RESUMEN
Study Objectives: Experimental evidence suggests that restorative processes depend on synaptic plasticity changes in the brain during sleep. We used the expression of plasticity-related genes to assess synaptic plasticity changes during drug-induced sleep. Methods: We first characterized sleep induced by eszopiclone in mice during baseline conditions and during the recovery from sleep deprivation. We then compared the expression of 18 genes and two miRNAs critically involved in synaptic plasticity in these mice. Gene expression was assessed in the cerebral cortex and hippocampus by the TaqMan reverse transcription polymerase chain reaction and correlated with sleep parameters. Results: Eszopiclone reduced the latency to nonrapid eye movement (NREM) sleep and increased NREM sleep amounts. Eszopiclone had no effect on slow wave activity (SWA) during baseline conditions but reduced the SWA increase during recovery sleep (RS) after sleep deprivation. Gene expression analyses revealed three distinct patterns: (1) four genes had higher expression either in the cortex or hippocampus in the group of mice with increased amounts of wakefulness; (2) a large proportion of plasticity-related genes (7 out of 18 genes) had higher expression during RS in the cortex but not in the hippocampus; and (3) six genes and the two miRNAs showed no significant changes across conditions. Even at a relatively high dose (20 mg/kg), eszopiclone did not reduce the expression of plasticity-related genes during RS period in the cortex. Conclusions: These results indicate that gene expression associated with synaptic plasticity occurs in the cortex in the presence of a hypnotic medication.
Asunto(s)
Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Eszopiclona/farmacología , Regulación de la Expresión Génica/efectos de los fármacos , Plasticidad Neuronal/efectos de los fármacos , Plasticidad Neuronal/genética , Fármacos Inductores del Sueño/farmacología , Sueño/efectos de los fármacos , Animales , Corteza Cerebral/efectos de los fármacos , Corteza Cerebral/metabolismo , Expresión Génica , Hipocampo/efectos de los fármacos , Hipocampo/metabolismo , Hipnóticos y Sedantes/farmacología , Masculino , Ratones , MicroARNs/genética , Sueño/genética , Privación de Sueño , Vigilia/efectos de los fármacosRESUMEN
Expression of the bacterial beta-galactosidase reporter gene (lacZ) in the vector used for the Knockout Mouse Project (KOMP) is driven by the endogenous promoter of the target gene. In tissues from KOMP mice, histochemical staining for LacZ enzyme activity can be used to determine gene expression patterns. With this technique, we have produced a comprehensive resource of gene expression using both whole mount (WM) and frozen section (FS) LacZ staining in 313 unique KOMP mutant mouse lines. Of these, â¼ 80% of mutants showed specific staining in one or more tissues, while â¼ 20% showed no specific staining, â¼ 13% had staining in only one tissue, and â¼ 25% had staining in >6 tissues. The highest frequency of specific staining occurred in the brain (â¼ 50%), male gonads (42%), and kidney (39%). The WM method was useful for rapidly identifying whole organ and some substructure staining, while the FS method often revealed substructure and cellular staining specificity. Both staining methods had >90% repeatability in biological replicates. Nonspecific LacZ staining occurs in some tissues due to the presence of bacteria or endogenous enzyme activity. However, this can be effectively distinguished from reporter gene activity by the combination of the WM and FS methods. After careful annotation, LacZ staining patterns in a high percentage of mutants revealed a unique structure-function not previously reported for many of these genes. The validation of methods for LacZ staining, annotation, and expression analysis reported here provides unique insights into the function of genes for which little is currently known.
