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1.
EMBO Rep ; 24(10): e56380, 2023 10 09.
Artigo em Inglês | MEDLINE | ID: mdl-37548091

RESUMO

Oxidative phosphorylation and glycolysis are the dominant ATP-generating pathways in mammalian metabolism. The balance between these two pathways is often shifted to execute cell-specific functions in response to stimuli that promote activation, proliferation, or differentiation. However, measurement of these metabolic switches has remained mostly qualitative, making it difficult to discriminate between healthy, physiological changes in energy transduction or compensatory responses due to metabolic dysfunction. We therefore present a broadly applicable method to calculate ATP production rates from oxidative phosphorylation and glycolysis using Seahorse XF Analyzer data and empirical conversion factors. We quantify the bioenergetic changes observed during macrophage polarization as well as cancer cell adaptation to in vitro culture conditions. Additionally, we detect substantive changes in ATP utilization upon neuronal depolarization and T cell receptor activation that are not evident from steady-state ATP measurements. This method generates a single readout that allows the direct comparison of ATP produced from oxidative phosphorylation and glycolysis in live cells. Additionally, the manuscript provides a framework for tailoring the calculations to specific cell systems or experimental conditions.


Assuntos
Smegmamorpha , Animais , Smegmamorpha/metabolismo , Mitocôndrias/metabolismo , Metabolismo Energético , Glicólise , Fosforilação Oxidativa , Trifosfato de Adenosina/metabolismo , Mamíferos/metabolismo
2.
Cancer Cell ; 41(6): 1048-1060.e9, 2023 06 12.
Artigo em Inglês | MEDLINE | ID: mdl-37236196

RESUMO

Malignant tumors exhibit heterogeneous metabolic reprogramming, hindering the identification of translatable vulnerabilities for metabolism-targeted therapy. How molecular alterations in tumors promote metabolic diversity and distinct targetable dependencies remains poorly defined. Here we create a resource consisting of lipidomic, transcriptomic, and genomic data from 156 molecularly diverse glioblastoma (GBM) tumors and derivative models. Through integrated analysis of the GBM lipidome with molecular datasets, we identify CDKN2A deletion remodels the GBM lipidome, notably redistributing oxidizable polyunsaturated fatty acids into distinct lipid compartments. Consequently, CDKN2A-deleted GBMs display higher lipid peroxidation, selectively priming tumors for ferroptosis. Together, this study presents a molecular and lipidomic resource of clinical and preclinical GBM specimens, which we leverage to detect a therapeutically exploitable link between a recurring molecular lesion and altered lipid metabolism in GBM.


Assuntos
Ferroptose , Glioblastoma , Metabolismo dos Lipídeos , Humanos , Inibidor p16 de Quinase Dependente de Ciclina/genética , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Ferroptose/genética , Ferroptose/fisiologia , Perfilação da Expressão Gênica , Glioblastoma/genética , Glioblastoma/metabolismo , Glioblastoma/patologia , Metabolismo dos Lipídeos/genética , Metabolismo dos Lipídeos/fisiologia , Recidiva Local de Neoplasia
3.
NMR Biomed ; 36(6): e4785, 2023 06.
Artigo em Inglês | MEDLINE | ID: mdl-35704275

RESUMO

Amine-weighted chemical exchange saturation transfer (CEST) magnetic resonance imaging (MRI) is particularly valuable as an amine- and pH-sensitive imaging technique in brain tumors, targeting the intrinsically high concentration of amino acids with exchangeable amine protons and reduced extracellular pH in brain tumors. Amine-weighted CEST MRI contrast is dependent on the glioma genotype, likely related to differences in degree of malignancy and metabolic behavior. Amine-weighted CEST MRI may provide complementary value to anatomic imaging in conventional and exploratory therapies in brain tumors, including chemoradiation, antiangiogenic therapies, and immunotherapies. Continual improvement and clinical testing of amine-weighted CEST MRI has the potential to greatly impact patients with brain tumors by understanding vulnerabilities in the tumor microenvironment that may be therapeutically exploited.


