RESUMO
CD82, also known as KAI1, was recently identified as a prostate cancer metastasis suppressor gene on human chromosome 11p1.2 (ref. 1). The product of CD82 is KAI1, a 40- to 75-kDa tetraspanin cell-surface protein also known as the leukocyte cell-surface marker CD82 (refs. 1,2). Downregulation of KAI1 has been found to be clinically associated with metastatic progression in a variety of cancers, whereas overexpression of CD82 specifically suppresses tumor metastasis in various animal models. To define the mechanism of action of KAI1, we used a yeast two-hybrid screen and identified an endothelial cell-surface protein, DARC (also known as gp-Fy), as an interacting partner of KAI1. Our results indicate that the cancer cells expressing KAI1 attach to vascular endothelial cells through direct interaction between KAI1 and DARC, and that this interaction leads to inhibition of tumor cell proliferation and induction of senescence by modulating the expression of TBX2 and p21. Furthermore, the metastasis-suppression activity of KAI1 was significantly compromised in DARC knockout mice, whereas KAI1 completely abrogated pulmonary metastasis in wild-type and heterozygous littermates. These results provide direct evidence that DARC is essential for the function of CD82 as a suppressor of metastasis.
Assuntos
Sistema do Grupo Sanguíneo Duffy/metabolismo , Endotélio Vascular/metabolismo , Proteína Kangai-1/metabolismo , Neoplasias Pulmonares/patologia , Glicoproteínas de Membrana/metabolismo , Metástase Neoplásica/prevenção & controle , Receptores de Superfície Celular/metabolismo , Alelos , Sequência de Aminoácidos , Animais , Sequência de Bases , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Senescência Celular/fisiologia , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Sistema do Grupo Sanguíneo Duffy/química , Feminino , Heterozigoto , Humanos , Proteína Kangai-1/química , Masculino , Glicoproteínas de Membrana/química , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Knockout , Dados de Sequência Molecular , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Estrutura Terciária de Proteína , Ratos , Receptores de Superfície Celular/química , Proteínas com Domínio T/metabolismoRESUMO
We examined the role of aldosterone-sensitive neurons in the nucleus tractus solitarius (NTS) in the arterial baroreceptor reflex (baroreflex) function. Baroreflex sensitivity was induced by phenylephrine in high sodium-loaded rats and was significantly reduced. This baroreflex sensitivity was reversed by microinjection of the mineralocorticoid receptor (MR) antagonist eplerenone into the NTS. 11beta-Hydroxysteroid dehydrogenase type 2 neurons and MR were also identified in the NTS. These data suggest that the aldosterone-sensitive neurons in the NTS may have an important role in baroreflex function.
Assuntos
Aldosterona/farmacologia , Neurônios/efeitos dos fármacos , Pressorreceptores/efeitos dos fármacos , Sódio/administração & dosagem , Núcleo Solitário/efeitos dos fármacos , Animais , Ratos , Núcleo Solitário/citologiaRESUMO
Mutations of genes encoding alpha4, beta2, or alpha2 subunits (CHRNA4, CHRNB2, or CHRNA2, respectively) of nAChR [neuronal nicotinic ACh (acetylcholine) receptor] cause nocturnal frontal lobe epilepsy (NFLE) in human. NFLE-related seizures are seen exclusively during sleep and are characterized by three distinct seizure phenotypes: "paroxysmal arousals," "paroxysmal dystonia," and "episodic wandering." We generated transgenic rat strains that harbor a missense mutation S284L, which had been identified in CHRNA4 in NFLE. The transgenic rats were free of biological abnormalities, such as dysmorphology in the CNS, and behavioral abnormalities. The mRNA level of the transgene (mutant Chrna4) was similar to the wild type, and no distorted expression was detected in the brain. However, the transgenic rats showed epileptic seizure phenotypes during slow-wave sleep (SWS) similar to those in NFLE exhibiting three characteristic seizure phenotypes and thus fulfilled the diagnostic criteria of human NFLE. The therapeutic response of these rats to conventional antiepileptic drugs also resembled that of NFLE patients with the S284L mutation. The rats exhibited two major abnormalities in neurotransmission: (1) attenuation of synaptic and extrasynaptic GABAergic transmission and (2) abnormal glutamate release during SWS. The currently available genetically engineered animal models of epilepsy are limited to mice; thus, our transgenic rats offer another dimension to the epilepsy research field.
