RESUMO
RATIONALE: The behavioural effects elicited by chemical constituents of Cannabis sativa, such as cannabidiol (CBD), on the ventromedial hypothalamus (VMH) are not well understood. There is evidence that VMH neurons play a relevant role in the modulation of unconditioned fear-related defensive behavioural reactions displayed by laboratory animals. OBJECTIVES: This study was designed to explore the specific pattern of distribution of the CB1 receptors in the VMH and to investigate the role played by this cannabinoid receptor in the effect of CBD on the control of defensive behaviours and unconditioned fear-induced antinociception. METHODS: A panic attack-like state was triggered in Wistar rats by intra-VMH microinjections of N-methyl-D-aspartate (NMDA). One of three different doses of CBD was microinjected into the VMH prior to local administration of NMDA. In addition, the most effective dose of CBD was used after pre-treatment with the CB1 receptor selective antagonist AM251, followed by NMDA microinjections in the VMH. RESULTS: The morphological procedures demonstrated distribution of labelled CB1 receptors on neuronal perikarya situated in dorsomedial, central and ventrolateral divisions of the VMH. The neuropharmacological approaches showed that both panic attack-like behaviours and unconditioned fear-induced antinociception decreased after intra-hypothalamic microinjections of CBD at the highest dose (100 nmol). These effects, however, were blocked by the administration of the CB1 receptor antagonist AM251 (100 pmol) in the VMH. CONCLUSION: These findings suggest that CBD causes panicolytic-like effects and reduces unconditioned fear-induced antinociception when administered in the VMH, and these effects are mediated by the CB1 receptor-endocannabinoid signalling mechanism in VMH.
Assuntos
Canabidiol/toxicidade , Medo/fisiologia , Medição da Dor/métodos , Transtorno de Pânico/metabolismo , Receptor CB1 de Canabinoide/metabolismo , Núcleo Hipotalâmico Ventromedial/metabolismo , Animais , Canabidiol/administração & dosagem , Medo/efeitos dos fármacos , Medo/psicologia , Injeções Intraventriculares , Masculino , N-Metilaspartato/administração & dosagem , Medição da Dor/efeitos dos fármacos , Medição da Dor/psicologia , Transtorno de Pânico/induzido quimicamente , Piperidinas/administração & dosagem , Pirazóis/administração & dosagem , Ratos , Ratos Wistar , Receptor CB1 de Canabinoide/antagonistas & inibidores , Núcleo Hipotalâmico Ventromedial/efeitos dos fármacosRESUMO
The severity score of quinolinic acid (QA)-induced seizures was investigated after N-methyl-D-aspartate (NMDA) preconditioning associated with adenosine receptors. Also, the levels of adenosine A1 and A2A receptors and subunits of NMDA receptors in the hippocampi of mice were determined to define components of the resistance mechanism. Adult CF-1 mice were treated intraperitoneally with saline or NMDA (75 mg/kg), and some mice were treated intracerebroventricularly (i.c.v.) with 0.1 pmol of adenosine receptor antagonists 8-cyclopentyltheophylline (CPT; receptor A1) or ZM241385 (receptor A2A) 0, 1, or 6 h after NMDA administration. These adenosine receptor antagonists were administered to block NMDA's protective effect. Seizures and their severity scores were evaluated during convulsions induced by QA (36.8 nmol) that was administered i.c.v. 24 h after NMDA. The cell viability and content of subunits of the NMDA receptors were analyzed 24 h after QA administration. NMDA preconditioning reduced the maximal severity 6 displayed in QA-administered mice, inducing protection in 47.6% of mice after QA-induced seizures. CPT increased the latency of seizures when administered 0 or 6 h, and ZM241385 generated the same effect when administered 6 h after NMDA administration. The GluN1 content was lower in the hippocampi of the QA mice and the NMDA-preconditioned animals without seizures. GluN2A content was unaltered in all groups. The results demonstrated the components of resistance evoked by NMDA, in which adenosine receptors participate in a time-dependent mode. Similarly, the reduction on GluN1 expression in the hippocampus may contribute to this effect during the preconditioning period.
Assuntos
Anticonvulsivantes/uso terapêutico , N-Metilaspartato/uso terapêutico , Fármacos Neuroprotetores/uso terapêutico , Receptores de N-Metil-D-Aspartato/metabolismo , Receptores Purinérgicos P1/metabolismo , Convulsões/tratamento farmacológico , Animais , Anticonvulsivantes/administração & dosagem , Anticonvulsivantes/farmacologia , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Injeções Intraperitoneais , Masculino , Camundongos , N-Metilaspartato/administração & dosagem , N-Metilaspartato/farmacologia , Fármacos Neuroprotetores/administração & dosagem , Fármacos Neuroprotetores/farmacologia , Ácido Quinolínico/toxicidade , Convulsões/etiologiaRESUMO
Taste memory recognition is crucial for species survival; thus, the acquisition of conditioned taste aversion (CTA) protects animals against consuming poisons or toxins. In nature, food and poison are confined in the same edible item; however, in the laboratory these food constituents are usually presented separately for experimental analysis. The taste, or conditioned stimulus (CS), can be hours apart from the gastric malaise, or unconditioned stimulus (US); this extended inter-stimulus interval (ISI) allows the analysis of a particular learning phase. Evidence indicates a relevant function of glutamatergic activity in the insular cortex (IC) throughout the ISI. N-methyl-D-aspartate receptors (NMDAR) are crucial during CTA acquisition and retrieval. However, the exact participation of NMDAR in the IC during the ISI has not been demonstrated. Thus, the aim of this work was to evaluate the effects of temporal NMDAR activation during four time frames throughout the ISI of conditioned sugar aversion with bilateral injections of NMDA at a physiological dose (1⯵g/µl) in the IC, given (1) immediately before or (2) immediately after sugar presentation, or (3) immediately before or (4) immediately after LiCl i.p. injection. The results showed that NMDAR activation in the IC had a specific ISI effect during CTA acquisition, increasing aversive memory formation and delaying extinction only after CS presentation. Overall, these results demonstrate that NMDAR in the IC have a particular enhancing associative effect after CS and suggest that there is a precise coincidence in neurochemical events in the IC that correlates with the stimulus to be associated and the glutamate NMDAR activity that must be finely tuned in the ISI during CTA acquisition.
