RESUMEN
Neuronal activity elicits a rapid increase in the expression of several immediate early genes (IEGs). To clarify a role for IEG response in activity-dependent development, we examined the contribution of the egr1/zif268 gene during visual cortical processing and plasticity in mice. We first analyzed the expression of egr1 mRNA in wild-type (WT) mice using Northern blot hybridization. In the visual cortex, expression of egr1 mRNA increased dramatically after eye opening, systemic injection of kainate, or 30 min of photostimulation after a brief (5 d) period of dark adaptation. Thus, the expression of egr1 is regulated by synaptic activity in the mouse visual cortex, as it is in other species (e.g., monkeys, cats, and rats). To evaluate whether this transcription factor is directly involved in activity-dependent plasticity, mice lacking Egr1 were deprived of the use of one eye during the developmental critical period [postnatal day 24 (P24)-P34]. Extracellular in vivo single-unit recordings from the binocular zone of the visual cortex revealed that visual responses developed normally in egr1 knock-out (KO) mice. Moreover, a similarly significant shift of responsiveness in favor of the open eye was produced in both KO and WT mice by either brief (4 d) or long-term (>2 weeks) occlusion of one eye. There was no apparent compensation among egr2, egr3, or c-fos mRNA and protein expression in the visual cortex of egr1 KO mice. Taken together, these results indicate that egr1 is a useful marker of sensory input in mice but is not intrinsically necessary for the experience-dependent plasticity of the visual cortex. Our findings underscore a mechanistic distinction between sensory plasticity and long-lasting forms of synaptic potentiation in the hippocampus, for which egr1/zif268 was recently found to be essential.
Asunto(s)
Proteínas de Unión al ADN/deficiencia , Predominio Ocular/fisiología , Proteínas Inmediatas-Precoces , Plasticidad Neuronal/fisiología , Neuronas/metabolismo , Factores de Transcripción/deficiencia , Corteza Visual/metabolismo , Envejecimiento/fisiología , Animales , Biomarcadores , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Adaptación a la Oscuridad/fisiología , Proteína 1 de la Respuesta de Crecimiento Precoz , Proteína 2 de la Respuesta de Crecimiento Precoz , Proteína 3 de la Respuesta de Crecimiento Precoz , Marcación de Gen , Ácido Kaínico/farmacología , Ratones , Ratones Noqueados , Neuronas/efectos de los fármacos , Estimulación Luminosa , Proteínas Proto-Oncogénicas c-fos/genética , Proteínas Proto-Oncogénicas c-fos/metabolismo , ARN Mensajero/metabolismo , Privación Sensorial/fisiología , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Corteza Visual/citología , Corteza Visual/efectos de los fármacos , Percepción Visual/fisiologíaRESUMEN
A series of 1,4-phenylene-bridged ZP-HP hybrid porphyrins (ZP = zinc porphyrin, HP = free-base porphyrin) 1-8 ZH have been prepared in which an electron-donating ZP moiety is kept constant and electron-accepting HP moieties are varied by introducing electron-accepting substituents, so that the energy gap for charge separation, ZP-1HP*--> ZP(+)-HP-, covers a range of about 0.9 eV in DMF. Here selective excitation at the HP moiety was employed to avoid complication in the determination of electron transfer rates derived from energy transfer, 1ZP*-HP --> ZP-1HP*. Definitive evidence for the electron transfer has been obtained in three solvents (benzene, THF, and DMF) through picosecond-femtosecond transient absorption studies, which have allowed the determination of the rates of the photoinduced charge separation, ZP-1HP* --> ZP(+)-HP-, and subsequent thermal charge recombination ZP(+)-HP- --> ZP-HP. Dyad 1ZH in THF exhibits a biphasic fluorescence decay that indicates thermal repopulation of the ZP-1HP* from ZP(+)-HP-; this has been also supported by the transient absorption spectra. On this ground, the energy levels of the ZP(+)-HP- ion pairs have been estimated. Similar biphasic fluorescence decay has been observed for 5 ZH in benzene; this allows furhter estimation of the energy level of the ZP(+)-HP- ion pairs. The free-energy-gap dependence (energy-gap law) has been probed from the normal to the upper limit region for the rate of the charge separation alone, and only the inverted region for the rate of the charge recombination. It was not possible to reproduce both energy-gap dependencies of the charge separation and the charge recombination assuming common parameter values for the reorganization energy and electronic interaction responsible for the electron transfer with the classical Marcus equation. Although both energy-gap dependencies can be approximately reproduced by means of the simplified semiclassical equation, which takes into consideration the effect of the high-frequency vibrations replaced by one mode of averaged frequency, many features, which include the effects of solvent polarity, electron-tunneling matrix element, and so forth on the energy-gap law, are considerably different from those of the previous studied porphyrin-quinone systems with weaker inter-chromophore electronic interactions.