Asunto(s)
Regulación de la Expresión Génica/genética , Genes Reporteros/genética , Operón Lac/genética , Regiones Promotoras Genéticas/genética , Animales , Atlas como Asunto , Femenino , Expresión Génica , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Coloración y Etiquetado , Relación Estructura-ActividadRESUMEN
Although the neural circuitry underlying homeostatic sleep regulation is little understood, cortical neurons immunoreactive for neuronal nitric oxide synthase (nNOS) and the neurokinin-1 receptor (NK1) have been proposed to be involved in this physiological process. By systematically manipulating the durations of sleep deprivation and subsequent recovery sleep, we show that activation of cortical nNOS/NK1 neurons is directly related to non-rapid eye movement (NREM) sleep time, NREM bout duration, and EEG δ power during NREM sleep, an index of preexisting homeostatic sleep drive. Conversely, nNOS knockout mice show reduced NREM sleep time, shorter NREM bouts, and decreased power in the low δ range during NREM sleep, despite constitutively elevated sleep drive. Cortical NK1 neurons are still activated in response to sleep deprivation in these mice but, in the absence of nNOS, they are unable to up-regulate NREM δ power appropriately. These findings support the hypothesis that cortical nNOS/NK1 neurons translate homeostatic sleep drive into up-regulation of NREM δ power through an NO-dependent mechanism.
Asunto(s)
Ondas Encefálicas/fisiología , Corteza Cerebral/fisiología , Interneuronas/metabolismo , Óxido Nítrico Sintasa de Tipo I/metabolismo , Sueño/fisiología , Animales , Recuento de Células , Electroencefalografía , Electromiografía , Inmunohistoquímica , Ratones , Ratas , Ratas Sprague-DawleyRESUMEN
The hypothalamus is a prominent target of nicotine action. We have previously shown that acute systemic nicotine treatment induces Fos expression in the lateral hypothalamus and perifornical area (LH/PFA), with orexin/hypocretin neurons being particularly responsive. However, the neurochemical correlates of acute nicotine treatment in the LH/PFA have not been described. Anatomical studies have revealed that this area receives afferents from cholinergic, glutamatergic, and GABAergic telencephalic brain regions, suggesting a potential role for these neurotransmitters in mediating the hypothalamic component of nicotine effects on homeostatic phenomena, such as arousal and appetite. Here, we used in vivo microdialysis to determine the effect of acute systemic or local nicotine on glutamate, acetylcholine, and GABA efflux in the LH/PFA of rats. Local administration of nicotine significantly increased acetylcholine and glutamate, but not GABA, in the LH/PFA. Thus, we further tested the role of afferent sources of glutamate and acetylcholine in mediating acute nicotine-induced activation of orexin neurons by unilaterally lesioning the prefrontal cortex or basal forebrain cholinergic regions. Lesioned animals showed reduced Fos-positive orexin neurons following nicotine treatment. These data suggest that both acetylcholine and glutamate may mediate the effects of acute nicotine on the activity of hypothalamic neurons, including orexin/hypocretin cells. Changes in cholinergic or glutamatergic transmission in this region with chronic nicotine may contribute to long-term alterations in functions mediated by LH/PFA neurons, including feeding and arousal.
Asunto(s)
Acetilcolina/metabolismo , Ácido Glutámico/metabolismo , Área Hipotalámica Lateral/efectos de los fármacos , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Neuronas/efectos de los fármacos , Neuropéptidos/metabolismo , Nicotina/farmacología , Animales , Apetito/efectos de los fármacos , Apetito/fisiología , Nivel de Alerta/efectos de los fármacos , Nivel de Alerta/fisiología , Esquema de Medicación , Líquido Extracelular/efectos de los fármacos , Líquido Extracelular/metabolismo , Área Hipotalámica Lateral/metabolismo , Masculino , Microdiálisis , Vías Nerviosas/efectos de los fármacos , Vías Nerviosas/fisiología , Neuronas/metabolismo , Agonistas Nicotínicos/farmacología , Orexinas , Ratas , Ratas Sprague-Dawley , Transmisión Sináptica/efectos de los fármacos , Transmisión Sináptica/fisiología , Tabaquismo/metabolismo , Tabaquismo/fisiopatología , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/fisiología , Ácido gamma-Aminobutírico/metabolismoRESUMEN
Caffeine is a widely self-administered psychostimulant with purported neuroprotective and procognitive effects in rodent models of aging. The cholinergic basal forebrain is important for arousal and attention and is implicated in age-related cognitive decline. Accordingly, we determined the effects of caffeine on cholinergic neuron activation in the rat basal forebrain. Young adult (age 2 months) male rats were treated with caffeine (0, 10, or 50 mg/kg) and killed 2 h later. Caffeine significantly increased c-Fos expression in cholinergic neurons of the horizontal limb of the diagonal band of Broca but not other basal forebrain regions such as the medial septum or substantia innominata. The horizontal limb of the diagonal band of Broca provides cholinergic innervation to the olfactory bulb, suggesting that deficits in this structure may contribute to diminished olfactory function observed in Alzheimer's disease patients. These results suggest that part of the cognitive-enhancing effects of caffeine may be mediated through activation of this part of the cholinergic basal forebrain.