Assuntos
Aprendizagem da Esquiva/fisiologia , Córtex Cerebral/fisiologia , Condicionamento Clássico/fisiologia , Receptores de N-Metil-D-Aspartato/fisiologia , Percepção Gustatória/fisiologia , Animais , Aprendizagem da Esquiva/efeitos dos fármacos , Córtex Cerebral/efeitos dos fármacos , Condicionamento Clássico/efeitos dos fármacos , Agonistas de Aminoácidos Excitatórios/administração & dosagem , Extinção Psicológica/efeitos dos fármacos , Extinção Psicológica/fisiologia , Masculino , N-Metilaspartato/administração & dosagem , Ratos Wistar , Receptores de N-Metil-D-Aspartato/agonistas , Fatores de TempoRESUMO
N-methyl D-aspartate (NMDA) preconditioning is evoked by the administration of a subtoxic dose of NMDA and is protective against neuronal excitotoxicity. This effect may involve a diversity of targets and cell signaling cascades associated to neuroprotection. Phosphatidylinositol-3 kinase/protein kinase B (PI3K/Akt) and mitogen-activated protein kinases (MAPKs) such as extracellular regulated protein kinase 1/2 (ERK1/2) and p38MAPK pathways play a major role in neuroprotective mechanisms. However, their involvement in NMDA preconditioning was not yet fully investigated. The present study aimed to evaluate the effect of NMDA preconditioning on PI3K/Akt, ERK1/2, and p38MAPK pathways in the hippocampus of mice and characterize the involvement of PI3K on NMDA preconditioning-evoked prevention of seizures and hippocampal cell damage induced by quinolinic acid (QA). Thus, mice received wortmannin (a PI3K inhibitor) and 15 min later a subconvulsant dose of NMDA (preconditioning) or saline. After 24 h of this treatment, an intracerebroventricular QA infusion was administered. Phosphorylation levels and total content of Akt, glycogen synthase protein kinase-3ß (GSK-3ß), ERK1/2, and p38MAPK were not altered after 24 h of NMDA preconditioning with or without wortmmanin pretreatment. Moreover, after QA administration, behavioral seizures, hippocampal neuronal degeneration, and Akt activation were evaluated. Inhibition of PI3K pathway was effective in abolishing the protective effect of NMDA preconditioning against QA-induced seizures, but did not modify neuronal protection promoted by preconditioning as evaluated by Fluoro-Jade B staining. The study confirms that PI3K participates in the mechanism of protection induced by NMDA preconditioning against QA-induced seizures. Conversely, NMDA preconditioning-evoked protection against neuronal degeneration is not altered by PI3K signaling pathway inhibition. These results point to differential mechanisms regarding protection against a behavioral and cellular manifestation of neural damage.
Assuntos
Agonistas de Aminoácidos Excitatórios/administração & dosagem , Hipocampo/patologia , N-Metilaspartato/administração & dosagem , Doenças Neurodegenerativas/induzido quimicamente , Fosfatidilinositol 3-Quinase/metabolismo , Ácido Quinolínico/toxicidade , Transdução de Sinais/efeitos dos fármacos , Animais , Modelos Animais de Doenças , Esquema de Medicação , Inibidores Enzimáticos/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Masculino , Camundongos , Doenças Neurodegenerativas/patologia , Neurônios/efeitos dos fármacos , Neurônios/patologia , Fatores de TempoRESUMO
microRNA profiling has identified cell-specific expression patterns that could represent molecular signatures triggering the acquisition of a specific phenotype; in other words, of cellular identity and its associated function. Several groups have hypothesized that retinal cell phenotyping could be achieved through the determination of the global pattern of miRNA expression across specific cell types in the adult retina. This is especially relevant for Müller glia in the context of retinal damage, as these cells undergo dramatic changes of gene expression in response to injury, that render them susceptible to acquire a progenitor-like phenotype and be a source of new neurons.We describe a method that combines an experimental protocol for excitotoxic-induced retinal damage through N-methyl-D-aspartate subretinal injection with magnetic-activated cell sorting (MACS) of Müller cells and RNA isolation for microRNA profiling. Comparison of microRNA patterns of expression should allow Müller cell phenotyping under different experimental conditions.
Assuntos
Células Ependimogliais/metabolismo , Perfilação da Expressão Gênica/métodos , Separação Imunomagnética/métodos , MicroRNAs/metabolismo , Doenças Retinianas/patologia , Animais , Modelos Animais de Doenças , Células Ependimogliais/efeitos dos fármacos , Células Ependimogliais/patologia , Transportador 1 de Aminoácido Excitatório/imunologia , Perfilação da Expressão Gênica/instrumentação , Humanos , Separação Imunomagnética/instrumentação , Injeções Intravítreas , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/isolamento & purificação , N-Metilaspartato/administração & dosagem , N-Metilaspartato/toxicidade , Reação em Cadeia da Polimerase em Tempo Real/instrumentação , Reação em Cadeia da Polimerase em Tempo Real/métodos , Doenças Retinianas/induzido quimicamente , Reação em Cadeia da Polimerase Via Transcriptase Reversa/instrumentaçãoRESUMO
Stimulation-evoked antinociception (SEA) from the anterior pretectal nucleus (APtN) activates mechanisms that descend to the spinal cord through the dorsolateral funiculus, but the encephalic route followed by the descending pathways from the APtN is not completely known. This study evaluated the changes in the SEA from the APtN in the Wistar rat tail-flick test after lidocaine-induced neural block or N-methyl-d-aspartate-induced neurotoxic lesion of the deep mesencephalic nucleus (DpMe), tegmental pedunculopontine nucleus (PPTg), or lateral paragigantocellular nucleus (LPGi). The SEA from the APtN was less intense after neural block of the contralateral DpMe or PPTg or the ipsilateral LPGi, but was not changed by the neural block of the ipsilateral DpMe or PPTg or the contralateral LPGi. Antinociception did not occur when APtN stimulation was carried out 5 minutes after lidocaine or 6 days after N-methyl-d-aspartate injections into the contralateral DpMe and the ipsilateral LPGi, or into the contralateral PPTg and the ipsilateral LPGi. We conclude that the SEA from the APtN activates 2 descending pain inhibitory pathways, one relaying in the ipsilateral LPGi and another relaying sequentially in the contralateral DpMe and PPTg. PERSPECTIVE: The antinociceptive effect of the APtN stimulation involves 2 descending pathways: one relaying in the ipsilateral LPGi and another descending contralaterally via relays in the DpMe and PPTg.