RESUMEN
PURPOSE: To evaluate the hypothesis that the expression of the calcium-binding protein parvalbumin (PV) in a subpopulation of gamma-aminobutyric acid (GABA)ergic neurons is an appropriate molecular marker for the effect on ocular dominance plasticity of monocular deprivation during the postnatal sensitive period. METHODS: Long-Evans rats underwent monocular enucleation immediately before eye opening (postnatal day [P] 14). Immunohistochemical analysis using anti-PV antibody was performed on the superior colliculus (SC) and lateral geniculate nucleus (LGN) at P45. In the visual cortex (VC) developmental changes in immunoreactivity were also examined at the ages of P17, P20, P27, and P45. Northern blot analysis for PV mRNA was also performed at P45. Changes in PV expression in the visual system of these rats were evaluated by use of a computer-based quantitative technique. RESULTS: PV-immunoreactive neurons were present in the SC and VC, whereas only a few were found in the LGN. The monocular enucleation at the onset of the sensitive period markedly reduced PV immunoreactivity in the neuropil of the SC, contralateral to the enucleated eye when examined one month later. No consistent and significant change in PV immunoreactivity was found in either the LGN or the VC. The number of PV-immunoreactive neurons in the VC rapidly decreased to the adult level during the middle of the sensitive period. The expression of PV mRNA in these central visual structures was not affected by early monocular enucleation. CONCLUSIONS: Expression of PV is developmentally regulated, and marked changes in its protein expression in the SC can be induced by monocular enucleation. Contrary to the original hypothesis, monocular enucleation did not consistently affect the expression of PV in the rat VC. The expression of PV is probably regulated by multiple factors, not merely by binocular competition.
Asunto(s)
Enucleación del Ojo , Regulación del Desarrollo de la Expresión Génica , Cuerpos Geniculados/crecimiento & desarrollo , Parvalbúminas/genética , Colículos Superiores/crecimiento & desarrollo , Visión Monocular/genética , Corteza Visual/crecimiento & desarrollo , Animales , Northern Blotting , Cuerpos Geniculados/metabolismo , Técnicas para Inmunoenzimas , Ácido Kaínico/farmacología , Neuronas/metabolismo , Parvalbúminas/metabolismo , ARN Mensajero/metabolismo , Ratas , Ratas Long-Evans , Colículos Superiores/metabolismo , Corteza Visual/metabolismoRESUMEN
A chondrogenic cell line, TC6, was established by using cells derived from articular cartilage of transgenic mice harboring a temperature-sensitive mutant simian virus (SV) 40 large T-antigen gene. TC6 cells express genes encoding proteins related to cartilage phenotypes such as type II collagen. To examine the in vivo behavior of the TC6 cells, these cells were implanted into cavity-shaped full-thickness defects made in the articular cartilage of the central part of the patellar grooves of mouse femora. One week after implantation, the morphology of the cells was still fibroblastic but these cells were just about to start to form a cartilage-like matrix. By 6 weeks after implantation, the cells had produced abundant cartilaginous matrix and their morphology became closer to that of authentic chondrocytes. This was in sharp contrast to the fibroblastic morphology of these cells in an in vitro environment even after long-term culture. These observations indicate that a cartilage-matrix environment provides a scaffold for the TC6 cells to form cartilage tissues. Our data show that the genetically engineered chondrocytic cell line, TC6, can form a cartilage-like matrix in vivo.
