RESUMEN
The cystine/glutamate antiporter system xc- has been identified as the major source of extracellular glutamate in several brain regions as well as a modulator of neuroinflammation, and genetic deletion of its specific subunit xCT (xCT-/-) is protective in mouse models for age-related neurological disorders. However, the previously observed oxidative shift in the plasma cystine/cysteine ratio of adult xCT-/- mice led to the hypothesis that system xc- deletion would negatively affect life- and healthspan. Still, till now the role of system xc- in physiological aging remains unexplored. We therefore studied the effect of xCT deletion on the aging process of mice, with a particular focus on the immune system, hippocampal function, and cognitive aging. We observed that male xCT-/- mice have an extended lifespan, despite an even more increased plasma cystine/cysteine ratio in aged compared to adult mice. This oxidative shift does not negatively impact the general health status of the mice. On the contrary, the age-related priming of the innate immune system, that manifested as increased LPS-induced cytokine levels and hypothermia in xCT+/+ mice, was attenuated in xCT-/- mice. While this was associated with only a very moderate shift towards a more anti-inflammatory state of the aged hippocampus, we observed changes in the hippocampal metabolome that were associated with a preserved hippocampal function and the retention of hippocampus-dependent memory in male aged xCT-/- mice. Targeting system xc- is thus not only a promising strategy to prevent cognitive decline, but also to promote healthy aging.
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Sistema de Transporte de Aminoácidos y+ , Cistina , Sistema de Transporte de Aminoácidos y+/genética , Sistema de Transporte de Aminoácidos y+/metabolismo , Animales , Cisteína , Cistina/metabolismo , Ácido Glutámico , Hipocampo/metabolismo , Longevidad , Masculino , Ratones , Ratones Endogámicos C57BLRESUMEN
The astrocytic cystine/glutamate antiporter system xc- represents an important source of extracellular glutamate in the central nervous system, with potential impact on excitatory neurotransmission. Yet, its function and importance in brain physiology remain incompletely understood. Employing slice electrophysiology and mice with a genetic deletion of the specific subunit of system xc-, xCT (xCT-/- mice), we uncovered decreased neurotransmission at corticostriatal synapses. This effect was partly mitigated by replenishing extracellular glutamate levels, indicating a defect linked with decreased extracellular glutamate availability. We observed no changes in the morphology of striatal medium spiny neurons, the density of dendritic spines, or the density or ultrastructure of corticostriatal synapses, indicating that the observed functional defects are not due to morphological or structural abnormalities. By combining electron microscopy with glutamate immunogold labeling, we identified decreased intracellular glutamate density in presynaptic terminals, presynaptic mitochondria, and in dendritic spines of xCT-/- mice. A proteomic and kinomic screen of the striatum of xCT-/- mice revealed decreased expression of presynaptic proteins and abnormal kinase network signaling, that may contribute to the observed changes in postsynaptic responses. Finally, these corticostriatal deregulations resulted in a behavioral phenotype suggestive of autism spectrum disorder in the xCT-/- mice; in tests sensitive to corticostriatal functioning we recorded increased repetitive digging behavior and decreased sociability. To conclude, our findings show that system xc- plays a previously unrecognized role in regulating corticostriatal neurotransmission and influences social preference and repetitive behavior.
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Trastorno del Espectro Autista , Ácido Glutámico , Animales , Antiportadores , Trastorno del Espectro Autista/genética , Cistina , Ratones , Proteómica , Interacción SocialRESUMEN
Lithium's inhibitory effect on enzymes involved in sulfation process, such as inhibition of 3'(2')-phosphoadenosine 5'-phosphate (PAP) phosphatase, is a possible mechanism of its therapeutic effect for bipolar disorder (BD). 3'-Phosphoadenosine 5'-phosphosulfate (PAPS) is translocated from cytosol to Golgi lumen by PAPS transporter 1 (PAPST1/SLC35B2), where it acts as a sulfa donor. Since SLC35B2 was previously recognized as a molecule that facilitates the release of D-serine, a co-agonist of N-methyl-D-aspartate type glutamate receptor, altered function of SLC35B2 might be associated with the pathophysiology of BD and schizophrenia (SCZ). We performed genetic association analyses of the SLC35B2 gene using Japanese cohorts with 366 BD cases and 370 controls and 2012 SCZ cases and 2170 controls. We then investigated expression of SLC35B2 mRNA in postmortem brains by QPCR using a Caucasian cohort with 33 BD and 34 SCZ cases and 34 controls and by in situ hybridization using a Caucasian cohort with 37 SCZ and 29 controls. We found significant associations between three SNPs (rs575034, rs1875324, and rs3832441) and BD, and significantly reduced SLC35B2 mRNA expression in postmortem dorsolateral prefrontal cortex (DLPFC) of BD. Moreover, we observed normalized SLC35B2 mRNA expression in BD subgroups who were medicated with lithium. While there was a significant association of SLC35B2 with SCZ (SNP rs2233437), its expression was not changed in SCZ. These findings indicate that SLC35B2 might be differentially involved in the pathophysiology of BD and SCZ by influencing the sulfation process and/or glutamate system in the central nervous system.