Assuntos
Mapeamento Encefálico , Estimulação Elétrica/métodos , Manejo da Dor , Dor , Pulvinar/fisiologia , Analgésicos/uso terapêutico , Animais , Agonistas de Aminoácidos Excitatórios/administração & dosagem , Lateralidade Funcional , Masculino , Microinjeções , N-Metilaspartato/administração & dosagem , Dor/tratamento farmacológico , Pulvinar/efeitos dos fármacos , Ratos , Ratos WistarRESUMO
Methylprednisolone sodium succinate (MPSS) has been proposed as a first-line treatment for acute spinal cord injury (SCI). Its clinical use remains, however, controversial because of the modest benefits and numerous side-effects. We investigated if MPSS could protect spinal neurons and glia using an in vitro model of the rat spinal cord that enables recording reflexes, fictive locomotion and morphological analysis of damage. With this model, a differential lesion affecting mainly either neurons or glia can be produced via kainate-evoked excitotoxicity or application of a pathological medium (lacking O2 and glucose), respectively. MPSS (6-10 µM) applied for 24 h after 1-h pathological medium protected astrocytes and oligodendrocytes especially in the ventrolateral white matter. This effect was accompanied by the return of slow, alternating oscillations (elicited by NMDA and 5-hydroxytryptamine (5-HT)) reminiscent of a sluggish fictive locomotor pattern. MPSS was, however, unable to reverse even a moderate neuronal loss and the concomitant suppression of fictive locomotion evoked by kainate (0.1 mM; 1 h). These results suggest that MPSS could, at least in part, contrast damage to spinal glia induced by a dysmetabolic state (associated to oxygen and glucose deprivation) and facilitate reactivation of spinal networks. Conversely, when even a minority of neurons was damaged by excitotoxicity, MPSS did not protect them nor did it restore network function in the current experimental model.
Assuntos
Metilprednisolona/farmacologia , Fármacos Neuroprotetores/farmacologia , Traumatismos da Medula Espinal/tratamento farmacológico , Animais , Animais Recém-Nascidos , Imuno-Histoquímica , Ácido Caínico , Vértebras Lombares , Potenciais da Membrana/efeitos dos fármacos , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , N-Metilaspartato/administração & dosagem , N-Metilaspartato/metabolismo , Vias Neurais/efeitos dos fármacos , Vias Neurais/patologia , Vias Neurais/fisiopatologia , Neuroglia/efeitos dos fármacos , Neuroglia/patologia , Neurônios/efeitos dos fármacos , Neurônios/patologia , Neurônios/fisiologia , Neurotransmissores/administração & dosagem , Ratos Wistar , Serotonina/administração & dosagem , Serotonina/metabolismo , Traumatismos da Medula Espinal/patologia , Traumatismos da Medula Espinal/fisiopatologia , Substância Branca/efeitos dos fármacos , Substância Branca/patologia , Substância Branca/fisiopatologiaRESUMO
Previously we showed that activation of the Nucleus of the Solitary Tract (NTS)-Nucleus Paragigantocellularis (PGi)-Locus coeruleus (LC) pathway, which theoretically culminates with norepinephrine (NE) release in dorsal hippocampus (CA1 region) and basolateral amygdala (BLA) is necessary for the consolidation of object recognition (OR) memory. Here we show that, while the microinjection of the beta-noradrenergic receptor blocker timolol into CA1 impairs OR memory consolidation, the microinjection of norepinephrine (NE) promotes the persistence of this type of memory. Further, we show that OR consolidation is attended by an increase of norepinephrine (NE) levels and of the expression of brain derived neurotrophic factor (BDNF) in hippocampus, which are impaired by inactivation of the NTS-PGi-LC pathway by the infusion of muscimol into the NTS.
Assuntos
Fator Neurotrófico Derivado do Encéfalo/metabolismo , Hipocampo/fisiologia , Consolidação da Memória/fisiologia , Norepinefrina/fisiologia , Reconhecimento Psicológico/fisiologia , Antagonistas Adrenérgicos beta/administração & dosagem , Animais , Agonistas de Aminoácidos Excitatórios/administração & dosagem , Comportamento Exploratório/efeitos dos fármacos , Comportamento Exploratório/fisiologia , Agonistas de Receptores de GABA-A/administração & dosagem , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Locus Cerúleo/efeitos dos fármacos , Locus Cerúleo/fisiologia , Masculino , Consolidação da Memória/efeitos dos fármacos , Muscimol/administração & dosagem , N-Metilaspartato/administração & dosagem , Norepinefrina/administração & dosagem , Ratos Wistar , Reconhecimento Psicológico/efeitos dos fármacos , Núcleo Solitário/efeitos dos fármacos , Núcleo Solitário/fisiologia , Timolol/administração & dosagemRESUMO
GABA (γ-amino butyric acid) is the major inhibitory transmitter in the central nervous system and its action is terminated by specific transporters (GAT), found in neurons and glial cells. We have previously described that GAT-3 is responsible for GABA uptake activity in cultured avian Müller cells and that it operates in a Na(+) and Cl(-) dependent manner. Here we show that glutamate decreases [(3)H] GABA uptake in purified cultured glial cells up to 50%, without causing cell death. This effect is mediated by ionotropic glutamatergic receptors. Glutamate inhibition on GABA uptake is not reverted by inhibitors of protein kinase C or modified by agents that modulate cyclic AMP/PKA. Biotinylation experiments demonstrate that this reduction in GABA uptake correlates with a decrease in GAT-3 plasma membrane levels. Interestingly, both GAT-1 and GAT-3 mRNA levels are also decreased by glutamate. Conditioned media (CM) prepared from retinal neurons could also decrease GABA influx, and glutamate receptor antagonists (MK-801 + CNQX) were able to prevent this effect. However, glutamate levels in CM were not different from those found in fresh media, indicating that a glutamatergic co-agonist or modulator could be regulating GABA uptake by Müller cells in this scenario. In the whole avian retina, GAT-3 is present from embryonic day 5 (E5) increasing up to the end of embryonic development and post-hatch period exclusively in neuronal layers. However, this pattern may change in pathological conditions, which drive GAT-3 expression in Müller cells. Our data suggest that in purified cultures and upon extensive neuronal lesion in vivo, shown as a Brn3a reduced neuronal cells and an GFAP increased gliosis, Müller glia may change its capacity to take up GABA due to GAT-3 up regulation and suggests a regulatory interplay mediated by glutamate between neurons and glial cells in this process.