Asunto(s)
Antígenos Transformadores de Poliomavirus/farmacología , Cartílago Articular/virología , Diferenciación Celular , Animales , Supervivencia Celular , Trasplante de Células , Células Cultivadas , Condrocitos/metabolismo , Colágeno/metabolismo , Fibroblastos/metabolismo , Rodilla/anatomía & histología , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Mutación , TemperaturaRESUMEN
We investigated the expression pattern of two immediate-early genes, zif268 and c-fos, under various visual conditions using immunohistochemical and northern blot analysis in the visual cortex of young rats. The basal expression of c-fos was low and was further reduced by dark rearing that lasted for one week. A marked and transient increase was induced upon visual stimulation applied immediately after dark rearing. Zif268 showed a relatively high basal level. Its expression was reduced by dark rearing of the animals, but returned rapidly to the basal expression level following the introduction of light. Administration of N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine, a selective noradrenergic neurotoxin, suppressed the basal expression of c-fos messenger RNA. The response of c-fos to photo-stimulation was also significantly lower in the visual cortex of N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine-treated young rats. In contrast, no significant change in zif268 expression was detected between normal and N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine-treated animals. These findings suggest that differential expression of these immediate-early genes is involved in the activity-dependent regulation of cortical function. One possibility is that the noradrenergic system controls cortical function, including plasticity, by modifying the expression of c-fos.
Asunto(s)
Proteínas de Unión al ADN/genética , Genes Inmediatos-Precoces/fisiología , Genes fos/fisiología , Proteínas Inmediatas-Precoces , Proteínas Proto-Oncogénicas c-fos/metabolismo , Factores de Transcripción/genética , Corteza Visual , Factores de Edad , Animales , Bencilaminas/farmacología , Proteínas de Unión al ADN/efectos de los fármacos , Oscuridad/efectos adversos , Proteína 1 de la Respuesta de Crecimiento Precoz , Expresión Génica , Genes Inmediatos-Precoces/efectos de los fármacos , Genes fos/efectos de los fármacos , Masculino , Neurotoxinas/farmacología , Norepinefrina/genética , Norepinefrina/fisiología , Estimulación Luminosa , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Factores de Transcripción/efectos de los fármacos , Corteza Visual/efectos de los fármacos , Corteza Visual/metabolismoRESUMEN
Sensory experience in early life shapes the mammalian brain. An impairment in the activity-dependent refinement of functional connections within developing visual cortex was identified here in a mouse model. Gene-targeted disruption of one isoform of glutamic acid decarboxylase prevented the competitive loss of responsiveness to an eye briefly deprived of vision, without affecting cooperative mechanisms of synapse modification in vitro. Selective, use-dependent enhancement of fast intracortical inhibitory transmission with benzodiazepines restored plasticity in vivo, rescuing the genetic defect. Specific networks of inhibitory interneurons intrinsic to visual cortex may detect perturbations in sensory input to drive experience-dependent plasticity during development.