Assuntos
Aminas , Neoplasias Encefálicas , Humanos , Aminas/química , Concentração de Íons de Hidrogênio , Imageamento por Ressonância Magnética/métodos , Neoplasias Encefálicas/diagnóstico por imagem , Neoplasias Encefálicas/química , Prótons , Microambiente Tumoral
4.
Cell Chem Biol ; 29(11): 1588-1600.e7, 2022 11 17.
Artigo em Inglês | MEDLINE | ID: mdl-36306785

RESUMO

Cancer cells need a steady supply of nutrients to evade cell death and proliferate. Depriving cancer cells of the amino acid cystine can trigger the non-apoptotic cell death process of ferroptosis. Here, we report that cancer cells can evade cystine deprivation-induced ferroptosis by uptake and catabolism of the cysteine-rich extracellular protein albumin. This protective mechanism is enhanced by mTORC1 inhibition and involves albumin degradation in the lysosome, predominantly by cathepsin B (CTSB). CTSB-dependent albumin breakdown followed by export of cystine from the lysosome via the transporter cystinosin fuels the synthesis of glutathione, which suppresses lethal lipid peroxidation. When cancer cells are grown under non-adherent conditions as spheroids, mTORC1 pathway activity is reduced, and albumin supplementation alone affords considerable protection against ferroptosis. These results identify the catabolism of extracellular protein within the lysosome as a mechanism that can inhibit ferroptosis in cancer cells.


Assuntos
Ferroptose , Cistina , Glutationa/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina , Lisossomos/metabolismo , Albuminas , Linhagem Celular Tumoral
5.
Neuroimage Clin ; 32: 102882, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34911188

RESUMO

PURPOSE: To quantify abnormal metabolism of diffuse gliomas using "aerobic glycolytic imaging" and investigate its biological correlation. METHODS: All subjects underwent a pH-weighted amine chemical exchange saturation transfer spin-and-gradient-echo echoplanar imaging (CEST-SAGE-EPI) and dynamic susceptibility contrast perfusion MRI. Relative oxygen extraction fraction (rOEF) was estimated as the ratio of reversible transverse relaxation rate R2' to normalized relative cerebral blood volume. An aerobic glycolytic index (AGI) was derived by the ratio of pH-weighted image contrast (MTRasym at 3.0 ppm) to rOEF. AGI was compared between different tumor types (N = 51, 30 IDH mutant and 21 IDH wild type). Metabolic MR parameters were correlated with 18F-FDG uptake (N = 8, IDH wild-type glioblastoma), expression of key glycolytic proteins using immunohistochemistry (N = 38 samples, 21 from IDH mutant and 17 from IDH wild type), and bioenergetics analysis on purified tumor cells (N = 7, IDH wild-type high grade). RESULTS: AGI was significantly lower in IDH mutant than wild-type gliomas (0.48 ± 0.48 vs. 0.70 ± 0.48; P = 0.03). AGI was strongly correlated with 18F-FDG uptake both in non-enhancing tumor (Spearman, ρ = 0.81; P = 0.01) and enhancing tumor (ρ = 0.81; P = 0.01). AGI was significantly correlated with glucose transporter 3 (ρ = 0.71; P = 0.004) and hexokinase 2 (ρ = 0.73; P = 0.003) in IDH wild-type glioma, and monocarboxylate transporter 1 (ρ = 0.59; P = 0.009) in IDH mutant glioma. Additionally, a significant correlation was found between AGI derived from bioenergetics analysis and that estimated from MRI (ρ = 0.79; P = 0.04). CONCLUSION: AGI derived from molecular MRI was correlated with glucose uptake (18F-FDG and glucose transporter 3/hexokinase 2) and cellular AGI in IDH wild-type gliomas, whereas AGI in IDH mutant gliomas appeared associated with monocarboxylate transporter density.


Assuntos
Neoplasias Encefálicas , Glioma , Neoplasias Encefálicas/diagnóstico por imagem , Neoplasias Encefálicas/genética , Glioma/diagnóstico por imagem , Glioma/genética , Humanos , Concentração de Íons de Hidrogênio , Imageamento por Ressonância Magnética , Mutação , Oxigênio , Perfusão
6.
Neurosurg Clin N Am ; 32(2): 159-169, 2021 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-33781499

RESUMO

Glioblastomas (GBMs) exhibit altered metabolism to support a variety of bioenergetic and biosynthetic demands for tumor growth, invasion, and drug resistance. Changes in glycolytic flux, oxidative phosphorylation, the pentose phosphate pathway, fatty acid biosynthesis and oxidation, and nucleic acid biosynthesis are observed in GBMs to help drive tumorigenesis. Both the genetic landscape of GBMs and the unique brain tumor microenvironment shape metabolism; therefore, an understanding of how both intrinsic and extrinsic factors modulate metabolism is becoming increasingly important for finding effect targets and therapeutics for GBM.