Asunto(s)
Cafeína/farmacología , Estimulantes del Sistema Nervioso Central/farmacología , Colina O-Acetiltransferasa/metabolismo , Neuronas/efectos de los fármacos , Prosencéfalo/efectos de los fármacos , Proteínas Proto-Oncogénicas c-fos/metabolismo , Animales , Cafeína/administración & dosificación , Recuento de Células , Estimulantes del Sistema Nervioso Central/administración & dosificación , Relación Dosis-Respuesta a Droga , Inmunohistoquímica , Masculino , Neuronas/metabolismo , Prosencéfalo/metabolismo , Ratas , Ratas Sprague-Dawley , Factores de TiempoRESUMEN
Orexin/hypocretin neurons of the lateral hypothalamus/perifornical area project to a diverse array of brain regions and are responsive to a variety of psychostimulant drugs. It has been shown that orexin neurons are activated by systemic nicotine administration suggesting a possible orexinergic contribution to the effects of this drug on arousal and cognitive function. The basal forebrain and paraventricular nucleus of the dorsal thalamus (PVT) both receive orexin inputs and have been implicated in arousal, attention and psychostimulant drug responses. However, it is unknown whether orexin inputs to these areas are activated by psychostimulant drugs such as nicotine. Here, we infused the retrograde tract tracer cholera toxin B subunit (CTb) into either the basal forebrain or PVT of adult male rats. Seven to 10 days later, animals received an acute systemic administration of (-) nicotine hydrogen tartrate or vehicle and were euthanized 2h later. Triple-label immunohistochemistry/immunofluorescence was used to detect Fos expression in retrogradely-labeled orexin neurons. Nicotine increased Fos expression in orexin neurons projecting to both basal forebrain and PVT. The relative activation in lateral and medial banks of retrogradely-labeled orexin neurons was similar following basal forebrain CTb deposits, but was more pronounced in the medial bank following PVT deposits of CTb. Our findings suggest that orexin inputs to the basal forebrain and PVT may contribute to nicotine effects on arousal and cognition and provide further support for the existence of functional heterogeneity across the medial-lateral distribution of orexin neurons.