Asunto(s)
Glutamato Descarboxilasa/metabolismo , Interneuronas/fisiología , Plasticidad Neuronal , Corteza Visual/fisiología , Ácido gamma-Aminobutírico/metabolismo , Animales , Diazepam/farmacología , Moduladores del GABA/farmacología , Marcación de Gen , Glutamato Descarboxilasa/genética , Potenciación a Largo Plazo , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Plasticidad Neuronal/efectos de los fármacos , Estimulación Luminosa , Receptores de GABA-A/metabolismo , Transmisión Sináptica , Corteza Visual/citología , Corteza Visual/metabolismo , Vías VisualesRESUMEN
Scleraxis is a basic helix-loop-helix-type transcription factor that is expressed in sclerotome. Fibroblast growth factor (FGF) is one of the cytokines produced by the cells in skeletal tissues and is a potent modulator of skeletogenesis. The aim of this study was to examine the effects of FGF on the expression of scleraxis in chondrocyte-like cells, TC6. In these cells, scleraxis mRNA was constitutively expressed as a 1 .2 kb message at a high level in contrast to its low levels of expression in fibroblast-like cells or osteoblast-like cells. Upon treatment with FGF, scleraxis mRNA level was decreased within 12 h. This effect was at its nadir at 24 h and the scleraxis mRNA level returned to its base line level by 48 h. The FGF effect was maximal at 1 ng/ml. FGF effects on scleraxis were blocked by actinomycin D but not by cycloheximide, suggesting the involvement of transcriptional events that do not require new protein synthesis. The FGF effects on scleraxis were blocked by genistein, suggesting the involvement of tyrosine kinase in the post-receptor signaling. TGFbeta treatment of TC6 cells enhanced scleraxis mRNA expression; however, combination of the saturation doses of FGF and TGFbeta resulted in suppression of scleraxis mRNA level. BMP2 also suppressed scleraxis mRNA expression in TC6 cells and no further suppression was observed in combination with FGF. These results indicate that scleraxis is expressed in chondrocyte-like TC6 cells and it is one of the targets of FGF action in these cells.
Asunto(s)
Condrocitos/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Factores de Crecimiento de Fibroblastos/farmacología , Factores de Transcripción/genética , Factores de Transcripción con Motivo Hélice-Asa-Hélice Básico , Línea Celular , Condrocitos/citología , Secuencias Hélice-Asa-Hélice , Humanos , ARN Mensajero/genética , Receptores de Factores de Crecimiento de Fibroblastos/genéticaRESUMEN
We have re-examined whether N-methyl-D-aspartate receptors play a specific role in experience-dependent plasticity in kitten visual cortex. A specific antagonist of this glutamate receptor subtype, D,L-2-amino-5-phosphonovaleric acid, was directly and continuously infused into kitten striate cortex for one week concurrently with monocular lid suture. In the hemisphere infused with 50 mM antagonist, we found the usual shift in ocular dominance toward the open eye with only a few binocular cells remaining. The changes were accompanied by an extremely high incidence (38%) of abnormal cells lacking orientation selectivity across different ocular dominance groups. In kitten cortex infused with 10 mM antagonist concurrently with monocular deprivation for a week, recording from a drug-affected region near the infusion centre, we again found the U-shaped ocular dominance distribution with the high incidence of non-selective cells. In antagonist-infused, otherwise normal striate cortex of adult cats, we found that the proportion of binocular cells decreased by one-half in two cellular populations: one recorded during the continuous infusion of 10 mM antagonist under general anaesthesia and paralysis, and the other about two days after stopping the infusion. We also established that in vivo concentrations of chronically infused 10 mM antagonist decreased, not near-exponentially, but linearly with increasing distance from the infusion site. Thus, the effects of a directly and continuously infused, concentrated antagonist of N-methyl-D-aspartate receptors on receptive-field properties of visuocortical cells are complex. The present findings strongly suggest that the antagonist effects in the developing cortex may be due primarily to blockade of normal synaptic transmission rather than specific disruption of an experience-dependent mechanism underlying ocular dominance plasticity.