Assuntos
Epilepsia do Lobo Frontal/genética , Leucina/genética , Mutação/genética , Receptores Nicotínicos/genética , Serina/genética , Transmissão Sináptica/genética , Ácido gama-Aminobutírico/metabolismo , Fatores Etários , Análise de Variância , Animais , Comportamento Animal/fisiologia , Córtex Cerebral/citologia , Córtex Cerebral/patologia , Relação Dose-Resposta a Droga , Eletroencefalografia , Epilepsia do Lobo Frontal/dietoterapia , Epilepsia do Lobo Frontal/fisiopatologia , Antagonistas de Aminoácidos Excitatórios/farmacologia , Antagonistas GABAérgicos/farmacologia , Ácido Glutâmico/metabolismo , Temperatura Alta/efeitos adversos , Técnicas In Vitro , Microscopia Imunoeletrônica , Atividade Motora/genética , Destreza Motora/fisiologia , Neurotransmissores/metabolismo , Nicotina/farmacologia , Medição da Dor/métodos , Pentilenotetrazol/farmacologia , Fenótipo , Ratos , Ratos Transgênicos , Receptores Nicotínicos/metabolismo , Receptores Nicotínicos/ultraestrutura , Transmissão Sináptica/efeitos dos fármacosRESUMO
Painful diabetic neuropathy causes hyperalgesia and does not respond to commonly used analgesics such as non-steroidal anti-inflammatory drugs or opioids at doses below those producing disruptive side effects. In the present study, we examined the effect of P2X receptor antagonists, which are known to modulate the pain pathway, on mechanical hyperalgesia in streptozotocin (STZ)-induced diabetic mice. The paw withdrawal frequency measured by von Frey filaments, began to significantly increase 5 days after STZ injection and was maintained for more than 14 days. Intrathecal administration of P2X receptor antagonists (PPADS and TNP-ATP) inhibited the mechanical allodynia in diabetic mice. The levels of P2X(2) and P2X(3) receptors mRNA were significantly increased in diabetic mice at 14 days after the intravenous injection of STZ. These results suggest that the upregulation of P2X(2), P2X(3) and/or P2X(2/3) receptor in DRG neurons is associated with mechanical allodynia in STZ-induced diabetic mice.
Assuntos
Diabetes Mellitus Experimental/metabolismo , Neuropatias Diabéticas/metabolismo , Gânglios Espinais/metabolismo , Nociceptores/metabolismo , Receptores Purinérgicos P2/metabolismo , Células Receptoras Sensoriais/metabolismo , Trifosfato de Adenosina/análogos & derivados , Trifosfato de Adenosina/farmacologia , Animais , Diabetes Mellitus Experimental/complicações , Diabetes Mellitus Experimental/fisiopatologia , Neuropatias Diabéticas/fisiopatologia , Modelos Animais de Doenças , Hiperalgesia/etiologia , Hiperalgesia/metabolismo , Hiperalgesia/fisiopatologia , Masculino , Camundongos , Nociceptores/patologia , Medição da Dor , Estimulação Física , Inibidores da Agregação Plaquetária , Antagonistas do Receptor Purinérgico P2 , Fosfato de Piridoxal/análogos & derivados , Fosfato de Piridoxal/farmacologia , RNA Mensageiro/efeitos dos fármacos , RNA Mensageiro/metabolismo , Receptores Purinérgicos P2/genética , Receptores Purinérgicos P2X2 , Receptores Purinérgicos P2X3 , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/fisiologiaRESUMO
Involvement of Na+/Ca2+ exchanger (NCX) in pentylenetetrazol (PTZ)-induced convulsion by use of NCX knockout mice and the selective ligand SEA0400 to NCX was examined. In the SEA0400-administered group, the latency to clonic convulsion was extended into 210 s, although the latency to clonic convulsion was observed until 100 s in control group. SEA0400 had little effect on bicuculline-induced clonic seizure nicotine-induced wild running and 4-aminopyridine-induced tonic flexion, respectively. Tonic flexion convulsion was occurred three fifth in the wild type mice group by administration of PTZ, but tonic flexion was not observed in NCX1 knockout mice groups. These results suggest that NCX is involved in inhibitory action in PTZ-induced convulsion.