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Trastorno Bipolar , Esquizofrenia , Trastorno Bipolar/tratamiento farmacológico , Trastorno Bipolar/genética , Trastorno Bipolar/metabolismo , Humanos , Litio/metabolismo , Polimorfismo de Nucleótido Simple , ARN Mensajero/metabolismo , Esquizofrenia/genética , Esquizofrenia/metabolismo , Transportadores de Sulfato/genéticaRESUMEN
OBJECTIVE: The cystine/glutamate antiporter system xc- could represent a new target for antiepileptogenic treatments due to its crucial roles in glutamate homeostasis and neuroinflammation. To demonstrate this, we compared epilepsy development and seizure susceptibility in xCT knockout mice (xCT-/- ) and in littermate controls (xCT+/+ ) in different chronic models of epilepsy. METHODS: Mice were surgically implanted with electrodes in the basolateral amygdala and chronically stimulated to develop self-sustained status epilepticus (SSSE); continuous video-electroencephalography monitoring was performed for 28 days after SE and hippocampal histopathology was assessed. Corneal kindling was induced by twice daily electrical stimulation at 6 Hz and maintenance of the fully kindled state was evaluated. Next, messenger RNA (mRNA) and protein levels of xCT and of the proteins involved in the phosphoinositide 3-kinase (PI3K)/Akt/glycogen synthase kinase 3ß (GSK-3ß)/eukaryotic initiation factor 2α (eIF2α)/activating transcription factor 4 (ATF4) signaling pathway were measured at different time points during epileptogenesis in NMRI mice treated with pilocarpine. Finally, the anticonvulsant effect of sulfasalazine (SAS), a nonselective system xc- inhibitor, was assessed against 6 Hz-evoked seizures in pilocarpine-treated mice. RESULTS: In the SSSE model, xCT-/- mice displayed a significant delayed epileptogenesis, a reduced number of spontaneous recurrent seizures, and less pronounced astrocytic and microglial activation. Moreover, xCT-/- mice showed reduced seizure severity during 6 Hz kindling development and a lower incidence of generalized seizures during the maintenance of the fully kindled state. In pilocarpine-treated mice, protein levels of the PI3K/Akt/GSK-3ß/eIF2α/ATF4 pathway were increased during the chronic phase of the model, consistent with previous findings in the hippocampus of patients with epilepsy. Finally, repeated administration of SAS protected pilocarpine-treated mice against acute 6 Hz seizure induction, in contrast to sham controls, in which system xc- is not activated. SIGNIFICANCE: Inhibition of system xc- could be an attractive target for the development of new therapies with a potential for disease modification in epilepsy.
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Sistema de Transporte de Aminoácidos y+/efectos de los fármacos , Anticonvulsivantes/farmacología , Epilepsia/tratamiento farmacológico , Sistema de Transporte de Aminoácidos y+/metabolismo , Animales , Anticonvulsivantes/uso terapéutico , Modelos Animales de Enfermedad , Electroencefalografía , Epilepsia/etiología , Epilepsia/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Pilocarpina/farmacología , Estado Epiléptico/tratamiento farmacológico , Estado Epiléptico/etiología , Estado Epiléptico/metabolismoRESUMEN
The communication between the immune and central nervous system (CNS) is affected in many neurological disorders. Peripheral injections of the endotoxin lipopolysaccharide (LPS) are widely used to study this communication: an LPS challenge leads to a biphasic syndrome that starts with acute sickness and is followed by persistent brain inflammation and chronic behavioral alterations such as depressive-like symptoms. In vitro, the response to LPS treatment has been shown to involve enhanced expression of system x c - . This cystine-glutamate antiporter, with xCT as specific subunit, represents the main glial provider of extracellular glutamate in mouse hippocampus. Here we injected male xCT knockout and wildtype mice with a single intraperitoneal dose of 5 mg/kg LPS. LPS-injection increased hippocampal xCT expression but did not alter the mainly astroglial localization of the xCT protein. Peripheral and central inflammation (as defined by cytokine levels and morphological activation of microglia) as well as LPS-induced sickness and depressive-like behavior were significantly attenuated in xCT-deficient mice compared with wildtype mice. Our study is the first to demonstrate the involvement of system x c - in peripheral and central inflammation in vivo and the potential therapeutic relevance of its inhibition in brain disorders characterized by peripheral and central inflammation, such as depression.
