RESUMEN
Recent evidence underlines the crucial role of neuronal cytoskeleton in the pathophysiology of psychiatric diseases. In this line, the deletion of STOP/MAP6 (Stable Tubule Only Polypeptide), a microtubule-stabilizing protein, triggers various neurotransmission and behavioral defects, suggesting that STOP knockout (KO) mice could be a relevant experimental model for schizoaffective symptoms. To establish the predictive validity of such a mouse line, in which the brain serotonergic tone is dramatically imbalanced, the effects of a chronic fluoxetine treatment on the mood status of STOP KO mice were characterized. Moreover, we determined the impact, on mood, of a chronic treatment by epothilone D, a taxol-like microtubule-stabilizing compound that has previously been shown to improve the synaptic plasticity deficits of STOP KO mice. We demonstrated that chronic fluoxetine was either antidepressive and anxiolytic, or pro-depressive and anxiogenic, depending on the paradigm used to test treated mutant mice. Furthermore, control-treated STOP KO mice exhibited paradoxical behaviors, compared with their clear-cut basal mood status. Paradoxical fluoxetine effects and control-treated STOP KO behaviors could be because of their hyper-reactivity to acute and chronic stress. Interestingly, both epothilone D and fluoxetine chronic treatments improved the short-term memory of STOP KO mice. Such treatments did not affect the serotonin and norepinephrine transporter densities in cerebral areas of mice. Altogether, these data demonstrated that STOP KO mice could represent a useful model to study the relationship between cytoskeleton, mood, and stress, and to test innovative mood treatments, such as microtubule-stabilizing compounds.
Asunto(s)
Afecto/efectos de los fármacos , Epotilonas/farmacología , Fluoxetina/farmacología , Memoria a Corto Plazo/efectos de los fármacos , Proteínas Asociadas a Microtúbulos/deficiencia , Proteínas Asociadas a Microtúbulos/genética , Afecto/fisiología , Animales , Antineoplásicos/farmacología , Depresión/inducido químicamente , Depresión/genética , Depresión/fisiopatología , Modelos Animales de Enfermedad , Femenino , Masculino , Memoria a Corto Plazo/fisiología , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos C57BL , Ratones Noqueados , Inhibidores Selectivos de la Recaptación de Serotonina/farmacologíaRESUMEN
The microtubule-associated Stable Tubulie Only Polypeptide (STOP; also known as MAP6) protein plays a key role in neuron architecture and synaptic plasticity, the dysfunctions of which are thought to be implicated in the pathophysiology of psychiatric diseases. The deletion of STOP in mice leads to severe disorders reminiscent of several schizophrenia-like symptoms, which are also associated with differential alterations of the serotonergic tone in somas versus terminals. In STOP knockout (KO) compared with wild-type mice, serotonin (5-HT) markers are found to be markedly accumulated in the raphe nuclei and, in contrast, deeply depleted in all serotonergic projection areas. In the present study, we carefully examined whether the 5-HT imbalance would lead to behavioral consequences evocative of mood and/or cognitive disorders. We showed that STOP KO mice exhibited depression-like behavior, associated with a decreased anxiety-status in validated paradigms. In addition, although STOP KO mice had a preserved very short-term memory, they failed to perform well in all other learning and memory tasks. We also showed that STOP KO mice exhibited regional imbalance of the norepinephrine tone as observed for 5-HT. As a consequence, mutant mice were hypersensitive to acute antidepressants with different selectivity. Altogether, these data indicate that the deletion of STOP protein in mice caused deep alterations in mood and cognitive performances and that STOP protein might have a crucial role in the 5-HT and norepinephrine networks development.
Asunto(s)
Trastornos del Conocimiento/genética , Eliminación de Gen , Proteínas Asociadas a Microtúbulos/deficiencia , Proteínas Asociadas a Microtúbulos/genética , Trastornos del Humor/genética , Proteínas del Tejido Nervioso/deficiencia , Proteínas del Tejido Nervioso/genética , Animales , Conducta Animal/fisiología , Trastornos del Conocimiento/metabolismo , Trastornos del Conocimiento/psicología , Femenino , Masculino , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/fisiología , Trastornos del Humor/metabolismo , Trastornos del Humor/psicología , Proteínas del Tejido Nervioso/fisiologíaRESUMEN
Triadin is a multiple proteins family, some isoforms being involved in muscle excitation-contraction coupling, and some having still unknown functions. To obtain clues on triadin functions, we engineered a triadin knock-out mouse line and characterized the physiological effect of triadin ablation on skeletal muscle function. These mice presented a reduced muscle strength, which seemed not to alter their survival and has been characterized in the present work. We first checked in these mice the expression level of the different proteins involved in calcium homeostasis and observed in fast muscles an increase in expression of dihydropyridine receptor, with a large reduction in calsequestrin expression. Electron microscopy analysis of KO muscles morphology demonstrated the presence of triads in abnormal orientation and a reduction in the sarcoplasmic reticulum terminal cisternae volume. Using calcium imaging on cultured myotubes, we observed a reduction in the total amount of calcium stored in the sarcoplasmic reticulum. Physiological studies have been performed to evaluate the influence of triadin deletion on skeletal muscle function. Muscle strength has been measured both on the whole animal model, using hang test or electrical stimulation combined with NMR analysis and strength measurement, or on isolated muscle using electrical stimulation. All the results obtained demonstrate an important reduction in muscle strength, indicating that triadin plays an essential role in skeletal muscle function and in skeletal muscle structure. These results indicate that triadin alteration leads to the development of a myopathy, which could be studied using this new animal model.