Assuntos
Convulsivantes/toxicidade , Pentilenotetrazol/toxicidade , Convulsões/induzido quimicamente , Trocador de Sódio e Cálcio/fisiologia , Animais , Camundongos , Camundongos Knockout , Convulsões/fisiopatologia , Trocador de Sódio e Cálcio/genéticaRESUMO
Fatty acid synthase (FAS) has been found to be overexpressed in a wide range of epithelial tumors, including breast cancer. Pharmacologic inhibitors of FAS cause apoptosis of breast cancer cells and result in decreased tumor size in vivo. However, how the inhibition of FAS induces apoptosis in tumor cells remains largely unknown. To understand the apoptotic pathway resulting from direct inhibition of FAS, we treated breast tumor cells with or without FAS small interfering RNA (siRNA) followed by a microarray analysis. Our results indicated that the proapoptotic genes BNIP3, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), and death-associated protein kinase 2 (DAPK2) were significantly up-regulated on direct inhibition of the FAS gene. We also found that the knockdown of FAS expression significantly increased ceramide level in the tumor cells, and this increase was abrogated by acetyl-CoA carboxylase inhibitor. In addition, carnitine palmitoyltransferase-1 (CPT-1) inhibitor up-regulated the ceramide and BNIP3 levels in these cells, whereas treatment of tumor cells with FAS siRNA in the presence of a ceramide synthase inhibitor abrogated the up-regulation of BNIP3 and inhibited apoptosis. Furthermore, we found that treatment of cells with BNIP3 siRNA significantly counteracted the effect of FAS siRNA-mediated apoptosis. Consistent with these results, a significant inverse correlation was observed in the expression of FAS and BNIP3 in clinical samples of human breast cancer. Collectively, our results indicate that inhibition of FAS in breast cancer cells causes accumulation of malonyl-CoA, which leads to inhibition of CPT-1 and up-regulation of ceramide and induction of the proapoptotic genes BNIP3, TRAIL, and DAPK2, resulting in apoptosis.
Assuntos
Apoptose/fisiologia , Neoplasias da Mama/enzimologia , Ácido Graxo Sintases/antagonistas & inibidores , Apoptose/genética , Proteínas Reguladoras de Apoptose/genética , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Proteínas Quinases Dependentes de Cálcio-Calmodulina/genética , Linhagem Celular Tumoral , Proteínas Quinases Associadas com Morte Celular , Ácido Graxo Sintases/genética , Humanos , Glicoproteínas de Membrana/genética , Proteínas de Membrana/biossíntese , Proteínas de Membrana/genética , Proteínas Proto-Oncogênicas/biossíntese , Proteínas Proto-Oncogênicas/genética , RNA Interferente Pequeno/genética , Ligante Indutor de Apoptose Relacionado a TNF , Fator de Necrose Tumoral alfa/genética , Regulação para CimaRESUMO
The tumor metastasis suppressor gene Drg-1 has been shown to suppress metastasis without affecting tumorigenicity in immunodeficient mouse models of prostate and colon cancer. Expression of Drg-1 has also been found to have a significant inverse correlation with metastasis or invasiveness in various types of human cancer. However, how Drg-1 exerts its metastasis suppressor function remains unknown. In the present study, to elucidate the mechanism of action of the Drg-1 gene, we did a microarray analysis and found that induction of Drg-1 significantly inhibited the expression of activating transcription factor (ATF) 3, a member of the ATF/cyclic AMP-responsive element binding protein family of transcription factors. We also showed that Drg-1 attenuated the endogenous level of ATF3 mRNA and protein in prostate cancer cells, whereas Drg-1 small interfering RNA up-regulated the ATF3 expression. Furthermore, Drg-1 suppressed the promoter activity of the ATF3 gene, indicating that Drg-1 regulates ATF3 expression at the transcriptional level. Our immunohistochemical analysis on prostate cancer specimens revealed that nuclear expression of ATF3 was inversely correlated to Drg-1 expression and positively correlated to metastases. Consistently, we have found that ATF3 overexpression promoted invasiveness of prostate tumor cells in vitro, whereas Drg-1 suppressed the invasive ability of these cells. More importantly, overexpression of ATF3 in prostate cancer cells significantly enhanced spontaneous lung metastasis of these cells without affecting primary tumorigenicity in a severe combined immunodeficient mouse model. Taken together, our results strongly suggest that Drg-1 suppresses metastasis of prostate tumor cells, at least in part, by inhibiting the invasive ability of the cells via down-regulation of the expression of the ATF3 gene.
