deficient mice when tested in vivo. In wild-type mice, stimulation of the medial perforant path produced paired-pulse depression of inter-pulse intervals (IPIs) up to 30 ms. Mutant mGluR1, on the other hand, showed a significantly longer IPI depression, up to 50 ms. Paired-pulse depression results from the activation of inhibitory interneurons. The GABA(B) agonist baclofen, acting presynaptically on the GABA interneurons, attenuated paired-pulse depression and allowed for a normal and stable LTP in mGluR1 mutant mice. These findings suggest an indirect role for mGluR1 in synaptic plasticity via a regulation of GABA inhibition.
Asunto(s)
Plasticidad Neuronal/fisiología , Receptores de Glutamato Metabotrópico/genética , Potenciales de Acción/efectos de los fármacos , Potenciales de Acción/fisiología , Animales , Autorreceptores/fisiología , Baclofeno/farmacología , Giro Dentado/química , Giro Dentado/citología , Giro Dentado/fisiología , Estimulación Eléctrica , Electrofisiología , Agonistas del GABA/farmacología , Agonistas de Receptores GABA-B , Interneuronas/química , Interneuronas/efectos de los fármacos , Interneuronas/fisiología , Potenciación a Largo Plazo/fisiología , Masculino , Ratones , Ratones Noqueados , Mutagénesis/fisiología , Inhibición Neural/fisiología , Receptores de Glutamato Metabotrópico/metabolismoRESUMEN
The middle cerebral artery occlusion (MCAO) in mice induces a focal cerebral ischaemia at the level of the tempo-parietal cortex. Histological staining and immunohistochemical markers were used to characterize the temporal progression of the cerebral infarct: both qualitative and quantitative analyses were performed at different days after the MCAO. At 3 days after MCAO, an extensive necrosis of the cerebral parenchyma was accompanied by extravasation and by massive oedema. After 7 days, GFAP marker showed a gliotic reaction with alteration of the astrocytes membrane permeability (S100 marker). Positivity for acid phosphatase staining indicated the presence of macrophages. At Day 14 and 21 following MCAO, the histological profile was essentially similar. Interestingly, at Day 7, 14 and 21, a previously unreported gliosis was observed in the subthalamic area. Quantitative analysis showed a significantly large infarct volume at Day 3 (7.88 +/- 1.95 mm3 +/- S.E.M.) compared to Day 7 (4.28 +/- 0.47 mm3 +/- S.E.M.). At Day 14 and Day 21 the infarct volumes were further decreased to 2.00 +/- 0.52 and 1.43 +/- 0.39 mm3 +/- S.E.M., respectively. These results suggest that it is important to consider the time of evaluation of cerebral ischaemia-induced cerebral infarct, especially in studies which aim to evaluate the neuroprotective effect of putative therapeutic agents.