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Sistema de Transporte de Aminoácidos y+/deficiencia , Depresión/metabolismo , Conducta de Enfermedad/fisiología , Inflamación/metabolismo , Sistema de Transporte de Aminoácidos y+/genética , Animales , Astrocitos/metabolismo , Astrocitos/patología , Citocinas/metabolismo , Depresión/patología , Transportador 2 de Aminoácidos Excitadores/metabolismo , Eliminación de Gen , Proteína Ácida Fibrilar de la Glía/metabolismo , Hipocampo/metabolismo , Hipocampo/patología , Inflamación/patología , Lipopolisacáridos , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Microglía/metabolismo , Microglía/patología , ARN Mensajero/metabolismoRESUMEN
The cystine-glutamate exchanger (xCT) promotes glutathione synthesis by catalyzing cystine uptake and glutamate release. The released glutamate may modulate normal neural signaling and contribute to excitotoxicity in pathological situations. Uncertainty, however, remains as neither the expression levels nor the distribution of xCT have been unambiguously determined. In fact, xCT has been reported in astrocytes, neurons, oligodendrocytes and microglia, but most of the information derives from cell cultures. Here, we show by immunohistochemistry and by Western blotting that xCT is widely expressed in the central nervous system of both sexes. The labeling specificity was validated using tissue from xCT knockout mice as controls. Astrocytes were selectively labeled, but showed greatly varying labeling intensities. This astroglial heterogeneity resulted in an astrocyte domain-like labeling pattern. Strong xCT labeling was also found in the leptomeninges, along some blood vessels, in selected circumventricular organs and in a subpopulation of tanycytes residing the lateral walls of the ventral third ventricle. Neurons, oligodendrocytes and resting microglia, as well as reactive microglia induced by glutamine synthetase deficiency, were unlabeled. The concentration of xCT protein in hippocampus was compared with that of the EAAT3 glutamate transporter by immunoblotting using a chimeric xCT-EAAT3 protein to normalize xCT and EAAT3 labeling intensities. The immunoblots suggested an xCT/EAAT3 ratio close to one (0.75 ± 0.07; average ± SEM; n = 4) in adult C57BL6 mice. CONCLUSIONS: xCT is present in select blood/brain/CSF interface areas and in an astrocyte subpopulation, in sufficient quantities to support the notion that system xc- provides physiologically relevant transport activity.
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Sistema de Transporte de Aminoácidos y+/metabolismo , Astrocitos/metabolismo , Encéfalo/metabolismo , Sistema de Transporte de Aminoácidos y+/genética , Animales , Astrocitos/citología , Western Blotting , Encéfalo/citología , Proteínas de Unión al Calcio/metabolismo , Transportador 2 de Aminoácidos Excitadores/metabolismo , Transportador 3 de Aminoácidos Excitadores/metabolismo , Femenino , Inmunohistoquímica , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas de Microfilamentos/metabolismoRESUMEN
BACKGROUND: Multiple sclerosis (MS) is an autoimmune demyelinating disease that affects the central nervous system (CNS), leading to neurodegeneration and chronic disability. Accumulating evidence points to a key role for neuroinflammation, oxidative stress, and excitotoxicity in this degenerative process. System xc- or the cystine/glutamate antiporter could tie these pathological mechanisms together: its activity is enhanced by reactive oxygen species and inflammatory stimuli, and its enhancement might lead to the release of toxic amounts of glutamate, thereby triggering excitotoxicity and neurodegeneration. METHODS: Semi-quantitative Western blotting served to study protein expression of xCT, the specific subunit of system xc-, as well as of regulators of xCT transcription, in the normal appearing white matter (NAWM) of MS patients and in the CNS and spleen of mice exposed to experimental autoimmune encephalomyelitis (EAE), an accepted mouse model of MS. We next compared the clinical course of the EAE disease, the extent of demyelination, the infiltration of immune cells and microglial activation in xCT-knockout (xCT-/-) mice and irradiated mice reconstituted in xCT-/- bone marrow (BM), to their proper wild type (xCT+/+) controls. RESULTS: xCT protein expression levels were upregulated in the NAWM of MS patients and in the brain, spinal cord, and spleen of EAE mice. The pathways involved in this upregulation in NAWM of MS patients remain unresolved. Compared to xCT+/+ mice, xCT-/- mice were equally susceptible to EAE, whereas mice transplanted with xCT-/- BM, and as such only exhibiting loss of xCT in their immune cells, were less susceptible to EAE. In none of the above-described conditions, demyelination, microglial activation, or infiltration of immune cells were affected. CONCLUSIONS: Our findings demonstrate enhancement of xCT protein expression in MS pathology and suggest that system xc- on immune cells invading the CNS participates to EAE. Since a total loss of system xc- had no net beneficial effects, these results have important implications for targeting system xc- for treatment of MS.