Assuntos
Fator 3 Ativador da Transcrição/genética , Proteínas de Ligação ao GTP/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias da Próstata/genética , Linhagem Celular Tumoral , Humanos , Masculino , Metástase Neoplásica/genética , Metástase Neoplásica/prevenção & controle , Plasmídeos , Neoplasias da Próstata/patologia , Mapeamento por Restrição , TransfecçãoRESUMO
Chronic morphine exposure upregulates adenylate cyclase signaling and reduces analgesic efficacy, a condition known as opioid tolerance. Nonopioid neurotransmitters can enhance morphine tolerance, but the mechanism for this is poorly understood. We show that morphine tolerance was delayed in mice lacking vasopressin 1b receptors (V1bRs) or after administration of V1bR antagonist into the rostral ventromedial medulla, where transcripts for V1bRs and µ-opioid receptors are co-localized. Vasopressin increased morphine-binding affinity in cells expressing both V1bR and µ-opioid receptors. Complex formation among V1bR, ß-arrestin-2, and µ-opioid receptor resulted in vasopressin-mediated upregulation of ERK phosphorylation and adenylate cyclase sensitization. A leucine-rich segment in the V1bR C-terminus was necessary for the association with ß-arrestin-2. Deletion of this leucine-rich segment increased morphine analgesia and reduced vasopressin-mediated adenylate cyclase sensitization. These findings indicate that inhibition of µ-opioid-receptor-associated V1bR provides an approach for enhancing morphine analgesia without increasing analgesic tolerance.
Assuntos
Tolerância a Medicamentos/genética , Morfina/farmacologia , Entorpecentes/farmacologia , Receptores Opioides mu/metabolismo , Receptores de Vasopressinas/metabolismo , beta-Arrestina 2/metabolismo , Adenilil Ciclases/metabolismo , Animais , Sinalização do Cálcio/efeitos dos fármacos , Sinalização do Cálcio/genética , Injeções , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Bulbo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Morfina/farmacocinética , Dependência de Morfina/psicologia , Entorpecentes/farmacocinética , Medição da Dor/efeitos dos fármacos , Limiar da Dor/efeitos dos fármacos , Fosforilação , Receptores Opioides mu/genética , Receptores de Vasopressinas/genética , beta-Arrestina 2/genéticaRESUMO
We analyzed the role of the Na+/Ca2+ exchanger (NCX) in endothelin-1-aggravated hypoxia/reoxygenation-induced injury in renal epithelial LLC-PK1 cells. KB-R7943, a selective NCX inhibitor, suppressed hypoxia/reoxygenation-induced cell damage, whereas overexpression of NCX1 into cells enhanced it. Endothelin-1 significantly aggravated hypoxia/reoxygenation-induced injury in parental and NCX1-overexpressing LLC-PK1 cells. Such aggravation by endothelin-1 was not observed in cells overexpressing a deregulated NCX1 mutant, which displays no protein kinase C-dependent activation. These results suggest that Ca2+ overload via NCX plays a critical role in hypoxia/reoxygenation-induced renal tubular injury, and that endothelin-1 aggravates the cell damage through the activation of NCX.
Assuntos
Endotelina-1/fisiologia , Hipóxia/fisiopatologia , Rim/patologia , Oxigênio/metabolismo , Animais , Células Epiteliais/patologia , Células LLC-PK1 , Suínos , Tioureia/análogos & derivados , Tioureia/farmacologiaRESUMO
The arginine vasopressin (AVP) system plays an important role in social behavior. Autism, with its hallmark disturbances in social behavior, has been associated with the V1a receptor (V1aR) gene. Furthermore, impairments of social function are often observed in symptoms of schizophrenia. Subchronic phencyclidine (PCP) produces behaviors relating to certain aspects of schizophrenic symptoms such as impairing social interaction in animals and it reduces the density of V1aR binding sites in several brain regions. Here, we report that V1aR knockout (KO) mice exhibited impairment of social behavior in a social interaction test, and showed reduced anxiety-related behavior in elevated plus-maze and marble-burying behavior tests. Given the current findings, the V1aR may be involved in the regulation of social interaction, and V1aR KO mice could be used as an animal model of psychiatric disorders associated with social behavior deficits, such as autism and schizophrenia.