Asunto(s)
Proteínas Portadoras , Eliminación de Gen , Proteínas Musculares , Músculo Esquelético/fisiología , Animales , Conducta Animal/fisiología , Calcio/metabolismo , Proteínas Portadoras/genética , Proteínas Portadoras/metabolismo , Células Cultivadas , Femenino , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Contracción Muscular/fisiología , Proteínas Musculares/genética , Proteínas Musculares/metabolismo , Músculo Esquelético/citología , Músculo Esquelético/ultraestructura , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Canal Liberador de Calcio Receptor de Rianodina/genética , Canal Liberador de Calcio Receptor de Rianodina/metabolismoRESUMEN
The deletion of microtubule-associated protein stable tubule only polypeptide (STOP) leads to neuroanatomical, biochemical and severe behavioral alterations in mice, partly alleviated by antipsychotics. Therefore, STOP knockout (KO) mice have been proposed as a model of some schizophrenia-like symptoms. Preliminary data showed decreased brain serotonin (5-HT) tissue levels in STOP KO mice. As literature data demonstrate various interactions between microtubule-associated proteins and 5-HT, we characterized some features of the serotonergic neurotransmission in STOP KO mice. In the brainstem, mutant mice displayed higher tissue 5-HT levels and in vivo synthesis rate, together with marked increases in 5-HT transporter densities and 5-HT1A autoreceptor levels and electrophysiological sensitivity, without modification of the serotonergic soma number. Conversely, in projection areas, STOP KO mice exhibited lower 5-HT levels and in vivo synthesis rate, associated with severe decreases in 5-HT transporter densities, possibly related to reduced serotonergic terminals. Mutant mice also displayed a deficit of adult hippocampal neurogenesis, probably related to both STOP deletion and 5-HT depletion. Finally, STOP KO mice exhibited a reduced anxiety- and, probably, an increased helpness-status, that could be because of the strong imbalance of the serotonin neurotransmission between somas and terminals. Altogether, these data suggested that STOP deletion elicited peculiar 5-HT disconnectivity.
Asunto(s)
Encéfalo/fisiología , Proteínas Asociadas a Microtúbulos/deficiencia , Proteínas Asociadas a Microtúbulos/genética , Red Nerviosa/fisiología , Serotonina/metabolismo , Animales , Femenino , Masculino , Aprendizaje por Laberinto/fisiología , Ratones , Ratones de la Cepa 129 , Ratones Endogámicos BALB C , Ratones NoqueadosRESUMEN
BACKGROUND: Mice deficient for the stable tubule only peptide (STOP) display altered dopaminergic neurotransmission associated with severe behavioural defects including disorganized locomotor activity. Endogenous morphine, which is present in nervous tissues and synthesized from dopamine, may contribute to these behavioral alterations since it is thought to play a role in normal and pathological neurotransmission. RESULTS: In this study, we showed that STOP null brain structures, including cortex, hippocampus, cerebellum and spinal cord, contain high endogenous morphine amounts. The presence of elevated levels of morphine was associated with the presence of a higher density of mu opioid receptor with a higher affinity for morphine in STOP null brains. Interestingly, STOP null mice exhibited significantly lower nociceptive thresholds to thermal and mechanical stimulations. They also had abnormal behavioural responses to the administration of exogenous morphine and naloxone. Low dose of morphine (1 mg/kg, i.p.) produced a significant mechanical antinociception in STOP null mice whereas it has no effect on wild-type mice. High concentration of naloxone (1 mg/kg) was pronociceptive for both mice strain, a lower concentration (0.1 mg/kg) was found to increase the mean mechanical nociceptive threshold only in the case of STOP null mice. CONCLUSIONS: Together, our data show that STOP null mice displayed elevated levels of endogenous morphine, as well as an increase of morphine receptor affinity and density in brain. This was correlated with hypernociception and impaired pharmacological sensitivity to mu opioid receptor ligands.