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Sistema de Transporte de Aminoácidos y+/deficiencia , Sistema Nervioso Central/metabolismo , Encefalomielitis Autoinmune Experimental/metabolismo , Inmunidad Celular/fisiología , Esclerosis Múltiple/metabolismo , Anciano , Anciano de 80 o más Años , Sistema de Transporte de Aminoácidos y+/genética , Sistema de Transporte de Aminoácidos y+/inmunología , Animales , Sistema Nervioso Central/inmunología , Sistema Nervioso Central/patología , Encefalomielitis Autoinmune Experimental/inmunología , Encefalomielitis Autoinmune Experimental/patología , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Ratones Transgénicos , Microglía/patología , Microglía/fisiología , Persona de Mediana Edad , Esclerosis Múltiple/inmunología , Esclerosis Múltiple/patologíaRESUMEN
Parkinson's disease (PD) is a neurodegenerative disorder, characterized by a loss of dopamine (DA) neurons in the substantia nigra pars compacta (SNc). Caloric restriction (CR) has been shown to exert ghrelin-dependent neuroprotective effects in the 1-methyl-4-phenyl-1,2,3,6-tetrathydropyridine (MPTP)-based animal model for PD. We here investigated whether CR is neuroprotective in the lactacystin (LAC) mouse model for PD, in which proteasome disruption leads to the destruction of the DA neurons of the SNc, and whether this effect is mediated via the ghrelin receptor. Adult male ghrelin receptor wildtype (WT) and knockout (KO) mice were maintained on an ad libitum (AL) diet or on a 30% CR regimen. After 3 weeks, LAC was injected unilaterally into the SNc, and the degree of DA neuron degeneration was evaluated 1 week later. In AL mice, LAC injection significanty reduced the number of DA neurons and striatal DA concentrations. CR protected against DA neuron degeneration following LAC injection. However, no differences were observed between ghrelin receptor WT and KO mice. These results indicate that CR can protect the nigral DA neurons from toxicity related to proteasome disruption; however, the ghrelin receptor is not involved in this effect.
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Acetilcisteína/análogos & derivados , Restricción Calórica , Neuronas Dopaminérgicas/efectos de los fármacos , Neuronas Dopaminérgicas/metabolismo , Fármacos Neuroprotectores , Receptores de Ghrelina/metabolismo , Acetilcisteína/administración & dosificación , Acetilcisteína/farmacología , Factores de Edad , Animales , Recuento de Células , Masculino , Ratones , Ratones Noqueados , Receptores de Ghrelina/genética , Sustancia Negra/efectos de los fármacos , Sustancia Negra/metabolismo , Sustancia Negra/patologíaRESUMEN
Genes associated with endolysosomal function have been recently associated with familial Parkinson's disease and described as risk factors for sporadic cases. This indicates that deficits in this pathway predispose to parkinsonism. To better understand the role of these genes in disease development, rodent models have been created by targeting genes playing a role in endolysosomal function, such as LRRK2, DNAJC6, SYNJ1, VPS35, GBA1, ATP13A2 and TMEM175. Here, we review the latest findings describing parkinsonian features in these animal models secondary to endolysosomal dysfunction. Also, we provide suggestions for further development and application of these animal models to better understand the contribution of endolysosomal dysfunction in Parkinson's disease and provide novel models for testing therapeutic approaches.
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Enfermedad de Parkinson , Animales , Endosomas , Lisosomas/genética , Lisosomas/metabolismo , Enfermedad de Parkinson/genética , RoedoresRESUMEN
Despite ample evidence for the therapeutic potential of inhibition of the cystine/glutamate antiporter system xc - in neurological disorders and in cancer, none of the proposed inhibitors is selective. In this context, a lot of research has been performed using the EMA- and FDA-approved drug sulfasalazine (SAS). Even though this molecule is already on the market for decades as an anti-inflammatory drug, serious side effects due to its use have been reported. Whereas for the treatment of the main indications, SAS needs to be cleaved in the intestine into the anti-inflammatory compound mesalazine, it needs to reach the systemic circulation in its intact form to allow inhibition of system xc -. The higher plasma levels of intact SAS (or its metabolites) might induce adverse effects, independent of its action on system xc -. Some of these effects have however been attributed to system xc - inhibition, calling into question the safety of targeting system xc -. In this study we chronically treated system xc - - deficient mice and their wildtype littermates with two different doses of SAS (160 mg/kg twice daily or 320 mg/kg once daily, i.p.) and studied some of the adverse effects that were previously reported. SAS had a negative impact on the survival rate, the body weight, the thermoregulation and/or stress reaction of mice of both genotypes, and thus independent of its inhibitory action on system xc -. While SAS decreased the total distance travelled in the open-field test the first time the mice encountered the test, it did not influence this parameter on the long-term and it did not induce other behavioral changes such as anxiety- or depressive-like behavior. Finally, no major histological abnormalities were observed in the spinal cord. To conclude, we were unable to identify any undesirable system xc --dependent effect of chronic administration of SAS.