Assuntos
Ansiedade/metabolismo , Comportamento Exploratório/fisiologia , Receptores de Vasopressinas/fisiologia , Comportamento Social , Animais , Ansiedade/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Vasopressinas/genética , Estatísticas não ParamétricasRESUMO
PTEN (phosphatase and tensin homologue deleted on chromosome 10) has been shown to be inactivated in a wide variety of cancers, and the role of this gene as a tumor suppressor has been well established. On the other hand, results of recent animal studies as well as clinical evidence indicate that PTEN is also involved in tumor metastasis suppression. Although PTEN is known to play a key role in controlling cell growth and apoptosis, how PTEN exerts the metastasis suppressor function remains largely unknown. Recently, a microarray analysis identified the Drg-1 gene (differentiation related gene 1) as one of the potential targets of PTEN. The Drg-1 gene has been shown to suppress tumor metastasis in animal models of prostate and colon cancer, and the expression of this gene is significantly reduced with advancement of prostate and breast cancers in clinical setting. In this study, we explored the possibility that PTEN controls tumor metastasis by regulating the expression of the Drg-1 gene. Our results indicate that overexpression of PTEN significantly augments the endogenous expression of Drg-1 protein, whereas inhibition of PTEN by small interfering RNA decreases Drg-1 in a dose- and time-dependent manner. We also found that the control of the Drg-1 gene by PTEN seems to be at the transcriptional level, and that a phospho-Akt inhibitor restores the Drg-1 expression, indicating that PTEN controls Drg-1 by an Akt-dependent pathway. Consistent with these results, our immunohistochemical analysis revealed that PTEN expression correlates significantly with Drg-1 in both prostate and breast cancer cases. Furthermore, combination of the two markers, PTEN and Drg-1, emerged as a significantly better predictor of prostate and breast cancer patient survival than either marker alone.
Assuntos
Neoplasias da Mama/genética , Proteínas de Ciclo Celular/genética , Genes Supressores de Tumor , Monoéster Fosfórico Hidrolases/fisiologia , Neoplasias da Próstata/genética , Proteínas Supressoras de Tumor/fisiologia , Neoplasias da Mama/mortalidade , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Peptídeos e Proteínas de Sinalização Intracelular , Masculino , PTEN Fosfo-Hidrolase , Neoplasias da Próstata/mortalidade , Taxa de Sobrevida , Regulação para CimaRESUMO
Arginine vasopressin (AVP) is a neurohypophyseal peptide best known as an antidiuretic hormone. AVP receptors have been classified into three subtypes: V1a, V1b, and V2 receptors. The V1a receptor (V1aR) and V1b receptor (V1bR) are widely distributed in the central nervous system, including the cortex and hippocampus. In the present study, we examined the performance of V1aR or V1bR knockout (KO) mice compared to wild-type (WT) mice in behavioral tests. V1aR KO mice exhibited impairments of spatial learning (eight-arm radial maze), prepulse inhibition (PPI) and social behavior in comparison to WT mice. On the other hand, V1bR KO mice also displayed impairments of PPI and social behavior. These results suggest that V1aR and V1bR may be involved in psychiatric disorders associated with impairments of sensorimotor gating and social behavior such as schizophrenia and autism.
Assuntos
Transtornos Mentais/etiologia , Receptores de Vasopressinas/deficiência , Animais , Modelos Animais de Doenças , Camundongos , Camundongos KnockoutRESUMO
The differentiation-related gene-1 (Drg-1) was first identified as a gene strongly upregulated by induction of differentiation in colon carcinoma cells in vitro, and later the same gene was shown to suppress tumorigenicity of human bladder cancer cells in vivo. On the other hand, we and others have demonstrated that the Drg-1 gene suppresses prostate and colon cancer metastases in mouse models. In the context of such potential organ-specific differential function of the Drg-1 gene, the present study was designed to clarify the expression status, regulation and function of Drg-1 in the case of human breast cancer. We found that the expression of the Drg-1 protein was significantly reduced in breast tumor cells, particularly in patients with lymph node or bone metastasis as compared to those with localized breast cancer. Drg-1 expression also exhibited significant inverse correlation with the disease-free survival rate of patients and emerged as an independent prognostic factor. The downregulation of the Drg-1 gene appeared to be largely at the RNA level, and the DNA methylation inhibitor, 5-Azacytidine, significantly elevated the Drg-1 gene expression in various breast tumor cell lines. Furthermore, we found that overexpression of the Drg-1 gene suppresses the invasiveness of breast cancer cells in vitro, and this suppression was also achieved by treatment of cells with 5-Azacytidine. Together, our results strongly suggest functional involvement of the Drg-1 gene in suppressing the metastatic advancement of human breast cancer.