Asunto(s)
Dominancia Cerebral/fisiología , Plasticidad Neuronal/fisiología , Fenómenos Fisiológicos Oculares , Receptores de N-Metil-D-Aspartato/fisiología , Corteza Visual/crecimiento & desarrollo , Corteza Visual/fisiología , 2-Amino-5-fosfonovalerato/farmacocinética , 2-Amino-5-fosfonovalerato/farmacología , Animales , Gatos , Antagonistas de Aminoácidos Excitadores/farmacocinética , Antagonistas de Aminoácidos Excitadores/farmacología , Plasticidad Neuronal/efectos de los fármacos , Estimulación Luminosa , Receptores de N-Metil-D-Aspartato/antagonistas & inhibidores , Corteza Visual/efectos de los fármacos , Campos Visuales/efectos de los fármacosRESUMEN
We have recently demonstrated that some tenascin (TN) gene-knockout mice display abnormal behaviors, and that these abnormal behaviors stem from a low level of dopamine transmission in the brain. In the present study, we elucidated that tyrosine hydroxylase (TH) activity in the frontal cortex, striatum, and hippocampus of TN-knockout mice which showed abnormal behavior was significantly decreased. Also, the TH mRNA level of the midbrain was decreased by 43% in these animals compared with values for wild-type mice. These results suggest that the low dopamine turnover rate in some areas of the brain of TN-knockout mice accompanied by motor defects is due, at least in part, to the reduction in TH activity caused by diminished TH mRNA expression, and that TN-knockout mice exhibit abnormal behaviors in the presence of low levels of TH-gene expression.
Asunto(s)
Dopamina/metabolismo , Neuronas/enzimología , ARN Mensajero/genética , Tenascina/genética , Tirosina 3-Monooxigenasa/metabolismo , Animales , Northern Blotting , Mesencéfalo/enzimología , Ratones , Ratones Noqueados , Actividad Motora/genética , ARN Mensajero/metabolismo , Tirosina 3-Monooxigenasa/genéticaRESUMEN
Tenascin is a large extracellular matrix glycoprotein which is highly expressed in the developing nervous system. To examine the role of tenascin in vivo, we have produced mice in which the tenascin-gene is inactivated. These animals did not easily habituate to unfamiliar circumstances and displayed hyperlocomotion. A dopamine receptor agonist, apomorphine, reduced this hyperlocomotion dose dependently, but this phenomenon was not due to the appearance of apomorphine-induced stereotypic behavior, suggesting that tenascin-gene mutant mice have a paradoxical behavioral response to apomorphine compared to wild-type mice.
Asunto(s)
Apomorfina/farmacología , Agonistas de Dopamina/farmacología , Conducta Estereotipada/efectos de los fármacos , Tenascina/fisiología , Animales , Relación Dosis-Respuesta a Droga , Interacciones Farmacológicas , Locomoción/efectos de los fármacos , Locomoción/genética , Ratones , Ratones Noqueados , Especificidad de la Especie , Conducta Estereotipada/fisiología , Tenascina/genéticaRESUMEN
The aim of this study was to investigate the effect of cholecystokinin (CCK) receptor antagonist on the abnormal behavior and dopamine (DA) transmission of tenascin (TN)-gene knockout mice. Recently, we demonstrated that TN-gene deficient mice show hyperlocomotion that is related to reduced DA transmission and tyrosine hydroxylase (TH) activities in the brain. In this report, we show that the intraperitoneal administration of a CCK-B receptor antagonist, PD135158 (0.1 mg/kg), but not a CCK-A receptor antagonist, lorglumide, inhibited hyperlocomotion. Moreover, PD135158 reversed the low levels of DA turnover rate and TH activities in the striatum of TN-gene knockout mouse brain. These results suggest that CCK-B receptor is involved in the behavior of TN-gene knockout mouse through striatal DA transmission.
Asunto(s)
Ácido 3,4-Dihidroxifenilacético/metabolismo , Dopamina/metabolismo , Indoles/farmacología , Meglumina/análogos & derivados , Receptores de Colecistoquinina/antagonistas & inhibidores , Transmisión Sináptica/efectos de los fármacos , Tenascina/toxicidad , Animales , Hipercinesia/genética , Meglumina/farmacología , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Actividad Motora/efectos de los fármacos , Proglumida/análogos & derivados , Proglumida/farmacología , Receptor de Colecistoquinina A , Receptor de Colecistoquinina B , Tirosina 3-Monooxigenasa/metabolismoRESUMEN
Tissue-type plasminogen activator (tPA) plays important roles in the regulation of synaptic plasticity in the hippocampus and cerebellum. We found that the expression of tPA mRNA in the visual cortex was increased significantly by the peripheral administration of L-threo-3,4-dihydroxyphenylserine (L-threo-DOPS; 100 mg/kg, i.p.), which we had previously shown to have a promotive effect on ocular dominance (OD) plasticity. When plasminogen activator inhibitor-1 (PAI-1; 100 muM in an osmotic minipump) was infused into the kitten visual cortex, OD plasticity was suppressed; i.e. a significantly large number of binocular cells was recorded in the PAI-1 infused cortex following monocular deprivation. These results, therefore, suggest that the PA system is involved in the promotive effect of L-threo-DOPS in OD plasticity.