Assuntos
Células Ependimogliais/fisiologia , Proteínas da Membrana Plasmática de Transporte de GABA/fisiologia , Ácido Glutâmico/fisiologia , Ácido gama-Aminobutírico/metabolismo , Animais , Transporte Biológico Ativo , Biotinilação , Cálcio/análise , Membrana Celular/metabolismo , Células Cultivadas , Embrião de Galinha , Galinhas , Meios de Cultivo Condicionados , Células Ependimogliais/efeitos dos fármacos , Proteínas da Membrana Plasmática de Transporte de GABA/genética , Perfilação da Expressão Gênica , Ácido Glutâmico/farmacologia , Ácido Caínico/farmacologia , N-Metilaspartato/administração & dosagem , N-Metilaspartato/farmacologia , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C/fisiologia , Inibidores de Proteínas Quinases/farmacologia , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Retina/crescimento & desenvolvimento , Acetato de Tetradecanoilforbol/farmacologiaRESUMO
BACKGROUND: The anterior pretectal nucleus (APtN) activates descending mechanisms of pain control. This study evaluated whether the APtN also controls neuropathic pain in rats. METHODS: The hypersensitivity to mechanical stimulation with an electronic von Frey apparatus and the number of Fos-immunoreactive (Fos-ir) neurons in the APtN were evaluated in rats before and after chronic constriction injury of the sciatic nerve. RESULTS: The tactile hypersensitivity was characterized by an initial phase (the 2 days following the injury) and a maintenance phase (the subsequent 7 days). The injection of 2% lidocaine (0.25 µL) or N-methyl-D-aspartate (2.5 µg/0.25 µL) into the APtN intensified the tactile hypersensitivity observed 2 days after injury but did not alter the tactile hypersensitivity observed 7 and 14 days after injury. The injection of naloxone (10 ng/0.25 µL) or methysergide (40 pg/0.25 µL) but not atropine (100 ng/0.25 µL) into the APtN also intensified the tactile hypersensitivity observed 2 days after the injury. A significant increase in the number of Fos-ir cells was found in the contralateral APtN 2 days but not 7 or 14 days after the injury. Electrical stimulation of the APtN reduced the tactile hypersensitivity at 2, 7 and 14 days after the nerve ligation. CONCLUSION: APtN exerts a tonic inhibitory influence on persistent pain. The results point out to an important role of opioid and serotonergic mediation into the APtN to inhibit hyperalgesia during the initial phase of neuropathic pain.
Assuntos
Vias Neurais/patologia , Neuralgia/patologia , Área Pré-Tectal/patologia , Anestésicos Locais/administração & dosagem , Anestésicos Locais/farmacologia , Animais , Constrição Patológica/complicações , Constrição Patológica/patologia , Hiperalgesia/fisiopatologia , Lidocaína/administração & dosagem , Lidocaína/farmacologia , Masculino , Metisergida/farmacologia , N-Metilaspartato/administração & dosagem , N-Metilaspartato/farmacologia , Naloxona/farmacologia , Antagonistas de Entorpecentes/farmacologia , Neurônios/patologia , Medição da Dor/efeitos dos fármacos , Estimulação Física , Proteínas Proto-Oncogênicas c-fos/metabolismo , Ratos , Ratos Wistar , Neuropatia Ciática/patologiaRESUMO
The inferior colliculus (IC), a midbrain structure that processes acoustic information of aversive nature, is distinguished from other auditory nuclei in the brainstem by its connections with structures of the motor system. Recent evidence relating the IC to motor behavior shows that glutamate-mediated mechanisms in the neural circuits at the IC level modulate haloperidol-induced catalepsy. It has been shown that N(G)-nitro-L-arginine (L-NOARG), inhibitor of enzyme nitric oxide synthase (NOS), can induce catalepsy after intraperitoneal (ip), intracerebroventricular or intrastriatal administration. The present study examined whether the catalepsy induced by L-NOARG (ip) can be influenced by collicular glutamatergic mechanisms and if a NO-dependent neural substrate into the IC plays a role in this immobility state. L-NOARG-induced catalepsy was challenged with prior intracollicular microinjections of glutamate NMDA receptor antagonists, AP7 (20 or 40 nmol/0.5 µl), or of the NMDA receptor agonist N-methyl-D-aspartate (NMDA, 30 nmol/0.5 µl). Catalepsy was evaluated by positioning both forepaws of the rats on an elevated horizontal wooden bar and recording the time for which the animal maintained this position. The results showed that intracollicular microinjection of AP7 previous to systemic injections of L-NOARG (90 mg/kg) significantly attenuated the catalepsy. Conversely, intracollicular microinjection of NMDA increased the time of catalepsy when administered 10 min before systemic L-NOARG (10 or 45 mg/kg). The microinjection of L-NOARG (50 or 100 nmol) directly into the IC was not able to induce catalepsy. These findings suggest that glutamate-mediated mechanisms in the neural circuits of the IC modulate L-NOARG-induced catalepsy and participate in the regulation of motor activity.