Assuntos
Neoplasias da Mama/genética , Proteínas de Ligação ao GTP/genética , Metilação de DNA , Progressão da Doença , Intervalo Livre de Doença , Feminino , Regulação Neoplásica da Expressão Gênica , Genes Supressores de Tumor , Humanos , Metástase Neoplásica , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células Tumorais CultivadasRESUMO
In the present study, we investigated whether mice lacking the arginine vasopressin (AVP) V1b receptor (V1bR) exhibit deficits of prepulse inhibition (PPI) of the startle reflex, reminiscent of the sensorimotor gating deficits observed in a large majority of schizophrenic patients. V1bR knockout (KO) mice displayed significantly reduced levels of PPI of the startle reflex. In addition to PPI deficits, V1bR KO mice showed increased acoustic startle response. However, acoustic startle response was not significantly correlated to the PPI of the startle reflex in V1bR KO mice. V1bR KO mice also showed a decrease in basal levels of extracellular dopamine (DA) in the medial prefrontal cortex, which is thought to be an important brain region for PPI. Moreover, PPI deficits observed in the V1bR KO mice are significantly reversed by atypical antipsychotics such as risperidone and clozapine but not by a typical neuroleptic haloperidol, like in schizophrenic patients. By contrast, we did not observe any significant differences between V1bR KO mice and wild-type mice in the open-field, light/dark, elevated plus maze, and forced swimming tests. The results of the present study indicate that V1bR may be involved in the regulation of PPI of the startle reflex. The V1bR has been considered an important molecular target for the development of antipsychotic drugs.
Assuntos
Antipsicóticos/farmacologia , Inibição Neural/efeitos dos fármacos , Receptores de Vasopressinas/deficiência , Reflexo de Sobressalto/efeitos dos fármacos , Estimulação Acústica/métodos , Análise de Variância , Animais , Comportamento Animal/efeitos dos fármacos , Relação Dose-Resposta a Droga , Relação Dose-Resposta à Radiação , Interações Medicamentosas , Comportamento Exploratório/efeitos dos fármacos , Masculino , Aprendizagem em Labirinto/efeitos dos fármacos , Camundongos , Camundongos Knockout , Microdiálise/métodos , Reflexo de Sobressalto/fisiologia , Estatística como Assunto , NataçãoRESUMO
Nociceptive behaviors were examined in the mice lacking alpha1d-adrenergic receptor (alpha1d-AR) and wild type littermates using tail-flick, hot-plate (hindpaw-licking and jumping), tail-pinch and formalin tests. The distribution of alpha1d-AR was studied using in situ hybridization in the wild type mice. Mutant mice showed longer tail-flick and hindpaw-licking latencies while their jumping latency was shorter. Mechanical and chemical nociception was not altered in alpha1d-knockout mice. In situ hybridization study revealed dense alpha1d-AR mRNA expression in the reticular thalamic nucleus, the hippocampus, the cingulate cortex and the spinal cord. These results suggest that alpha1d-AR in the spinal cord contributes to thermal pronociception; and that the jump behavior seen when escaping from heat is inhibited via the supraspinal alpha1d-AR.