Asunto(s)
Antiparkinsonianos/farmacología , Droxidopa/farmacología , Plasticidad Neuronal/efectos de los fármacos , Activadores Plasminogénicos/genética , Activador de Tejido Plasminógeno/genética , Animales , Benserazida/farmacología , Northern Blotting , Gatos , Expresión Génica/efectos de los fármacos , Masculino , Plasticidad Neuronal/fisiología , Activadores Plasminogénicos/farmacología , ARN Mensajero/metabolismo , Ratas , Ratas Wistar , Privación Sensorial/fisiología , Activador de Tejido Plasminógeno/farmacología , Visión Monocular/efectos de los fármacos , Visión Monocular/fisiología , Corteza Visual/efectos de los fármacos , Corteza Visual/fisiología , Vías Visuales/fisiologíaRESUMEN
We established a clonal chondrocyte-like cell line (TC6, TC stands for large T immortalized chondrocyte-like cell line) derived from articular cartilage of transgenic mice harboring a temperature-sensitive simian virus 40 large T-antigen gene. TC6 cells exhibited spindle-like or polygonal morphology and grew well at 33 degrees C in alpha-minimal essential medium supplemented with 0.5% fetal bovine serum. After confluence, these cells formed nodules that were positive for staining with alcian blue. Northern blot analysis demonstrated that these cells expressed messenger RNAs (mRNA) of the genes encoding cartilage-specific proteins such as type II procollagen, link protein, and aggrecan. Furthermore, the expression of type II procollagen and link protein genes in TC6 cells was regulated by parathyroid hormone and basic fibroblast growth factor, suggesting the presence of the receptors for the hormone and cytokine. The expression of link protein mRNA in TC6 cells was regulated in a time-dependent manner and was enhanced in culture within a week and increased continuously up to 10-fold by the end of 4 weeks. Expression of mRNAs encoding type II procollagen and versican/PG-M also increased moderately during the culture period. TC6 cells expressed type I procollagen mRNA, however, its level declined along with time in culture in contrast to the enhancement of the genes encoding cartilage-specific molecules in these cells. Interestingly, alkaline phosphatase mRNA expression was barely detectable in the TC6 cells in their growing phase while it was enhanced dramatically more than 7-fold by day 14 in culture. These results indicate that the TC6 cells could serve as an excellent model for the studies on chondrocyte physiology.
Asunto(s)
Antígenos Transformadores de Poliomavirus/genética , Cartílago Articular/fisiología , Genes Virales , Animales , Cartílago Articular/citología , Diferenciación Celular/genética , Línea Celular , Células Clonales , Proteínas de la Matriz Extracelular/genética , Expresión Génica , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Fenotipo , ARN Mensajero/biosíntesis , TemperaturaRESUMEN
The regional difference in the expression of c-fos mRNA induced by typical and atypical antipsychotics was determined in prefrontal cortex, striatum, N. accumbens and lateral septum in rats by in situ hybridization. Two typical antipsychotics, haloperidol (2 mg/kg) and fluphenazine (2 mg/kg), and three atypical antipsychotics, (-)sulpiride (100 mg/kg), clozapine (20 mg/kg) and OPC-14597 (40 mg/kg), were used. Brains were fixed with 4% paraformaldehyde 45 min after drug administration (i.p.). Brain sections of 30 microns-thickness were made in a cryostat and hybridized with 35S-labelled for c-fos oligonucleotide probe. These sections were apposed to X-ray films and the autoradiograms were semi-quantitatively analysed by computer-assisted densitometry. All antipsychotics used increased c-fos mRNA expression in N. accumbens shell, a region of the forebrain associated with limbic systems. On the other hand, two typical antipsychotics (haloperidol and fluphenazine) that cause a high incidence of acute motor side effects increased the expression of c-fos mRNA in the dorsolateral striatum, an extrapyramidal region primarily involved in motor control. Only clozapine induced c-fos mRNA in the medial prefrontal cortex and lateral septum. These results strongly suggest that the shell region of N. accumbens may be a common site of therapeutic action of antipsychotics.