Assuntos
Catalepsia/induzido quimicamente , Catalepsia/patologia , Inibidores Enzimáticos/toxicidade , Colículos Inferiores/metabolismo , Nitroarginina/toxicidade , Receptores de N-Metil-D-Aspartato/metabolismo , 2-Amino-5-fosfonovalerato/administração & dosagem , 2-Amino-5-fosfonovalerato/análogos & derivados , Animais , Catalepsia/prevenção & controle , Relação Dose-Resposta a Droga , Esquema de Medicação , Interações Medicamentosas , Antagonistas de Aminoácidos Excitatórios/administração & dosagem , Colículos Inferiores/efeitos dos fármacos , Masculino , Microinjeções , N-Metilaspartato/administração & dosagem , Ratos , Ratos Wistar , Estatísticas não Paramétricas , Fatores de TempoRESUMO
UNLABELLED: We investigated the antinociceptive effects of AR-A014418, a selective inhibitor of glycogen synthase kinase-3ß (GSK-3ß) in mice. A 30-minute pretreatment with AR-A014418 (.1 and 1 mg/kg, intraperitoneal [ip]) inhibited nociception induced by an ip injection of acetic acid. AR-A014418 pretreatment (.1 and .3 mg/kg, ip) also decreased the late (inflammatory) phase of formalin-induced licking, without affecting responses of the first (neurogenic) phase. In a different set of experiments, AR-A014418 (.1-10 µg/site) coinjected intraplantarly (ipl) with formalin inhibited the late phase of formalin-induced nociception. Furthermore, AR-A014418 administration (1 and 10 ng/site, intrathecal [it]) inhibited both phases of formalin-induced licking. In addition, AR-A014418 coinjection (10 ng/site, it) inhibited nociception induced by glutamate, N-methyl-D-aspartate (NMDA), (±)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (trans-ACPD), tumor necrosis factor-alpha (TNF-α), and interleukin-1beta (IL-1ß) by 47 ± 12%, 48 ± 11%, 31 ± 8%, 46 ± 13%, and 44 ± 11%, respectively. In addition, a 30-minute pretreatment with NP031115 (3 and 10 mg/kg, ip), a different GSK-3 ß inhibitor, also attenuated the late phase of formalin-induced nociception. Collectively, these results provide convincing evidence that AR-A014418, given by local, systemic, and central routes, produces antinociception in several mouse models of nociception. The AR-A014418-dependent antinociceptive effects were induced by modulation of the glutamatergic system through metabotropic and ionotropic (NMDA) receptors and the inhibition of the cytokine (TNF-α and IL-1ß) signaling. PERSPECTIVE: These results suggest that GSK-3ß may be a novel pharmacological target for the treatment of pain.
Assuntos
Dor Abdominal/prevenção & controle , Analgésicos/administração & dosagem , Quinase 3 da Glicogênio Sintase/antagonistas & inibidores , Tiazóis/administração & dosagem , Ureia/análogos & derivados , Dor Abdominal/induzido quimicamente , Agressão/efeitos dos fármacos , Análise de Variância , Animais , Anti-Inflamatórios não Esteroides , Azidas/administração & dosagem , Citocinas/administração & dosagem , Citocinas/metabolismo , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Vias de Administração de Medicamentos , Formaldeído/efeitos adversos , Ácido Glutâmico/efeitos adversos , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Masculino , Camundongos , N-Metilaspartato/administração & dosagem , Medição da Dor/métodos , Açúcares Ácidos/efeitos adversos , Ureia/administração & dosagem , Xilose/efeitos adversos , Xilose/análogos & derivadosRESUMO
Traumatic brain injury (TBI) causes impairment of fine motor functions in humans and nonhuman mammals that often persists for months after the injury occurs. Neuroprotective strategies for prevention of the sequelae of TBI and understanding the molecular mechanisms and cellular pathways are related to the glutamatergic system. It has been suggested that cellular damage subsequent to TBI is mediated by the excitatory neurotransmitters, glutamate and aspartate, through the excessive activation of the N-methyl-D-aspartate (NMDA) receptors. Thus, preconditioning with a low dose of NMDA was used as a strategy for protection against locomotor deficits observed after TBI in mice. Male adult mice CF-1 were preconditioned with NMDA (75 mg/kg) 24 hr before the TBI induction. Under anesthesia with O(2)/N(2)O (33%: 66%) inhalation, the animals were subjected to the experimental model of trauma that occurs by the impact of a 25 g weight on the skull. Sensorimotor gating was evaluated at 1.5, 6, or 24 hr after TBI induction by using footprint and rotarod tests. Cellular damage also was assessed 24 hr after occurrence of cortical trauma. Mice preconditioned with NMDA were protected against all motor deficits revealed by footprint tests, but not those observed in rotarod tasks. Although mice showed motor deficits after TBI, no cellular damage was observed. These data corroborate the hypothesis that glutamatergic excitotoxicity, especially via NMDA receptors, contributes to severity of trauma. They also point to a putative neuroprotective mechanism induced by a sublethal dose of NMDA to improve motor behavioral deficits after TBI.