Assuntos
Camundongos Knockout/fisiologia , Dor/metabolismo , Receptores Adrenérgicos alfa 1/fisiologia , Animais , Comportamento Animal , Encéfalo/anatomia & histologia , Encéfalo/metabolismo , Hibridização In Situ/métodos , Masculino , Camundongos , Medição da Dor/métodos , RNA Mensageiro/metabolismo , Tempo de Reação , Receptores Adrenérgicos alfa 1/deficiência , Receptores Adrenérgicos alfa 1/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/métodos , Medula Espinal/metabolismo , Fatores de TempoRESUMO
To investigate the functional role of alpha1d-adrenergic receptor (alpha1d-AR) in the CNS, we have generated mutant mice lacking the alpha1d-AR using a gene targeting approach and examined in detail the effects of alpha1d-AR knockout mice on motor function, sensory function, and learning and memory. alpha1d-AR knockout mice showed better motor coordination at the highest rotating speed of the rotarod performance and stronger muscle tone using the traction meter, but their locomotor activity and swimming ability in the water maze were not affected. In the water maze requiring reference memory, alpha1d-AR knockout mice showed normal spatial learning. In the Y-maze task requiring working memory or attention, alpha1d-AR knockout mice displayed an impaired spontaneous alternation performance. The alpha1d-AR knockout mice tended to display lower levels of acoustic startle responses than the wild-type group at lower pulse intensities, although the acoustic prepulse inhibition was not impaired in the alpha1d-AR knockout mice. Furthermore, the NMDA receptor antagonist, MK-801-induced deficits of acoustic prepulse inhibition were not observed in the alpha1d-AR knockout mice. These results clearly demonstrate that the alpha1d-AR receptor plays an important role in the process of auditory sensory function, attention or working memory rather than reference memory, and the sensorimotor gating deficits induced by the NMDA receptor antagonist.
Assuntos
Transtornos Mentais/fisiopatologia , Desempenho Psicomotor/fisiologia , Receptores Adrenérgicos alfa 1/deficiência , Receptores Adrenérgicos alfa 1/fisiologia , Estimulação Acústica/métodos , Análise de Variância , Animais , Atenção/fisiologia , Relação Dose-Resposta à Radiação , Inibição Psicológica , Masculino , Aprendizagem em Labirinto/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout/fisiologia , Atividade Motora/fisiologia , Tono Muscular/fisiologia , Tempo de Reação , Receptores Adrenérgicos alfa 1/genética , Reflexo Acústico/fisiologia , Comportamento Espacial/fisiologia , Estatísticas não Paramétricas , Fatores de TempoRESUMO
We examined the involvement of the spinal muscarinic receptors in the clonidine-induced antiallodynic effects. Mechanical sensitivity was assessed by stimulating the hind paw with von Frey filaments. In streptozotocin-treated (200 mg/kg, i.v.) diabetic mice, hypersensitivity to mechanical stimulation appeared 3 days after streptozotocin administration, and persisted for 11 days. This mechanical hypersensitivity (allodynia) was inhibited by the intrathecal (i.t.) injection of clonidine. The muscarinic receptor antagonist atropine (i.t.) and alpha2-adrenoreceptor antagonist yohimbine (i.t. or subcutaneous injection) abolished the antiallodynic effect of clonidine. The effect was mimicked by the muscarinic M1 receptor antagonist pirenzepine, but not by the muscarinic M2 receptor antagonist methoctoramine or the muscarinic M3 receptor antagonist 4-DAMP (4-diphenyl-acetoxy-N-methylpiperidine methiodide). In addition, the mechanical hypersensitivity in diabetic mice was reduced by the selective muscarinic M1 receptor agonist McN-A-343 (4-(m-chlorophenyl-carbamoyloxy)-2-butynyltrimethylammonium chloride) (i.t.). These results suggest that spinal muscarinic M1 receptors participate in the antiallodynic effect of clonidine in diabetic mice.