Asunto(s)
Proteínas Asociadas a Microtúbulos/deficiencia , Morfina/farmacología , Dolor/fisiopatología , Analgésicos Opioides , Animales , Ratones , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/fisiología , Morfina/análisis , Naloxona/administración & dosificación , Naloxona/farmacología , Antagonistas de Narcóticos , Proteínas del Tejido Nervioso , Alcaloides Opiáceos , Dolor/tratamiento farmacológico , Receptores Opioides mu/análisis , Receptores Opioides mu/metabolismoRESUMEN
Regulation of microtubule dynamics is crucial for axon growth and guidance as well as for the establishment of synaptic connections. STOPs (Stable Tubule Only Polypeptides) are microtubule-associated proteins that regulate microtubule stabilization but are also able to interact with actin or Golgi membranes. Here, we have investigated the involvement of STOPs during the development of the olfactory system. We first describe the spatio-temporal expression patterns of N- and E-STOP, the two neuronal-specific isoforms of STOP. E- and N-STOP are expressed in the axonal compartment of olfactory sensory neurons, but are differentially regulated during development. Interestingly, each neuronal isoform displays a specific gradient distribution within the olfactory nerve layer. Then, we have examined the development of the olfactory system in the absence of STOPs. Olfactory axons display a normal outgrowth and targeting in STOP-null mice, but maturation of the synapses in the glomerular neuropil is altered.
Asunto(s)
Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Vías Olfatorias/fisiología , Animales , Femenino , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/genética , Proteínas del Tejido Nervioso/genética , Neuronas Receptoras Olfatorias/citología , Neuronas Receptoras Olfatorias/fisiología , Embarazo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Ratas , Ratas Wistar , Técnicas de Cultivo de Tejidos , Órgano Vomeronasal/citología , Órgano Vomeronasal/fisiologíaRESUMEN
The microtubule-associated stable tubule only polypeptide (STOP) protein plays a key-role in neuron architecture and synaptic plasticity. Recent studies suggest that schizophrenia is associated with alterations in the synaptic connectivity. Mice invalidated for the STOP gene display phenotype reminiscent of some schizophrenic-like symptoms, such as behavioral disturbances, dopamine (DA) hyper-reactivity, and possible hypoglutamatergia, partly improved by antipsychotic treatment. In the present work, we examined potential alterations in some DAergic key proteins and behaviors in STOP knockout mice. Whereas the densities of the DA transporter, the vesicular monoamine transporter and the D(1) receptor were not modified, the densities of the D(2) and D(3) receptors were decreased in some DAergic regions in mutant versus wild-type mice. Endogenous DA levels were selectively decreased in DAergic terminals areas, although the in vivo DA synthesis was diminished both in cell bodies and terminal areas. The DA uptake was decreased in accumbic synaptosomes, but not significantly altered in striatal synaptosomes. Finally, STOP knockout mice were hypersensitive to acute and subchronic locomotor effects of cocaine, although the drug equally inhibited DA uptake in mutant and wild-type mice. Altogether, these data showed that deletion of the ubiquitous STOP protein elicited restricted alterations in DAergic neurotransmission, preferentially in the meso-limbic pathway.
Asunto(s)
Dopamina/metabolismo , Proteínas Asociadas a Microtúbulos/deficiencia , Receptores Dopaminérgicos/metabolismo , Transmisión Sináptica/fisiología , Análisis de Varianza , Animales , Ganglios Basales/metabolismo , Cocaína/farmacología , Inhibidores de Captación de Dopamina/farmacología , Relación Dosis-Respuesta a Droga , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Actividad Motora/efectos de los fármacos , Actividad Motora/genética , Radiografía , Transmisión Sináptica/efectos de los fármacos , Sinaptosomas/metabolismo , Tirosina 3-Monooxigenasa/metabolismo , Proteínas de Transporte Vesicular de Monoaminas/metabolismoRESUMEN
In mice, deletion of the STOP protein leads to subtle anatomic changes and induces depleted synaptic vesicle pools, impaired synaptic plasticity, hyperdopaminergy, and major behavioral disorders alleviated by neuroleptics, hence leading to a schizophrenic-like phenotype. In this study, we applied the quantitative autoradiographic [(14)C]2-deoxyglucose technique to study to what extent the basal rate of cerebral glucose utilization in STOP-knockout (STOP-KO) mice occurs in regions where metabolic changes have been reported in schizophrenic patients. Studies were performed on wild-type, heterozygous, and homozygous STOP-KO mice (7-8 per group). Mice were implanted with femoral artery and vein catheters, and cerebral glucose utilization was quantified over 45 min. Compared with that in wild-type mice, glucose utilization in STOP-KO mice was significantly increased in the olfactory cortex, ventromedial and anterolateral hypothalamus, ventral tegmental area, and substantia nigra pars compacta. Nonsignificant increases, ranging between 9% and 19%, were recorded in the whole auditory system, CA1 pyramidal cell layer, and dorsal raphe. Glucose utilization was also significantly increased in heterozygous mice compared with that in wild-type mice in olfactory cortex. These data might reflect hyperdopaminergic activity, olfactory deficits, and sleep disturbances in STOP-KO mice that have also been reported in schizophrenic patients.