Assuntos
Lesões Encefálicas/tratamento farmacológico , Discinesias/tratamento farmacológico , N-Metilaspartato/uso terapêutico , Fármacos Neuroprotetores/uso terapêutico , Animais , Lesões Encefálicas/complicações , Lesões Encefálicas/patologia , Contagem de Células , Cerebelo/efeitos dos fármacos , Cerebelo/patologia , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/patologia , Fragmentação do DNA/efeitos dos fármacos , Modelos Animais de Doenças , Discinesias/etiologia , Discinesias/patologia , Masculino , Camundongos , Atividade Motora/efeitos dos fármacos , N-Metilaspartato/administração & dosagem , Fármacos Neuroprotetores/administração & dosagem , Equilíbrio Postural/efeitos dos fármacos , Índice de Gravidade de Doença , Fatores de Tempo , Resultado do TratamentoRESUMO
RATIONALE: The midbrain periaqueductal gray (PAG) is part of the brain system involved in active defense reactions to threatening stimuli. Glutamate N-methyl-D: -aspartate (NMDA) receptor activation within the dorsal column of the PAG (dPAG) leads to autonomic and behavioral responses characterized as the fear reaction. Nitric oxide (NO) has been proposed to be a mediator of the aversive action of glutamate, since the activation of NMDA receptors in the brain increases NO synthesis. OBJECTIVES: We investigated the effects of intra-dPAG infusions of NMDA on defensive behaviors in mice pretreated with a neuronal nitric oxide synthase (nNOS) inhibitor [Nomega-propyl-L: -arginine (NPLA)], in the same midbrain site, during a confrontation with a predator in the rat exposure test (RET). MATERIALS AND METHODS: Male Swiss mice received intra-dPAG injections of NPLA (0.1 or 0.4 nmol/0.1 microl), and 10 min later, they were infused with NMDA (0.04 nmol/0.1 microl) into the dPAG. After 10 min, each mouse was placed in the RET. RESULTS: NMDA treatment enhanced avoidance behavior from the predator and markedly increased freezing behavior. These proaversive effects of NMDA were prevented by prior injection of NPLA. Furthermore, defensive behaviors (e.g., avoidance, risk assessment, freezing) were consistently reduced by the highest dose of NPLA alone, suggesting an intrinsic effect of nitric oxide on defensive behavior in mice exposed to the RET. CONCLUSIONS: These findings suggest a potential role of glutamate NMDA receptors and NO in the dPAG in the regulation of defensive behaviors in mice during a confrontation with a predator in the RET.
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Medo/fisiologia , N-Metilaspartato/administração & dosagem , Óxido Nítrico/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Animais , Arginina/administração & dosagem , Arginina/análogos & derivados , Arginina/farmacologia , Aprendizagem da Esquiva/fisiologia , Aprendizagem da Esquiva/efeitos da radiação , Comportamento Animal/efeitos dos fármacos , Comportamento Animal/fisiologia , Relação Dose-Resposta a Droga , Medo/efeitos dos fármacos , Reação de Congelamento Cataléptica/fisiologia , Masculino , Camundongos , Óxido Nítrico Sintase Tipo I/antagonistas & inibidores , Óxido Nítrico Sintase Tipo I/metabolismo , Substância Cinzenta Periaquedutal/efeitos dos fármacos , Substância Cinzenta Periaquedutal/metabolismo , Comportamento Predatório , Ratos , Ratos Long-EvansRESUMO
Glutamate-NMDA (N-methyl-D-aspartate) receptor activation within the periaqueductal gray (PAG) leads to antinociceptive, autonomic and behavioral responses characterized as the fear reaction. We have recently demonstrated that the vigorous defensive-like behaviors (e.g. jumping and running) and antinociception induced by intra-PAG injection of N-methyl-D-aspartate (NMDA) were completely blocked by prior infusion of N(omega)-propyl-L-arginine (NPLA), a specific neuronal nitric oxide synthesis (nNOS) enzyme inhibitor, into the same midbrain structure. It remains unclear however, whether the inhibition of nNOS within the mouse PAG changes the anxiety-like behavior per se or the effects of the inhibition of nNOS depend on the suppression of downstream of glutamate-NMDA receptor activation. This study investigated whether intra-PAG infusion of NPLA (i) attenuates anxiety in the elevated plus-maze (EPM) and (ii) antagonizes the anxiogenic-like effects induced by intra-PAG injection of NMDA. Test sessions were videotaped and subsequently scored for conventional indices of anxiety (percentage of open arm entries and percentage of open arm time) and locomotor activity (closed arm entries). Results showed that intra-PAG infusions of NPLA (0.2, 0.4 or 0.8 nmol/0.1 microl) did not alter significantly any behavioral response in the EPM when compared to control group (Experiment 1). Intra-PAG infusion of NMDA (0 and 0.02 nmol/0.1 microl; a dose that does not provoke vigorous defensive behaviors per se in mice) significantly reduced open arm exploration, confirming an anxiogenic-like effect (Experiment 2). When injected into the PAG 10 min prior local NMDA injection (0.02 nmol/0.1 microl), NPLA (0.4 nmol/0.1 microl) was able to revert the anxiogenic-like effect of glutamate-NMDA receptor activation. Neither intra-PAG infusion of NMDA nor NPLA altered closed arm entries, a widely used measure of locomotor activity in the EPM. These results suggest that intra-PAG nitric oxide synthesis does not play a role on anxiety-like behavior elicited during EPM exposure; however its synthesis is important for the proaversive effects produced by activation of glutamate-NMDA receptors located within this limbic midbrain structure.