RESUMEN
The astrocytic cystine/glutamate antiporter system x c - (with xCT as the specific subunit) imports cystine in exchange for glutamate and has been shown to interact with multiple pathways in the brain that are dysregulated in age-related neurological disorders, including glutamate homeostasis, redox balance, and neuroinflammation. In the current study, we investigated the effect of genetic xCT deletion on lactacystin (LAC)- and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced degeneration of the nigrostriatal pathway, as models for Parkinson's disease (PD). Dopaminergic neurons of adult xCT knock-out mice (xCT-/-) demonstrated an equal susceptibility to intranigral injection of the proteasome inhibitor LAC, as their wild-type (xCT+/+) littermates. Contrary to adult mice, aged xCT-/- mice showed a significant decrease in LAC-induced degeneration of nigral dopaminergic neurons, depletion of striatal dopamine (DA) and neuroinflammatory reaction, compared to age-matched xCT+/+ littermates. Given this age-related protection, we further investigated the sensitivity of aged xCT-/- mice to chronic and progressive MPTP treatment. However, in accordance with our previous observations in adult mice (Bentea et al., 2015a), xCT deletion did not confer protection against MPTP-induced nigrostriatal degeneration in aged mice. We observed an increased loss of nigral dopaminergic neurons, but equal striatal DA denervation, in MPTP-treated aged xCT-/- mice when compared to age-matched xCT+/+ littermates. To conclude, we reveal age-related protection against proteasome inhibition-induced nigrostriatal degeneration in xCT-/- mice, while xCT deletion failed to protect nigral dopaminergic neurons of aged mice against MPTP-induced toxicity. Our findings thereby provide new insights into the role of system x c - in mechanisms of dopaminergic cell loss and its interaction with aging.
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Phosphorylation by serine-threonine and tyrosine kinases is critical for determining protein function. Array-based platforms for measuring reporter peptide signal levels allow for differential phosphorylation analysis between conditions for distinct active kinases. Peptide array technologies like the PamStation12 from PamGene allow for generating high-throughput, multi-dimensional, and complex functional proteomics data. As the adoption rate of such technologies increases, there is an imperative need for software tools that streamline the process of analyzing such data. We present Kinome Random Sampling Analyzer (KRSA), an R package and R Shiny web-application for analyzing kinome array data to help users better understand the patterns of functional proteomics in complex biological systems. KRSA is an All-In-One tool that reads, formats, fits models, analyzes, and visualizes PamStation12 kinome data. While the underlying algorithm has been experimentally validated in previous publications, we demonstrate KRSA workflow on dorsolateral prefrontal cortex (DLPFC) in male (n = 3) and female (n = 3) subjects to identify differential phosphorylation signatures and upstream kinase activity. Kinase activity differences between males and females were compared to a previously published kinome dataset (11 female and 7 male subjects) which showed similar global phosphorylation signals patterns.
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Corteza Prefontal Dorsolateral/enzimología , Familia de Multigenes , Fosfoproteínas/metabolismo , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Tirosina Quinasas/metabolismo , Programas Informáticos , Algoritmos , Autopsia , Benchmarking , Conjuntos de Datos como Asunto , Corteza Prefontal Dorsolateral/química , Femenino , Expresión Génica , Humanos , Masculino , Fosfoproteínas/clasificación , Fosfoproteínas/genética , Fosforilación , Análisis de Componente Principal , Análisis por Matrices de Proteínas , Proteínas Serina-Treonina Quinasas/clasificación , Proteínas Serina-Treonina Quinasas/genética , Proteínas Tirosina Quinasas/clasificación , Proteínas Tirosina Quinasas/genética , Proteómica/métodosRESUMEN
INTRODUCTION: Besides proteasome dysfunction, neuroinflammation is a common feature in the pathogenesis of Parkinson's disease (PD). Accordingly, peripheral inflammation has been shown to increase the susceptibility of the brain for nigrostriatal degeneration by inducing activation of glial cells and release of pro-inflammatory cytokines in the brain. Given that current animal models of PD fail to recapitulate the pathophysiology occurring in idiopathic PD, the aim of this study was to combine two pathogenic mechanisms (i.e. neuroinflammation and proteasome inhibition) to create a dual-hit mouse model of PD. METHODS: We repeatedly injected mice with a low dose of LPS (250⯵g/kg/day i. p. for four days) to induce neuroinflammation, followed by a unilateral intranigral injection of lactacystin (LAC; 3⯵g). Seven days later, mice were evaluated behaviorally to assess locomotion, anxiety- and depressive-like behavior. Nigrostriatal degeneration was analyzed by measuring striatal dopamine loss as well as loss of nigral dopaminergic neurons. Neuroinflammation was confirmed by quantifying microglial cells in the substantia nigra (SN) and cytokine expression in the striatum. RESULTS: Repeated systemic LPS injections increase the number of microglial cells in the SN and induce a mixed profile of pro- and anti-inflammatory cytokines in the striatum without affecting the integrity of the nigrostriatal pathway. Systemic LPS-induced neuroinflammation, however, increases the susceptibility of the nigrostriatal pathway for LAC-induced degeneration. CONCLUSION: Recapitulating two relevant etiopathogenic mechanisms of PD - neuroinflammation and proteasome inhibition-, we propose this dual-hit model as a relevant mouse model for PD that could be used to investigate potential therapeutic targets.