Asunto(s)
Encéfalo/metabolismo , Proteínas Asociadas a Microtúbulos/metabolismo , Animales , Autorradiografía , Glucosa/metabolismo , Masculino , Ratones , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/genéticaRESUMEN
Mice deficient in the microtubule stabilizing protein STOP (stable tubule only polypeptide) show synaptic plasticity anomalies in hippocampus, dopamine hyper-reactivity in the limbic system and severe behavioral deficits. Some of these disturbances are alleviated by long-term antipsychotic treatment. Therefore, this mouse line represents a pertinent model for some aspects of schizophrenia symptomatology. Numerous data support dysfunction of nicotinic neurotransmission in schizophrenia and epidemiological studies show increased tobacco use in schizophrenic patients, in whom nicotine has been reported to improve cognitive deficits and impairment in sensory gating. In this study, we examined potential alterations in cholinergic (ACh) and nicotinic components and functions in STOP mutant mice. STOP KO mice displayed no variation of the density of ACh esterase and beta2* nicotinic receptors (nAChRs), large reductions in the density of vesicular ACh transporter and alpha6* nAChRs and marked increases in the density of alpha7 nAChRs, in some brain areas. STOP KO mice were hypersensitive to the stimulating locomotor effect of nicotine and, interestingly, their impaired performance in learning the cued version of the water maze were improved by administration of the preferential alpha7 nAChR agonist choline. Altogether, our data show that the deletion of the ubiquitous STOP protein elicited restricted alterations in ACh components. They also suggest that nicotinic neurotransmission can be deficient in STOP KO mice and that mutant mice can represent a meaningful model to study some nicotinic dysfunctions and therapeutic treatments.
Asunto(s)
Colina/uso terapéutico , Regulación de la Expresión Génica/genética , Discapacidades para el Aprendizaje , Proteínas Asociadas a Microtúbulos/deficiencia , Nootrópicos/uso terapéutico , Receptores Nicotínicos/metabolismo , Acetilcolinesterasa/metabolismo , Análisis de Varianza , Animales , Autorradiografía , Conducta Animal , Relación Dosis-Respuesta a Droga , Discapacidades para el Aprendizaje/tratamiento farmacológico , Discapacidades para el Aprendizaje/genética , Discapacidades para el Aprendizaje/metabolismo , Aprendizaje por Laberinto/efectos de los fármacos , Aprendizaje por Laberinto/fisiología , Ratones , Ratones Noqueados , Actividad Motora/efectos de los fármacos , Actividad Motora/genética , Tiempo de Reacción/efectos de los fármacos , Receptores Nicotínicos/genética , Proteínas de Transporte Vesicular de Acetilcolina/metabolismo , Receptor Nicotínico de Acetilcolina alfa 7RESUMEN
BACKGROUND: Recent data suggest that cytoskeletal defects may play a role in schizophrenia. We previously imitated features of schizophrenia in an animal model by disrupting gene coding for a microtubule-associated protein called STOP. STOP-null mice display synaptic defects in glutamatergic neurons, hyper-dopaminergy, and severe behavioral disorders. Synaptic and behavioral deficits are amended by neuroleptic treatment in STOP-null mice, providing an attractive model to test new antipsychotic agents. We examined the effects of a taxol-related microtubule stabilizer, epothilone D. METHODS: Mice were treated either with vehicle alone or with epothilone D. Treatment effects on synaptic function were assessed using electron-microscopy quantification of synaptic vesicle pools and electrophysiology in the CA1 region of the hippocampus. Dopamine transmission was investigated using electrochemical assays. Behavior was principally assessed using tests of maternal skills. RESULTS: In STOP-null mice, treatment with epothilone D increased synaptic vesicle pools, ameliorated both short- and long-term forms of synaptic plasticity in glutamatergic neurons, and had a dramatic beneficial effect on mouse behavior. CONCLUSIONS: A microtubule stabilizer can have a beneficial effect on synaptic function and behavior, suggesting new possibilities for treatment of schizophrenia.