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Ansiedade/metabolismo , Óxido Nítrico Sintase Tipo I/metabolismo , Substância Cinzenta Periaquedutal/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Animais , Comportamento Animal , Inibidores Enzimáticos/administração & dosagem , Injeções Intraventriculares , Camundongos , Microinjeções , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , N-Metilaspartato/administração & dosagem , N-Metilaspartato/metabolismo , Óxido Nítrico Sintase Tipo I/efeitos dos fármacos , Substância Cinzenta Periaquedutal/efeitos dos fármacos , Receptores de N-Metil-D-Aspartato/efeitos dos fármacosRESUMO
The involvement of the dorsolateral periaqueductal gray in the regulation of fear-related behaviors such as escape and freezing is well established. It is still a matter of investigation, however, whether this midbrain area may have a relevant role in the modulation of more subtle defensive responses associated with anxiety such as risk assessment and inhibitory avoidance. By stimulating N-methyl-D-aspartic acid glutamate receptors located in the dorsolateral periaqueductal gray with its prototypical agonist N-methyl-D-aspartic acid (50 pmol), we report here an increase in both risk assessment and inhibitory avoidance behaviors of male Wistar rats tested in the elevated T-maze. These results are indicative of an anxiogenic-like effect. The selective N-methyl-D-aspartic acid receptor antagonist DL-2-amino-7-phosphonoheptanoic acid (2.0 and 4.0 nmol) had the opposite effect on both defensive tasks. Pretreatment with an ineffective dose of DL-2-amino-7-phosphonoheptanoic acid (1.0 nmol) prevented the N-methyl-D-aspartic acid anxiogenic-like effect. At the dose range of DL-2-amino-7-phosphonoheptanoic acid and/or N-methyl-D-aspartic acid tested, neither the escape response from one of the elevated T-maze open arms nor the general exploratory activity as assessed in the open-field test was affected. The present results suggest that the dorsolateral periaqueductal gray column is also involved in the regulation of defensive behaviors related to anxiety, and N-methyl-D-aspartic acid glutamate receptors are recruited for this action.
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Ansiedade/tratamento farmacológico , Ansiedade/psicologia , Medo/efeitos dos fármacos , Medo/psicologia , Substância Cinzenta Periaquedutal/fisiologia , Receptores de N-Metil-D-Aspartato/efeitos dos fármacos , Assunção de Riscos , 2-Amino-5-fosfonovalerato/administração & dosagem , 2-Amino-5-fosfonovalerato/análogos & derivados , 2-Amino-5-fosfonovalerato/farmacologia , Animais , Anticonvulsivantes/farmacologia , Etossuximida/farmacologia , Agonistas de Aminoácidos Excitatórios/administração & dosagem , Agonistas de Aminoácidos Excitatórios/farmacologia , Antagonistas de Aminoácidos Excitatórios/administração & dosagem , Antagonistas de Aminoácidos Excitatórios/farmacologia , Comportamento Exploratório/efeitos dos fármacos , Hipocampo/fisiologia , Masculino , Microinjeções , N-Metilaspartato/administração & dosagem , N-Metilaspartato/farmacologia , Ratos , Ratos WistarRESUMO
The changes in mean arterial pressure (MAP) and respiratory frequency (RF) in response to microinjection of NMDA or AMPA into the commissural subnucleus of the NTS (comNTS) at the calamus scriptorius level of awake rats were evaluated. Under tribromoethanol anesthesia, the rats received guide-cannulae in direction of the NTS and a catheter was inserted into the femoral artery for measurement of arterial pressure. Changes in RF were evaluated with the rats inside a plethysmographic chamber. Randomly microinjections of 5 doses of NMDA (0.001, 0.01, 0.1, 1.0 and 2 nmol/50 nL; n = 10) or AMPA (1, 5, 10, 25 and 50 pmol/50 nL; n = 8) into the comNTS were performed at 15 min intervals and produced a dose-dependent increase in MAP [NMDA (3 +/- 2, 4 +/- 3, 25 +/- 4, 41 +/- 4 and 51 +/- 4 mm Hg) and AMPA (0 +/- 1, 14 +/- 4, 17 +/- 3, 27 +/- 5 and 34 +/- 3 mm Hg)]. Microinjection of NMDA (1 nmol/50 nL; n = 7) or AMPA (50 pmol/50 nL; n = 4) into the comNTS produced a long lasting apnea. The pressor responses to microinjection of NMDA or AMPA into the comNTS were blocked by prazosin, a alpha(1)-adrenoceptor antagonist, indicating that the increase in arterial pressure in both cases was sympathetically mediated. The data show that microinjection of NMDA and AMPA into the comNTS produced pressor response and apnea, indicating that both ionotropic l-glutamate receptors may play a role in the neurotransmission/neuromodulation of the autonomic and respiratory components of the cardiovascular reflexes at this level.
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Pressão Sanguínea/fisiologia , N-Metilaspartato/fisiologia , Respiração , Núcleo Solitário/fisiologia , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiônico/administração & dosagem , Análise de Variância , Animais , Apneia/induzido quimicamente , Sistema Nervoso Autônomo/efeitos dos fármacos , Sistema Nervoso Autônomo/fisiologia , Pressão Sanguínea/efeitos dos fármacos , Fenômenos Fisiológicos Cardiovasculares/efeitos dos fármacos , Sistema Cardiovascular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Agonistas de Aminoácidos Excitatórios/administração & dosagem , Masculino , Microinjeções , N-Metilaspartato/administração & dosagem , Distribuição Aleatória , Ratos , Ratos Wistar , Respiração/efeitos dos fármacos , Núcleo Solitário/efeitos dos fármacosRESUMO
Freezing and escape responses induced by gradual increases in the intensity of the electrical current applied to dorsal regions of the periaqueductal gray (dPAG) cause a distinct pattern of Fos distribution in the brain. From these studies, it has been suggested that a pathway involving the dPAG itself, dorsomedial hypothalamus and the cuneiform nucleus (CnF) would mediate responses to immediate danger and another one involving the amygdala and ventrolateral periaqueductal gray (vlPAG) would mediate cue-elicited responses. As electrical stimulation activates body cells and fibers of passage the need of studies with chemical stimulation of only post-synaptic fibers of the dPAG is obvious. To examine further this issue we measured Fos protein expression in brain areas activated by stimulation of the dPAG with glutamate (5 nmol/0.2 microL) and N-methyl-D-aspartate (NMDA) at doses that provoke either freezing (4 nmol/0.2 microL) or escape (7 nmol/0.2 microL) responses, respectively. The results showed that glutamate-induced freezing caused a selective increase in Fos expression in the superior and inferior colliculi as well as in the laterodorsal nucleus of the thalamus. On the other hand, NMDA-induced escape led to widespread increases in Fos labeling in almost all structures studied. Differently from glutamate, NMDA at doses provoking freezing caused significant increase of Fos labeling in the dPAG and CnF. Therefore, the present data support the notion that freezing behavior induced by activation of either non-NMDA or NMDA receptors in the dorsolateral periaqueductal gray (dlPAG) is neurally segregated: glutamate activates only structures that are mainly involved in the sensorial processing and NMDA-induced freezing structures involved in the motor output of defensive behavior. Therefore, the freezing elicited by the activation of non-NMDA receptors seem to be related to the acquisition of aversive information, whereas that resulting from the activation of NMDA receptors could serve as a preparatory response for flight.