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Cuerpo Estriado/efectos de los fármacos , Inhibidores de Cisteína Proteinasa/farmacología , Conducta de Enfermedad/efectos de los fármacos , Inflamación/inducido químicamente , Lipopolisacáridos/farmacología , Microglía/efectos de los fármacos , Degeneración Nerviosa/inducido químicamente , Inhibidores de Proteasoma/farmacología , Sustancia Negra/efectos de los fármacos , Acetilcisteína/análogos & derivados , Acetilcisteína/farmacología , Animales , Conducta Animal , Inhibidores de Cisteína Proteinasa/administración & dosificación , Modelos Animales de Enfermedad , Lipopolisacáridos/administración & dosificación , Masculino , Ratones , Ratones Endogámicos C57BL , Vías Nerviosas/efectos de los fármacos , Inhibidores de Proteasoma/administración & dosificaciónRESUMEN
Converging evidence suggests bioenergetic defects contribute to the pathophysiology of schizophrenia and may underlie cognitive dysfunction. The transport and metabolism of lactate energetically couples astrocytes and neurons and supports brain bioenergetics. We examined the concentration of lactate in postmortem brain (dorsolateral prefrontal cortex) in subjects with schizophrenia, in two animal models of schizophrenia, the GluN1 knockdown mouse model and mutant disrupted in schizophrenia 1 (DISC1) mouse model, as well as inducible pluripotent stem cells (iPSCs) from a schizophrenia subject with the DISC1 mutation. We found increased lactate in the dorsolateral prefrontal cortex (p = 0.043, n = 16/group) in schizophrenia, as well as in frontal cortical neurons differentiated from a subject with schizophrenia with the DISC1 mutation (p = 0.032). We also found a decrease in lactate in mice with induced expression of mutant human DISC1 specifically in astrocytes (p = 0.049). These results build upon the body of evidence supporting bioenergetic dysfunction in schizophrenia, and suggests changes in lactate are a key feature of this often devastating severe mental illness.
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Encéfalo/metabolismo , Células Madre Pluripotentes Inducidas/metabolismo , Lactatos/metabolismo , Esquizofrenia/metabolismo , Animales , Astrocitos/metabolismo , Encéfalo/citología , Diagnóstico , Modelos Animales de Enfermedad , Lóbulo Frontal/citología , Lóbulo Frontal/metabolismo , Humanos , Células Madre Pluripotentes Inducidas/citología , Masculino , Ratones , Mutación , Proteínas del Tejido Nervioso/metabolismo , Neuronas/citología , Neuronas/metabolismo , Corteza Prefrontal/citología , Corteza Prefrontal/metabolismo , Ratas , Ratas Sprague-DawleyRESUMEN
Objectives: The cystine/glutamate antiporter (system xc-) is believed to contribute to nonvesicular glutamate release from glial cells in various brain areas. Although recent investigations implicate system xc- in mood disorders, unambiguous evidence has not yet been established. Therefore, we evaluated the possible role of system xc- in the depressive state. Methods: We conducted a protein expression analysis of the specific subunit of system xc- (xCT) in brain regions of the corticosterone mouse model, Flinders Sensitive Line rat model and post-mortem tissue of depressed patients. We next subjected system xc- deficient mice to the corticosterone model and analysed their behaviour in several tests. Lastly, we subjected additional cohorts of xCT-deficient and wild-type mice to N-acetylcysteine treatment to unveil whether the previously reported antidepressant-like effects are dependent upon system xc-. Results: We did not detect any changes in xCT expression levels in the animal models or patients compared to proper controls. Furthermore, loss of system xc- had no effect on depression- and anxiety-like behaviour. Finally, the antidepressant-like effects of N-acetylcysteine are not mediated via system xc-. Conclusions: xCT protein expression is not altered in the depressed brain and system xc- deficiency does not affect depression-associated behaviour in the corticosterone mouse model.