Asunto(s)
Conducta Animal/efectos de los fármacos , Epotilonas/administración & dosificación , Neuronas/efectos de los fármacos , Esquizofrenia , Transmisión Sináptica/efectos de los fármacos , Moduladores de Tubulina/administración & dosificación , Animales , Conducta Animal/fisiología , Células Cultivadas , Modelos Animales de Enfermedad , Estimulación Eléctrica/métodos , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Conducta Exploratoria/efectos de los fármacos , Femenino , Hipocampo/patología , Masculino , Conducta Materna/efectos de los fármacos , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/deficiencia , Plasticidad Neuronal/efectos de los fármacos , Plasticidad Neuronal/fisiología , Plasticidad Neuronal/efectos de la radiación , Esquizofrenia/tratamiento farmacológico , Esquizofrenia/patología , Esquizofrenia/fisiopatología , Transmisión Sináptica/fisiologíaRESUMEN
INTRODUCTION: Recent studies have suggested that schizophrenia is associated with alterations in the synaptic connectivity involving cytoskeletal proteins. The microtubule-associated protein stable tubule only polypeptide (STOP) plays a key role in neuronal architecture and synaptic plasticity, and it has been demonstrated that STOP gene deletion in mice leads to a phenotype mimicking aspects of positive and negative symptoms and cognitive deficits classically observed in schizophrenic patients. In STOP null mice, behavioral defects are associated with synaptic plasticity abnormalities including defects in long-term potentiation. In these mice, long-term administration of typical antipsychotics has been shown to partially alleviate behavioral defects but, as in humans, such a treatment was poorly active on deficits related to negative symptoms and cognitive impairments. Here, we assessed the effects of risperidone and clozapine, two atypical antipsychotics, on STOP null mice behavior and synaptic plasticity. RESULTS: Long-term administration of either drug results in alleviation of behavioral alterations mimicking some negative symptoms and partial amelioration of some cognitive defects in STOP null mice. Interestingly, clozapine treatment also improves synaptic plasticity of the STOP null animals by restoring long-term potentiation in the hippocampus. DISCUSSION: All together, the pharmacological reactivity of STOP null mice to antipsychotics evokes the pharmacological response of humans to such drugs. Totally, our study suggests that STOP null mice may provide a useful preclinical model to evaluate pharmacological properties of antipsychotic drugs.
Asunto(s)
Antipsicóticos/farmacología , Clozapina/farmacología , Plasticidad Neuronal/efectos de los fármacos , Risperidona/farmacología , Animales , Conducta Animal/efectos de los fármacos , Evaluación Preclínica de Medicamentos/métodos , Hipocampo/efectos de los fármacos , Hipocampo/metabolismo , Masculino , Ratones , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/genéticaRESUMEN
BACKGROUND: STOP (Stable Tubulin-Only Polypeptide) null mice show behavioral deficits, impaired synaptic plasticity, decrease in synaptic vesicular pools and disturbances in dopaminergic transmission, and are considered a neurodevelopmental model of schizophrenia. Olfactory neurons highly express STOP protein and are continually generated throughout life. Experimentally-induced loss of olfactory neurons leads to epithelial regeneration within two months, providing a useful model to evaluate the role played by STOP protein in adult olfactory neurogenesis. METHODOLOGY/PRINCIPAL FINDINGS: Immunocytochemistry and electron microscopy were used to study the structure of the glomerulus in the main olfactory bulb and neurogenesis in the neurosensorial epithelia. In STOP null mice, olfactory neurons showed presynaptic swellings with tubulovesicular profiles and autophagic-like structures. In olfactory and vomeronasal epithelia, there was an increase in neurons turnover, as shown by the increase in number of proliferating, apoptotic and immature cells with no changes in the number of mature neurons. Similar alterations in peripheral olfactory neurogenesis have been previously described in schizophrenia patients. In STOP null mice, regeneration of the olfactory epithelium did not modify these anomalies; moreover, regeneration resulted in abnormal organisation of olfactory terminals within the olfactory glomeruli in STOP null mice. CONCLUSIONS/SIGNIFICANCE: In conclusion, STOP protein seems to be involved in the establishment of synapses in the olfactory glomerulus. Our results indicate that the olfactory system of STOP null mice is a well-suited experimental model (1) for the study of the mechanism of action of STOP protein in synaptic function/plasticity and (2) for pathophysiological studies of the mechanisms of altered neuronal connections in schizophrenia.