Asunto(s)
Antipsicóticos/farmacología , Encéfalo/metabolismo , Cuerpo Estriado/metabolismo , Genes fos/genética , ARN Mensajero/metabolismo , Animales , Aripiprazol , Clozapina/farmacología , Flufenazina/farmacología , Haloperidol/farmacología , Hibridación in Situ , Núcleo Accumbens/metabolismo , Piperazinas/farmacología , Corteza Prefrontal/metabolismo , Quinolonas/farmacología , Ratas , Tabique Pelúcido/metabolismo , Sulpirida/farmacologíaRESUMEN
NCB-20 cells expressed m1- and m4-muscarinic acetylcholine receptor (mAChR) mRNAs, while NG108-15 cells expressed only m4-mAChR mRNA. Butyrate induced a time-dependent increase in the level of m1-mAChR mRNA with no change in the m4-mAChR mRNA level in NCB-20 cells. Similarly, butyrate did not affect the m4-mAChR mRNA level in NG108-15 cells. In contrast, dibutyryl cAMP caused a significant time-dependent decrease in the level of m4-mAChR mRNA in NCB-20 and NG108-15 cells as well as m1-mAChR mRNA in NCB-20 cells. Our results suggest that these two differentiating agents are important physiological regulators of the transcription and/or stability of the mRNA of certain mAChR subtypes expressed in these two neurohybrid cell lines.
Asunto(s)
Bucladesina/farmacología , Butiratos/farmacología , Células Híbridas/efectos de los fármacos , Receptores Muscarínicos/genética , Animales , Northern Blotting , Ácido Butírico , Sondas de ADN , Regulación hacia Abajo/fisiología , Expresión Génica/efectos de los fármacos , Células Híbridas/química , Células Híbridas/fisiología , Ratones , Neuroblastoma , ARN Mensajero/metabolismo , Células Tumorales Cultivadas/química , Células Tumorales Cultivadas/efectos de los fármacos , Células Tumorales Cultivadas/fisiología , Regulación hacia Arriba/fisiologíaRESUMEN
To examine the role of tenascin (TN) in vivo, we have produced mice in which the TN gene is inactivated. In behavioral studies, TN-knockout mice showed abnormal behavior such as hyperlocomotion and poor swimming ability. Biochemical analysis revealed that serotonin (5-HT) and dopamine (DA) transmission was decreased in the cerebral cortex, the hippocampus, or the striatum of TN-knockout mouse brain. The intraperitoneal administration of the DA receptor agonist, LY171555 (0.5 mg/kg, BW), inhibited the hyperlocomotion, and swimming behavior was transiently improved by the treatment with the 5-HT receptor agonist, 1-(4-iodo-2,5-dimethoxyphenyl)-2-aminopropane hydrochloride. These findings suggest that TN may play an important role in neurotransmissions related to behavior.