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Sistema de Transporte de Aminoácidos y+/deficiencia , Encéfalo/metabolismo , Depresión/genética , Depresión/fisiopatología , Anciano de 80 o más Años , Sistema de Transporte de Aminoácidos y+/genética , Animales , Antiinflamatorios , Encéfalo/patología , Corticosterona , Modelos Animales de Enfermedad , Conducta Exploratoria , Femenino , Humanos , Masculino , Ratones , Ratones Endogámicos C57BL , Actividad Motora , RatasRESUMEN
We utilized a cell-level approach to examine glycolytic pathways in the DLPFC of subjects with schizophrenia (n = 16) and control (n = 16) and found decreased mRNA expression of glycolytic enzymes in pyramidal neurons, but not astrocytes. To replicate these novel bioenergetic findings, we probed independent datasets for bioenergetic targets and found similar abnormalities. Next, we used a novel strategy to build a schizophrenia bioenergetic profile by a tailored application of the Library of Integrated Network-Based Cellular Signatures data portal (iLINCS) and investigated connected cellular pathways, kinases, and transcription factors using Enrichr. Finally, with the goal of identifying drugs capable of "reversing" the bioenergetic schizophrenia signature, we performed a connectivity analysis with iLINCS and identified peroxisome proliferator-activated receptor (PPAR) agonists as promising therapeutic targets. We administered a PPAR agonist to the GluN1 knockdown model of schizophrenia and found it improved long-term memory. Taken together, our findings suggest that tailored bioinformatics approaches, coupled with the LINCS library of transcriptional signatures of chemical and genetic perturbagens, may be employed to identify novel treatment strategies for schizophrenia and related diseases.
Asunto(s)
Metabolismo Energético , Redes Reguladoras de Genes , Esquizofrenia/metabolismo , Esquizofrenia/terapia , Animales , Análisis por Conglomerados , Regulación hacia Abajo/efectos de los fármacos , Regulación hacia Abajo/genética , Descubrimiento de Drogas , Metabolismo Energético/efectos de los fármacos , Técnicas de Silenciamiento del Gen , Redes Reguladoras de Genes/efectos de los fármacos , Humanos , Captura por Microdisección con Láser , Masculino , Ratones , Actividad Motora/efectos de los fármacos , Proteínas del Tejido Nervioso/metabolismo , Pioglitazona/farmacología , Inhibición Prepulso/efectos de los fármacos , Células Piramidales/efectos de los fármacos , Células Piramidales/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Reflejo de Sobresalto/efectos de los fármacos , Reproducibilidad de los Resultados , Esquizofrenia/genética , Esquizofrenia/fisiopatología , Conducta Estereotipada/efectos de los fármacos , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/genéticaRESUMEN
Protein kinases orchestrate signal transduction pathways involved in central nervous system functions ranging from neurodevelopment to synaptic transmission and plasticity. Abnormalities in kinase-mediated signaling are involved in the pathophysiology of neurological disorders, including neuropsychiatric disorders. Here, we expand on the hypothesis that kinase networks are dysregulated in schizophrenia. We investigated changes in serine/threonine kinase activity in cortical excitatory neurons differentiated from induced pluripotent stem cells (iPSCs) from a schizophrenia patient presenting with a 4 bp mutation in the disrupted in schizophrenia 1 (DISC1) gene and a corresponding control. Using kinome peptide arrays, we demonstrate large scale abnormalities in DISC1 cells, including a global depression of serine/threonine kinase activity, and changes in activity of kinases, including AMP-activated protein kinase (AMPK), extracellular signal-regulated kinases (ERK), and thousand-and-one amino acid (TAO) kinases. Using isogenic cell lines in which the DISC1 mutation is either introduced in the control cell line, or rescued in the schizophrenia cell line, we ascribe most of these changes to a direct effect of the presence of the DISC1 mutation. Investigating the gene expression signatures downstream of the DISC1 kinase network, and mapping them on perturbagen signatures obtained from the Library of Integrated Network-based Cellular Signatures (LINCS) database, allowed us to propose novel drug targets able to reverse the DISC1 kinase dysregulation gene expression signature. Altogether, our findings provide new insight into abnormalities of kinase networks in schizophrenia and suggest possible targets for disease intervention.