Asunto(s)
Proteínas Asociadas a Microtúbulos/deficiencia , Neurogénesis , Neuronas Receptoras Olfatorias/metabolismo , Animales , Modelos Animales de Enfermedad , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/genética , Bulbo Olfatorio/citología , Bulbo Olfatorio/metabolismo , Mucosa Olfatoria/inervación , Mucosa Olfatoria/metabolismo , Esquizofrenia/genética , Esquizofrenia/metabolismo , Sinapsis/metabolismoRESUMEN
In mice, the deletion of the STOP protein leads to hyperdopaminergia and major behavioral disorders that are alleviated by neuroleptics, representing a potential model of schizophrenia. The reduction of the glutamatergic synaptic vesicle pool in the hippocampus could reflect a disturbance in glutamatergic neurotransmission in this model. Here we examined potential disturbances in energy metabolism and interactions between neurons and glia in 15-week-old STOP KO, wild-type, and heterozygous mice. Animals received [1-(13)C]glucose and [1,2-(13)C]acetate, the preferential substrates of neurons and astrocytes, respectively. Extracts from the whole forebrain and midbrain were analyzed by HPLC, (13)C and (1)H NMR spectroscopy. Amounts and labeling of most metabolites were unchanged. However, glutamine concentration and amount of [4,5-(13)C]glutamine derived from [1,2-(13)C]acetate significantly decreased by 17% and 18%, respectively, in STOP KO compared with wild-type mice. The amount of [4-(13)C]glutamate was decreased in STOP KO and heterozygous compared with wild-type mice. gamma-Aminobutyric acid labeling was not influenced by the genotype. Because STOP-deficient mice have a lower synaptic vesicle density, less glutamate is released to the synaptic cleft, leading to decreased stimulation of the postsynaptic glutamate receptors, reflecting increased glutamine metabolism only in the vicinity of the postsynapse of STOP KO mice.
Asunto(s)
Encéfalo/metabolismo , Modelos Animales de Enfermedad , Ácido Glutámico/deficiencia , Proteínas Asociadas a Microtúbulos/genética , Esquizofrenia/fisiopatología , Animales , Astrocitos/metabolismo , Química Encefálica , Cromatografía Líquida de Alta Presión , Enfermedad Crónica , Dopamina/deficiencia , Metabolismo Energético , Glutamina/deficiencia , Espectroscopía de Resonancia Magnética , Masculino , Ratones , Ratones Noqueados , Neuronas/metabolismo , Esquizofrenia/metabolismo , Vesículas Sinápticas/metabolismoRESUMEN
Stable tubulin-only polypeptide (STOP) proteins are microtubule-associated proteins responsible for microtubule stabilization in neurons. STOP null mice show apparently normal cerebral anatomy but display synaptic defects associated with neuroleptic-sensitive behavioral disorders. STOP null mice have therefore been proposed as an animal model for the study of schizophrenia. In the present study, the expression pattern of STOP gene in developing and adult brain has been examined by using lacZ gene inserted in the STOP locus, as a reporter gene. beta-Galactosidase (beta-gal) immunostaining was confined to neuronal cells and projections. Strong labeling was observed in the whole olfactory system, cortical layer VII, hippocampus, hypothalamus, cerebellum, habenula, fasciculus retroflexus, and interpeduncular nucleus in adults. Additionally, ventral thalamic nucleus, clusters of positive cells in striatum, and Cajal-Retzius cells of cortical layer I were labeled in young mice. The strong expression of STOP lacZ reporter gene observed in brain is confined to areas that may be involved in the schizophrenia-related symptoms observed in STOP-deficient mice.
Asunto(s)
Encéfalo/metabolismo , Regulación del Desarrollo de la Expresión Génica , Proteínas Asociadas a Microtúbulos/metabolismo , Esquizofrenia/metabolismo , Animales , Encéfalo/embriología , Encéfalo/crecimiento & desarrollo , Genes Reporteros , Inmunohistoquímica , Análisis por Apareamiento , Ratones , Ratones Endogámicos BALB C , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/genética , Proteínas del Tejido Nervioso/genética , Proteínas del Tejido Nervioso/metabolismo , Distribución Tisular , beta-Galactosidasa/genética , beta-Galactosidasa/metabolismoRESUMEN
Overt schizophrenia is preceded by a prodromal phase during which juvenile patients display attenuated schizophrenia-related symptoms. Here, we have looked for evidence of a prodromal phase in juvenile STOP null mice, which, during adulthood, imitate features of schizophrenia. We have principally examined locomotor activity, which is abnormal in adult STOP null mice, and its apparent relationship with perturbed glutamatergic and dopaminergic transmission. When compared to corresponding wild-type mice, juvenile STOP null mice did not exhibit the basal hyperlocomotion or locomotor hypersensitivity to mild stress observed in adult mice. Juvenile STOP null mice also lacked disturbed locomotor sensitivity to MK-801, which was evident in adult mice. In contrast, juvenile STOP null mice exhibited a similar hypersensitivity to amphetamine as that found in adult mice. Thus, STOP null mice exhibited both a progression of locomotor activity defects over time and subtle alterations in the prepubertal period. We suggest that the pattern of locomotor disturbances observed in this study is related to altered dopaminergic reactivity in juvenile mice without major disturbance in glutamatergic transmission, whereas both neurotransmitter systems are impaired in adult mice.