Asunto(s)
Vías Eferentes/metabolismo , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Núcleos Talámicos de la Línea Media/metabolismo , Neuronas/metabolismo , Nicotina/farmacología , Animales , Toxina del Cólera/administración & dosificación , Toxina del Cólera/metabolismo , Vías Eferentes/citología , Vías Eferentes/efectos de los fármacos , Técnica del Anticuerpo Fluorescente/métodos , Globo Pálido/citología , Globo Pálido/efectos de los fármacos , Globo Pálido/metabolismo , Inmunohistoquímica/métodos , Inyecciones Intraperitoneales , Péptidos y Proteínas de Señalización Intracelular/administración & dosificación , Péptidos y Proteínas de Señalización Intracelular/efectos de los fármacos , Masculino , Núcleos Talámicos de la Línea Media/citología , Núcleos Talámicos de la Línea Media/efectos de los fármacos , Neuronas/citología , Neuronas/efectos de los fármacos , Neuropéptidos/administración & dosificación , Neuropéptidos/metabolismo , Neurotransmisores/administración & dosificación , Neurotransmisores/metabolismo , Neurotransmisores/farmacología , Nicotina/administración & dosificación , Orexinas , Proteínas Proto-Oncogénicas c-fos/biosíntesis , Proteínas Proto-Oncogénicas c-fos/metabolismo , Ratas , Ratas Sprague-Dawley , Sustancia Innominada/citología , Sustancia Innominada/efectos de los fármacos , Sustancia Innominada/metabolismo , Tálamo/citología , Tálamo/efectos de los fármacos , Tálamo/metabolismoRESUMEN
Sleep deprivation (SD) results in increased electroencephalographic (EEG) delta power during subsequent non-rapid eye movement sleep (NREMS) and is associated with changes in the expression of circadian clock-related genes in the cerebral cortex. The increase of NREMS delta power as a function of previous wake duration varies among inbred mouse strains. We sought to determine whether SD-dependent changes in circadian clock gene expression parallel this strain difference described previously at the EEG level. The effects of enforced wakefulness of incremental durations of up to 6 h on the expression of circadian clock genes (bmal1, clock, cry1, cry2, csnk1epsilon, npas2, per1, and per2) were assessed in AKR/J, C57BL/6J, and DBA/2J mice, three strains that exhibit distinct EEG responses to SD. Cortical expression of clock genes subsequent to SD was proportional to the increase in delta power that occurs in inbred strains: the strain that exhibits the most robust EEG response to SD (AKR/J) exhibited dramatic increases in expression of bmal1, clock, cry2, csnkIepsilon, and npas2, whereas the strain with the least robust response to SD (DBA/2) exhibited either no change or a decrease in expression of these genes and cry1. The effect of SD on circadian clock gene expression was maintained in mice in which both of the cryptochrome genes were genetically inactivated. cry1 and cry2 appear to be redundant in sleep regulation as elimination of either of these genes did not result in a significant deficit in sleep homeostasis. These data demonstrate transcriptional regulatory correlates to previously described strain differences at the EEG level and raise the possibility that genetic differences underlying circadian clock gene expression may drive the EEG differences among these strains.
Asunto(s)
Ritmo alfa , Corteza Cerebral/fisiología , Ritmo Circadiano/genética , Regulación de la Expresión Génica/fisiología , Privación de Sueño/metabolismo , Análisis de Varianza , Animales , Proteínas CLOCK , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Criptocromos , Flavoproteínas/genética , Flavoproteínas/metabolismo , Ratones , Ratones Endogámicos , Ratones Noqueados , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Proteínas Circadianas Period , Especificidad de la Especie , Transactivadores/genética , Transactivadores/metabolismo , Factores de Transcripción/genética , Factores de Transcripción/metabolismoRESUMEN
Depressive illness is associated with changes in amygdalar volume, and stressful life events are known to precipitate depressive episodes in this patient population. Stress affects amygdalar synaptic plasticity and several neurotransmitter systems have been implicated in stress-mediated changes in the brain, including the glutamatergic system. However, the role of the glutamatergic system in stress-mediated plasticity in the amygdala remains to be determined. Accordingly the current study examined the stress modulation of extracellular glutamate levels in the basolateral nucleus (BLA) and the central nucleus (CeA) of the amygdala by in vivo microdialysis. Acute stress increased extracellular glutamate levels in the BLA and CeA, although the dynamics of these stress-mediated changes were dramatically different in these amygdalar nuclei. Tetrodotoxin administration reduced basal, and completely eliminated stress-mediated increases in glutamate efflux in the amygdala, demonstrating that stress effects are dependent on local axonal depolarization. Moreover, stress-mediated increases in glutamate efflux in the BLA were inhibited by the antidepressant tianeptine but not by the selective serotonin-reuptake inhibitor fluoxetine. Collectively, these data demonstrate that stress-induced modulation of glutamate neurochemistry reflects a fundamental pathological change that may contribute to the aetiology and progression of depressive illness, and suggest that some antidepressants such as tianeptine may elicit their clinical effects by modulation of glutamatergic neurotransmission.