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Reação de Fuga/fisiologia , Reação de Congelamento Cataléptica/fisiologia , Vias Neurais/metabolismo , Neurônios/metabolismo , Substância Cinzenta Periaquedutal/metabolismo , Proteínas Proto-Oncogênicas c-fos/metabolismo , Receptores de Glutamato/metabolismo , Tonsila do Cerebelo/citologia , Tonsila do Cerebelo/metabolismo , Animais , Relação Dose-Resposta a Droga , Ácido Glutâmico/administração & dosagem , Ácido Glutâmico/fisiologia , Hipotálamo/citologia , Hipotálamo/metabolismo , Masculino , Microinjeções , Atividade Motora/fisiologia , N-Metilaspartato/administração & dosagem , N-Metilaspartato/fisiologia , Vias Neurais/citologia , Neurônios/efeitos dos fármacos , Percepção/fisiologia , Substância Cinzenta Periaquedutal/citologia , Substância Cinzenta Periaquedutal/efeitos dos fármacos , Ratos , Ratos Wistar , Receptores de Glutamato/efeitos dos fármacos , Formação Reticular/citologia , Formação Reticular/metabolismo , Estimulação Química , Distribuição TecidualRESUMO
The neuromodulatory effect of NO on glutamatergic transmission has been studied in several brain areas. Our previous single-cell studies suggested that NO facilitates glutamatergic transmission in the nucleus of the solitary tract (NTS). In this study, we examined the effect of the nitric oxide synthase (NOS) inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME) on glutamatergic and reflex transmission in the NTS. We measured mean arterial pressure (MAP), heart rate (HR), and renal sympathetic nerve activity (RSNA) from Inactin-anesthetized Sprague-Dawley rats. Bilateral microinjections of L-NAME (10 nmol/100 nl) into the NTS did not cause significant changes in basal MAP, HR, or RSNA. Unilateral microinjection of (RS)-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA, 1 pmol/100 nl) into the NTS decreased MAP and RSNA. Fifteen minutes after L-NAME microinjections, AMPA-evoked cardiovascular changes were significantly reduced. N-methyl-D-aspartate (NMDA, 0.5 pmol/100 nl) microinjection into the NTS decreased MAP, HR, and RSNA. NMDA-evoked falls in MAP, HR, and RSNA were significantly reduced 30 min after L-NAME. To examine baroreceptor and cardiopulmonary reflex function, L-NAME was microinjected at multiple sites within the rostro-caudal extent of the NTS. Baroreflex function was tested with phenylephrine (PE, 25 microg iv) before and after L-NAME. Five minutes after L-NAME the decrease in RSNA caused by PE was significantly reduced. To examine cardiopulmonary reflex function, phenylbiguanide (PBG, 8 microg/kg) was injected into the right atrium. PBG-evoked hypotension, bradycardia, and RSNA reduction were significantly attenuated 5 min after L-NAME. Our results indicate that inhibition of NOS within the NTS attenuates baro- and cardiopulmonary reflexes, suggesting that NO plays a physiologically significant neuromodulatory role in cardiovascular regulation.
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Barorreflexo/fisiologia , Ácido Glutâmico/fisiologia , Coração/inervação , Pulmão/inervação , Óxido Nítrico/fisiologia , Núcleo Solitário/fisiologia , Transmissão Sináptica/fisiologia , Animais , Biguanidas/farmacologia , Pressão Sanguínea/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Agonistas de Aminoácidos Excitatórios/farmacologia , Frequência Cardíaca/efeitos dos fármacos , Rim/inervação , Masculino , Microinjeções , N-Metilaspartato/administração & dosagem , N-Metilaspartato/farmacologia , NG-Nitroarginina Metil Éster/farmacologia , Óxido Nítrico/antagonistas & inibidores , Fenilefrina/farmacologia , Pressorreceptores/efeitos dos fármacos , Pressorreceptores/fisiologia , Ratos , Ratos Sprague-Dawley , Sistema Nervoso Simpático/efeitos dos fármacos , Sistema Nervoso Simpático/fisiologia , Transmissão Sináptica/efeitos dos fármacos , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiônico/administração & dosagem , Ácido alfa-Amino-3-hidroxi-5-metil-4-isoxazol Propiônico/farmacologiaRESUMO
PURPOSE: N-methyl D-aspartate (NMDA) preconditioning has been used to prevent cellular death induced by glutamate or NMDA in cultured neurons. Quinolinic acid (QA)-induced seizures are used to average NMDA receptors-evoked neurotoxicity in animal models. The purpose of this study was to investigate the potential neuroprotective effects of NMDA preconditioning against QA-induced seizures and hippocampal damage in vivo. METHODS: Mice were pretreated with nonconvulsant doses of NMDA for different times before i.c.v. QA infusion and observed for the occurrence of seizures. Hippocampal slices from mice were assayed to measure cellular viability. RESULTS: NMDA preconditioning presented 53% protection against QA-induced seizures, as well as QA-induced cellular death in the hippocampus. The NMDA receptor antagonist, MK-801, prevented the protection evoked by NMDA preconditioning. The adenosine A1 receptor antagonist, CPT, prevented the protection evoked by NMDA preconditioning against QA-induced seizures, but not against QA-induced hippocampal cellular damage. The adenosine A1 receptor agonist, CPA, did not mimic the NMDA preconditioning-evoked protective effects. CONCLUSIONS: These results suggest that in vivo preconditioning with subtoxic doses of NMDA protected mice against seizures and cellular hippocampal death elicited by QA, probably through mechanisms involving NMDA receptors operating with adenosine A1 receptors.