Asunto(s)
Envejecimiento/fisiología , Conducta Animal/fisiología , Proteínas Asociadas a Microtúbulos/deficiencia , Actividad Motora/genética , Anfetamina/farmacología , Análisis de Varianza , Animales , Conducta Animal/efectos de los fármacos , Estimulantes del Sistema Nervioso Central/farmacología , Maleato de Dizocilpina/farmacología , Relación Dosis-Respuesta a Droga , Antagonistas de Aminoácidos Excitadores/farmacología , Conducta Exploratoria/efectos de los fármacos , Conducta Exploratoria/fisiología , Masculino , Ratones , Ratones Noqueados , Actividad Motora/efectos de los fármacosRESUMEN
PURPOSE: To evaluate whether static high magnetic fields (HMFs), in the range of 10-17 T, affect the cytoskeleton and cell organization in different types of mammalian cells, including fibroblasts, epithelial cells, and differentiating neurons. MATERIALS AND METHODS: Cells were exposed to HMF for 30 or 60 minutes and subsequently assessed for viability. Cytoskeleton arrays and focal adhesions were visualized using immunofluorescence microscopy. RESULTS: Cell exposure to HMF over 10 T in the case of cycling cells, and over 15 T in the case of neurons, affected cell viability, apparently because of cell detachment from culture dishes. In the remaining adherent cells, the organization of actin assemblies was perturbed, and both cell adhesion and spreading were impaired. Moreover, in the case of neurons, exposure to HMF induced growth cone retraction and delayed cell differentiation. CONCLUSION: Cell exposure to HMF (over 10T and 15 T in the case of cycling cells and neurons, respectively) affects the cell cytoskeleton, with deleterious effects on cell viability, organization, and differentiation. Further studies are needed to determine whether such perturbations, as observed here in cultured cells, have consequences in whole animals.
Asunto(s)
Fenómenos Fisiológicos Celulares , Citoesqueleto/fisiología , Magnetismo , Animales , Apoptosis/fisiología , Ciclo Celular/fisiología , Diferenciación Celular/fisiología , División Celular/fisiología , Supervivencia Celular/fisiología , Células Cultivadas , Humanos , Ratones , Neuronas/fisiología , Ratas , TransfecciónRESUMEN
Neuroleptics are thought to exert their anti-psychotic effects by counteracting a hyper-dopaminergic transmission. Here, we have examined the dopaminergic status of STOP (stable tubule only polypeptide) null mice, which lack a microtubule-stabilizing protein and which display neuroleptic-sensitive behavioural disorders. Dopamine transmission was investigated using both behavioural analysis and measurements of dopamine efflux in different conditions. Compared to wild-type mice in basal conditions or following mild stress, STOP null mice showed a hyper-locomotor activity, which was erased by neuroleptic treatment, and an increased locomotor reactivity to amphetamine. Such a behavioural profile is indicative of an increased dopaminergic transmission. In STOP null mice, the basal dopamine concentrations, measured by quantitative microdialysis, were normal in both the nucleus accumbens and the striatum. When measured by electrochemical techniques, the dopamine efflux evoked by electrical stimulations mimicking physiological stimuli was dramatically increased in the nucleus accumbens of STOP null mice, apparently due to an increased dopamine release, whereas dopaminergic uptake and auto-inhibition mechanisms were normal. In contrast, dopamine effluxes were slightly diminished in the striatum. Together with previous results, the present study indicates the association in STOP null mice of hippocampal hypo-glutamatergy and of limbic hyper-dopaminergy. Such neurotransmission defects are thought to be central to mental diseases such as schizophrenia.
Asunto(s)
Dopamina/fisiología , Proteínas Asociadas a Microtúbulos/deficiencia , Proteínas Asociadas a Microtúbulos/genética , Proteínas del Tejido Nervioso/deficiencia , Proteínas del Tejido Nervioso/genética , Transmisión Sináptica/genética , Potenciales de Acción/genética , Potenciales de Acción/fisiología , Animales , Antipsicóticos/farmacología , Ritmo Circadiano/efectos de los fármacos , Ritmo Circadiano/genética , Ritmo Circadiano/fisiología , Oscuridad , Estimulación Eléctrica , Luz , Masculino , Ratones , Ratones Noqueados , Proteínas Asociadas a Microtúbulos/fisiología , Actividad Motora/efectos de los fármacos , Actividad Motora/genética , Actividad Motora/fisiología , Proteínas del Tejido Nervioso/fisiología , Transmisión Sináptica/fisiologíaRESUMEN
Tubulin is subject to a special cycle of detyrosination/tyrosination in which the C-terminal tyrosine of alpha-tubulin is cyclically removed by a carboxypeptidase and readded by a tubulin-tyrosine-ligase (TTL). This tyrosination cycle is conserved in evolution, yet its physiological importance is unknown. Here, we find that TTL suppression in mice causes perinatal death. A minor pool of tyrosinated (Tyr-)tubulin persists in TTL null tissues, being present mainly in dividing TTL null cells where it originates from tubulin synthesis, but it is lacking in postmitotic TTL null cells such as neurons, which is apparently deleterious because early death in TTL null mice is, at least in part, accounted for by a disorganization of neuronal networks, including a disruption of the cortico-thalamic loop. Correlatively, cultured TTL null neurons display morphogenetic anomalies including an accelerated and erratic time course of neurite outgrowth and a premature axonal differentiation. These anomalies may involve a mislocalization of CLIP170, which we find lacking in neurite extensions and growth cones of TTL null neurons. Our results demonstrate a vital role of TTL for neuronal organization and suggest a requirement of Tyr-tubulin for proper control of neurite extensions.