Assuntos
Clonidina/farmacologia , Diabetes Mellitus Experimental/fisiopatologia , Dor/prevenção & controle , Receptor Muscarínico M1/metabolismo , Medula Espinal/efeitos dos fármacos , Cloreto de (4-(m-Clorofenilcarbamoiloxi)-2-butinil)trimetilamônio/farmacologia , Analgésicos/administração & dosagem , Analgésicos/farmacologia , Animais , Atropina/farmacologia , Glicemia/efeitos dos fármacos , Glicemia/metabolismo , Clonidina/administração & dosagem , Diabetes Mellitus Experimental/sangue , Diabetes Mellitus Experimental/metabolismo , Diaminas/farmacologia , Injeções Espinhais , Masculino , Camundongos , Atividade Motora/efeitos dos fármacos , Agonistas Muscarínicos/farmacologia , Antagonistas Muscarínicos/farmacologia , Dor/etiologia , Limiar da Dor/efeitos dos fármacos , Piperidinas/farmacologia , Pirenzepina/farmacologia , Receptor Muscarínico M1/agonistas , Receptor Muscarínico M1/antagonistas & inibidores , Receptor Muscarínico M2/antagonistas & inibidores , Receptor Muscarínico M3/antagonistas & inibidores , Medula Espinal/metabolismo , Medula Espinal/fisiopatologia , Estreptozocina/administração & dosagem , Estresse Mecânico , Fatores de TempoRESUMO
The present study was undertaken to clarify how spinal muscarinic receptors can be involved in the antinociceptive effects induced by morphine in thermal stimulation. The morphine-induced antinociceptive effects (26.6 micromol/kg, s.c.) was inhibited by an intrathecal (i.t.) injection of the muscarinic antagonist (M) atropine and the M(1)/M(4) antagonist pirenzepine in a dose-dependent manner. In contrast, the M(2) antagonist methoctramine and the M(3) antagonist 4-DAMP did not inhibit the morphine-induced antinociceptive effects. Injection (i.t.) of the putative M(1) agonist McN-A-343 resulted in dose-dependent antinociceptive effects in thermal stimuli. In addition, antinociceptive effects induced by the i.t. injection of morphine were not inhibited by the M(1)/M(4) antagonist pirenzepine, although pirenzepine did inhibit the intracerebroventricular (i.c.v.) injection of morphine-induced antinociceptive effects. These results suggest that the morphine-induced antinociceptive effects in thermal stimuli are regulated by the M(1) or M(4) receptor in the spinal cord.
Assuntos
Analgésicos/farmacologia , Temperatura Alta , Morfina/farmacologia , Medição da Dor/efeitos dos fármacos , Receptores Muscarínicos/classificação , Receptores Muscarínicos/fisiologia , Medula Espinal/efeitos dos fármacos , Animais , Relação Dose-Resposta a Droga , Camundongos , Medição da Dor/métodos , Receptor Muscarínico M1/fisiologia , Medula Espinal/fisiologiaRESUMO
We examined the role of the spinal muscarinic receptor subtype in the anti-nociceptive effect of intrathecal (i.t.) alpha2 adrenoceptor agonist clonidine in mice. I.t. injection of the muscarinic receptor antagonist atropine completely inhibited i.t. clonidine-induced increase in the mechanical threshold, but did not affect the increase in tail-flick latency induced by i.t. clonidine. The clonidine-induced increase in mechanical threshold was inhibited by i.t. injection of the M1 receptor antagonist pirenzepine in a dose-dependent manner, and by the M3 receptor antagonist 4-DAMP, but not by the M2 receptor antagonist methoctramine. The potency of pirenzepine was greater than that of 4-DAMP. These results suggest that the clonidine-induced increase in mechanical threshold is mediated via the activation of M1 receptors in the spinal cord.
Assuntos
Agonistas de Receptores Adrenérgicos alfa 2 , Agonistas alfa-Adrenérgicos/farmacologia , Clonidina/farmacologia , Mecanorreceptores/efeitos dos fármacos , Receptores Muscarínicos/efeitos dos fármacos , Medula Espinal/efeitos dos fármacos , Transmissão Sináptica/efeitos dos fármacos , Agonistas alfa-Adrenérgicos/administração & dosagem , Antagonistas Adrenérgicos alfa/farmacologia , Analgésicos/farmacologia , Animais , Clonidina/administração & dosagem , Temperatura Alta , Injeções Espinhais , Masculino , Camundongos , Antagonistas Muscarínicos/farmacologia , Medição da Dor/efeitos dos fármacos , Limiar da Dor/efeitos dos fármacos , Estimulação Física , Tempo de Reação/efeitos dos fármacos , Receptor Muscarínico M1 , Ioimbina/farmacologiaRESUMO
In this study, we examined the performance of vasopressin V1a receptor (V1aR) and vasopressin V1b receptor (V1bR) knockout (KO) mice compared to wild-type (WT) mice in an eight-arm radial maze. V1aR KO mice exhibited an impairment of spatial memory in comparison to WT mice. By contrast, we did not observe any significant differences between the V1bR KO mice and the WT mice in the eight-arm radial maze. Moreover, OPC-21268, a selective V1aR antagonist, impaired spatial memory in the eight-arm radial maze in WT mice characterized by an increased number of errors. These results suggest that the V1aR controls spatial memory in mice.