Asunto(s)
Células Madre Pluripotentes Inducidas/metabolismo , Proteínas del Tejido Nervioso/metabolismo , Esquizofrenia/metabolismo , Simulación por Computador , Humanos , Modelos Biológicos , Mutación , Proteínas del Tejido Nervioso/genética , Neuronas , Transducción de Señal , Sinapsis/fisiología , Transmisión SinápticaRESUMEN
Parkinson's disease (PD) is an age-related neurodegenerative condition characterized by a progressive loss of dopaminergic neurons in the substantia nigra pars compacta (SNpc). A loss of proteasome function participates to the pathogenesis of PD, leading to the development of rodent models in which a proteasome inhibitor is applied to the nigrostriatal pathway. We recently characterized the intranigral lactacystin (LAC) mouse model, leading to nigrostriatal degeneration, motor dysfunction and alpha-synuclein accumulation. In the present study, we compared the effect of two commonly used anesthetics for generating animal models of PD-i.e., ketamine (KET) and isoflurane (ISO)-on the vulnerability of mouse dopaminergic neurons to proteasome inhibition-induced degeneration. Both anesthetics have the potential to affect the susceptibility of the nigrostriatal pathway for toxin-induced degeneration, and are known to modulate dopamine (DA) homeostasis. Yet, their impact on nigrostriatal degeneration in the proteasome inhibition model has not been evaluated. Unilateral injection with LAC in the SNpc of mice induced motor impairment and significantly reduced the number of dopaminergic cells to ~55%, irrespective of the anesthetic used. However, LAC-induced striatal DA depletion was slightly affected by the choice of anesthetic, resulting in a significant increase in DA turnover in the ISO- but not in KET-treated mice. These results suggest that the extent of nigrostriatal dopaminergic neural loss caused by LAC is not influenced by the choice of anesthetic, and that compared to other PD models, KET is not neuroprotective in the LAC model.
RESUMEN
The pathological hallmarks of Parkinson's disease are the progressive loss of nigral dopaminergic neurons and the formation of intracellular inclusion bodies, termed Lewy bodies, in surviving neurons. Accumulation of proteins in large insoluble cytoplasmic aggregates has been proposed to result, partly, from a failure in the function of intracellular protein degradation pathways. Evidence in support for such a hypothesis emerged in the beginning of the years 2000 with studies demonstrating structural and functional deficits in the ubiquitin-proteasome pathway in post-mortem nigral tissue of patients with Parkinson's disease. These fundamental findings have inspired the development of a new generation of animal models based on the use of proteasome inhibitors to disturb protein homeostasis and trigger nigral dopaminergic neurodegeneration. In this review, we provide an updated overview of the current approaches in employing proteasome inhibitors to model Parkinson's disease, with particular emphasis on rodent studies. In addition, the mechanisms underlying proteasome inhibition-induced cell death and the validity criteria (construct, face and predictive validity) of the model will be critically discussed. Due to its distinct, but highly relevant mechanism of inducing neuronal death, the proteasome inhibition model represents a useful addition to the repertoire of toxin-based models of Parkinson's disease that might provide novel clues to unravel the complex pathogenesis of this disorder.
Asunto(s)
Modelos Animales de Enfermedad , Enfermedad de Parkinson/metabolismo , Complejo de la Endopetidasa Proteasomal/metabolismo , Inhibidores de Proteasoma , Animales , HumanosRESUMEN
Despite the numerous benefits of microwave applications in our daily life, microwaves were associated with diverse neurological complaints such as headaches and impaired sleep patterns, and changes in the electroencephalogram (EEG). To which extent microwaves influence the brain function remains unclear. This exploratory study assessed the behavior and neurochemistry in mice immediately or 4weeks after a 6-day exposure to low-intensity 10-GHz microwaves with an amplitude modulation (AM) of 2 or 8Hz. These modulation frequencies of 2 and 8Hz are situated within the delta and theta-alpha frequency bands in the EEG spectrum and are associated with sleep and active behavior, respectively. During these experiments, the specific absorbance rate was 0.3W/kg increasing the brain temperature with 0.23°C. For the first time, exposing mice to 8-Hz AM significantly reduced locomotor activity in an open field immediately after exposure which normalized after 4weeks. This in contrast to 2-Hz AM which didn't induce significant changes in locomotor activity immediately and 4weeks after exposure. Despite this difference in motor behavior, no significant changes in striatal dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) levels and DOPAC/DA turnover nor in cortical glutamate (GLU) concentrations were detected. In all cases, no effects on motor coordination on a rotarod, spatial working memory, anxiety nor depressive-like behavior were observed. The outcome of this study indicates that exposing mice to low-intensity 8-Hz AM microwaves can alter the locomotor activity in contrast to 2-Hz AM which did not affect the tested behaviors.