Asunto(s)
Conducta Animal/efectos de los fármacos , Neurotransmisores/metabolismo , Tenascina/genética , Animales , Dopaminérgicos/metabolismo , Dopaminérgicos/farmacología , Ratones , Ratones Noqueados , Actividad Motora/efectos de los fármacos , Ensayo de Unión Radioligante , Serotoninérgicos/metabolismo , Serotoninérgicos/farmacologíaRESUMEN
A system for positron emission tomography study of conscious monkeys was newly developed. By use of this system in combination with a microdialysis technique, the effect of ketamine on the binding and release of dopamine was investigated. The administration of ketamine (5 mg/kg) caused sedation accompanied by psychotic symptoms such as nystagmus and stereotyped movements of extremities. During this psychotomimetic period produced by ketamine, a significant increase in the accumulation of the dopamine D2 receptor ligand N-[11C]methylspiperone was observed in the striatum compared with the level in the conscious state, while no significant change was observed in the frontal cortex and cerebellum. In contrast to the use of ketamine as the anesthetic, pentobarbital (25 mg/kg), which produced deeper anesthesia but no psychotic symptoms, caused a decrease in the accumulation of N-[11C]methylspiperone in the striatum. Kinetic analysis, conducted by a graphical method, revealed that the value of the association constant (K3) for N-[11C]methylspiperone binding in the striatum was increased to approximately 130% by ketamine and decreased to approximately 70% by pentobarbital compared with the control values. Furthermore, the release of dopamine from the striatum measured by microdialysis was not affected by ketamine anesthesia. These results indicate that ketamine facilitates striatal dopaminergic neurotransmission through increasing the binding activity of dopamine D2 receptors in the striatum, and suggest that these changes may be related to the psychotomimetic behavioral symptoms of this drug.
Asunto(s)
Cuerpo Estriado/efectos de los fármacos , Dopamina/fisiología , Ketamina/farmacología , Espiperona/análogos & derivados , Animales , Conducta Animal/efectos de los fármacos , Radioisótopos de Carbono , Cuerpo Estriado/metabolismo , Macaca mulatta , Masculino , Microdiálisis , Ensayo de Unión Radioligante , Espiperona/metabolismo , Tomografía Computarizada de EmisiónRESUMEN
A mode of internal motion of single tryptophan, Trp 86, of Streptomyces subtilisin inhibitor, was analyzed from its time-resolved fluorescence. The intensity and anisotropy decays of Trp 86 were measured in the picosecond range. These decays were analyzed with theoretical expressions derived assuming that the indole ring of tryptophan as an asymmetric rotor rotates around covalent bonds connecting indole with the peptide chain and an effective quencher of fluorescence of Trp 86 is the nearby SS bond of Cys 35-Cys 50. First, the intensity decays at 6 degrees, 20 degrees, and 40 degrees C were analyzed, and then the both decays of the intensity and anisotropy at 20 degrees C were simultaneously simulated with common parameters. Constants concerning geometrical structures of the protein used for the analysis were obtained from x-ray crystallographic data. Best fit between the observed and calculated decay curves was obtained by a nonlinear least squares method by adjusting a quenching constant averaged over the rotational angles, koq height of the potential energy, p, and three of six diffusion coefficients, Dxx, Dyy, Dzz, Dxy, Dyz, and Dzx, as variable parameters. The obtained results revealed that the internal motion of the indole ring became faster, the quenching rate of the fluorescence of Trp 86 was enhanced and the height of potential energy became lower at higher temperatures, and suggested that Trp 86 was wobbling around the long axis of the indole ring in the protein.
Asunto(s)
Proteínas Bacterianas/química , Conformación Proteica , Inhibidores de Serina Proteinasa/química , Triptófano , Secuencia de Aminoácidos , Proteínas Bacterianas/aislamiento & purificación , Polarización de Fluorescencia , Cinética , Modelos Moleculares , Espectrometría de Fluorescencia , StreptomycesRESUMEN
The locomotor activity and grooming of conscious freely moving rats were recorded during a 60-min unilateral perfusion of the preoptic area with neuroactive compounds using the microdialysis technique. The GABA agonist, muscimol (10, 20 and 100 microM) induced a dose-dependent increase in locomotor activity and grooming which was attenuated by co-perfusion with the GABA antagonist, bicuculline (10 microM), and was blocked by systemic injection of haloperidol, a preferential dopamine D2 receptor antagonist (0.25 mg/kg). Muscimol-induced hyperactivity was associated with a simultaneous increase of striatal extracellular dopamine. These data suggest that the preoptic area is functionally linked with the extrapyramidal dopaminergic system possibly via GABAergic system.