Asunto(s)
Amígdala del Cerebelo/efectos de los fármacos , Antidepresivos/farmacología , Trastorno Depresivo/tratamiento farmacológico , Ácido Glutámico/metabolismo , Estrés Psicológico/metabolismo , Enfermedad Aguda/terapia , Amígdala del Cerebelo/metabolismo , Animales , Antidepresivos de Segunda Generación/farmacología , Antidepresivos Tricíclicos/farmacología , Axones/efectos de los fármacos , Axones/metabolismo , Trastorno Depresivo/etiología , Trastorno Depresivo/fisiopatología , Líquido Extracelular/efectos de los fármacos , Líquido Extracelular/metabolismo , Fluoxetina/farmacología , Masculino , Potenciales de la Membrana/efectos de los fármacos , Potenciales de la Membrana/fisiología , Microdiálisis , Ratas , Ratas Sprague-Dawley , Bloqueadores de los Canales de Sodio/farmacología , Estrés Psicológico/complicaciones , Estrés Psicológico/fisiopatología , Tiazepinas/farmacología , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/fisiologíaRESUMEN
Several convergent lines of evidence point to the amygdala as a key site of plasticity underlying most forms of fear conditioning. Studies have shown that chronic physical activity, such as wheel running, can alter learning in a variety of contexts, including aversive conditioning. The ability of chronic wheel running (WR) to alter both behavioral correlates of fear conditioning and indices of amygdalar activation, however, has not been simultaneously assessed. Here, rats were given constant access to either free-turning or--as a control--locked (LC) running wheels in their home cages. After 8 weeks of housing under these conditions, animals were exposed to a series of shocks in a separate testing chamber. Twenty-four hours later, the animals were returned to the shock chamber and freezing behavior was measured as an indicator of contextual fear conditioning. The animals were then sacrificed and their brains processed for immunohistochemical detection of Fos to assess patterns of putative neuronal activation. WR rats spent significantly more time freezing than their LC counterparts upon return to the shock-paired context. The enhanced conditioned freezing response was most pronounced in animals showing high levels of nightly wheel running activity. WR animals also had significantly higher levels of neuronal activation, as indicated by Fos expression in the central nucleus of the amygdala, but less activation in the basolateral nucleus, compared to sedentary controls. These data demonstrate the ability of chronic physical activity to alter contextual fear conditioning and implicate the amygdala as a potential site of plasticity underlying this phenomenon.
Asunto(s)
Amígdala del Cerebelo/fisiología , Conducta Animal/fisiología , Condicionamiento Psicológico/fisiología , Miedo/fisiología , Actividad Motora/fisiología , Animales , Masculino , Proteínas Proto-Oncogénicas c-fos/biosíntesis , Ratas , Ratas Sprague-DawleyRESUMEN
The hypothalamus is a prominent central site of action of nicotine but the phenotype of nicotine-sensitive neurons in this region has not been fully described. Hypothalamic orexin neurons are important regulators of state-dependent behavior, arousal and feeding. Here, we treated rats with acute nicotine and quantitated Fos expression as a marker of neuronal activation. Nicotine increased the percentage of orexin neurons expressing Fos without a significant effect on non-orexin neurons. This effect was attenuated by the nicotinic antagonists mecamylamine and dihydro-beta-erythroidine, implicating alpha4beta2-containing nicotinic receptors. The orexin system is likely to play an important role in the coordination of physiological and behavioral responses to acute nicotine treatment.