Asunto(s)
Neuritas/metabolismo , Neuronas/metabolismo , Péptido Sintasas/metabolismo , Tubulina (Proteína)/metabolismo , Animales , Secuencia de Bases , Western Blotting , Encéfalo/anatomía & histología , Carbocianinas , Diferenciación Celular/fisiología , Células Cultivadas , Técnicas Histológicas , Ratones , Ratones Noqueados , Microscopía Fluorescente , Microscopía por Video , Proteínas Asociadas a Microtúbulos/metabolismo , Proteínas de Neoplasias/metabolismo , Red Nerviosa/anatomía & histología , Neuronas/citología , Péptido Sintasas/genética , ARN Interferente Pequeño/genéticaRESUMEN
Cyclic ADP-ribose, a metabolite of NAD+, is known to modulate intracellular calcium levels and signaling in various cell types, including neural cells. The enzymes responsible for producing cyclic ADP-ribose in the cytoplasm of mammalian cells remain unknown; however, two mammalian enzymes that are capable of producing cyclic ADP-ribose extracellularly have been identified, CD38 and CD157. The present study investigated whether an ADP-ribosyl cyclase/NAD+-glycohydrolase independent of CD38 is present in brain tissue. To address this question, NAD+ metabolizing activities were accurately examined in developing and adult Cd38-/- mouse brain protein extracts and cells. Low ADP-ribosyl cyclase and NAD+-glycohydrolase activities (in the range of pmol of product formed/mg of protein/min) were detected in Cd38-/- brain at all developmental stages studied. Both activities were found to be associated with cell membranes. The activities were significantly higher in Triton X-100-treated neural cells compared with intact cells, suggesting an intracellular location of the novel cyclase. The cyclase and glycohydrolase activities were optimal at pH 6.0 and were inhibited by zinc, properties which are distinct from those of CD157. Both activities were enhanced by guanosine 5'-O-(3-thiotriphosphate), a result suggesting that the novel enzyme may be regulated by a G protein-dependent mechanism. Altogether our results indicate the presence of an intracellular membrane-bound ADP-ribosyl cyclase/NAD+-glycohydrolase distinct from CD38 and from CD157 in mouse brain. This novel enzyme, which is more active in the developing brain than in the adult tissue, may play an important role in cyclic ADP-ribose-mediated calcium signaling during brain development as well as in adult tissue.
Asunto(s)
ADP-Ribosil Ciclasa/genética , ADP-Ribosil Ciclasa/metabolismo , Antígenos CD/genética , Encéfalo/metabolismo , NAD+ Nucleosidasa/química , ADP-Ribosil Ciclasa 1 , Animales , Calcio/metabolismo , Membrana Celular/metabolismo , Cloruros/farmacología , Cromatografía Líquida de Alta Presión , Detergentes/farmacología , Relación Dosis-Respuesta a Droga , Concentración de Iones de Hidrógeno , Glicoproteínas de Membrana , Ratones , Neuronas/metabolismo , Octoxinol/farmacología , Transducción de Señal , Factores de Tiempo , Zinc/metabolismo , Compuestos de Zinc/farmacologíaRESUMEN
Neurons contain abundant subsets of highly stable microtubules that resist depolymerizing conditions such as exposure to the cold. Stable microtubules are thought to be essential for neuronal development, maintenance, and function. Previous work has indicated an important role of the microtubule-associated protein STOP in the induction of microtubule cold stability. Here, we developed STOP null mice. These mice were devoid of cold-stable microtubules. In contrast to our expectations, STOP-/- mice had no detectable defects in brain anatomy but showed synaptic defects, with depleted synaptic vesicle pools and impaired synaptic plasticity, associated with severe behavioral disorders. A survey of the effects of psychotropic drugs on STOP-/- mice behavior showed a remarkable and specific effect of long-term administration of neuroleptics in alleviating these disorders. This study demonstrates that STOP is a major factor responsible for the intriguing stability properties of neuronal microtubules and is important for synaptic plasticity. Additionally, STOP-/- mice may yield a pertinent model for study of neuroleptics in illnesses such as schizophrenia, currently thought to result from synaptic defects.