Assuntos
Neoplasias Encefálicas , Glioblastoma , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/genética , Glioblastoma/tratamento farmacológico , Glioblastoma/genética , Glicólise , Humanos , Microambiente Tumoral
7.
J Biol Chem ; 295(35): 12330-12342, 2020 08 28.
Artigo em Inglês | MEDLINE | ID: mdl-32620552

RESUMO

GABA type A receptors (GABAARs) mediate fast synaptic inhibition and are trafficked to functionally diverse synapses. However, the precise molecular mechanisms that regulate the synaptic targeting of these receptors are unclear. Whereas it has been previously shown that phosphorylation events in α4, ß, and γ subunits of GABAARs govern their function and trafficking, phosphorylation of other subunits has not yet been demonstrated. Here, we show that the α2 subunit of GABAARs is phosphorylated at Ser-359 and enables dynamic regulation of GABAAR binding to the scaffolding proteins gephyrin and collybistin. We initially identified Ser-359 phosphorylation by MS analysis, and additional experiments revealed that it is regulated by the activities of cAMP-dependent protein kinase (PKA) and the protein phosphatase 1 (PP1) and/or PP2A. GST-based pulldowns and coimmunoprecipitation experiments demonstrate preferential binding of both gephyrin and collybistin to WT and an S359A phosphonull variant, but not to an S359D phosphomimetic variant. Furthermore, the decreased capacity of the α2 S359D variant to bind collybistin and gephyrin decreased the density of synaptic α2-containing GABAAR clusters and caused an absence of α2 enrichment in the axon initial segment. These results suggest that PKA-mediated phosphorylation and PP1/PP2A-dependent dephosphorylation of the α2 subunit play a role in the dynamic regulation of GABAAR accumulation at inhibitory synapses, thereby regulating the strength of synaptic inhibition. The MS data have been deposited to ProteomeXchange, with the data set identifier PXD019597.


Assuntos
Regulação para Baixo , Potenciais Pós-Sinápticos Inibidores , Receptores de GABA-A/metabolismo , Sinapses/metabolismo , Substituição de Aminoácidos , Animais , Proteínas Quinases Dependentes de AMP Cíclico/genética , Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Transgênicos , Mutação de Sentido Incorreto , Fosforilação , Proteína Fosfatase 2/genética , Proteína Fosfatase 2/metabolismo , Ratos , Ratos Wistar , Receptores de GABA-A/genética , Fatores de Troca de Nucleotídeo Guanina Rho/genética , Fatores de Troca de Nucleotídeo Guanina Rho/metabolismo , Sinapses/genética
8.
Int J Group Psychother ; 68(3): 428-457, 2018 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38449148

RESUMO

The zeitgeist for brief services psychotherapy efficacy is well underway within the individual and family therapy treatment modalities. However, this paradigm shift, to produce clinically significant mental health outcomes in a much shorter time, has evolved to a much lesser degree within the treatment group format. Longer-term treatment group protocols typically do not match treatment-seeking behaviors with high dropout rates for clients. The current authors describe a structured, four-session treatment protocol that integrates the tenets of Externalizing Metaphors Therapy (EMT) with Innovative Moments (IMs) in addressing anxiety for children and youth. EMT is based upon the externalization of problems, transformation of metaphoric imagery, and the shifting of underlying maladaptive emotional schemas. It is suggested that treatment outcomes are enhanced through the integration of three IMs between-session exercises.

10.
J Biol Chem ; 291(23): 12394-407, 2016 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-27044742

RESUMO

The accumulation of γ-aminobutyric acid receptors (GABAARs) at the appropriate postsynaptic sites is critical for determining the efficacy of fast inhibitory neurotransmission. Although we know that the majority of synaptic GABAAR subtypes are assembled from α1-3, ß, and γ2 subunits, our understanding of how neurons facilitate their targeting to and stabilization at inhibitory synapses is rudimentary. To address these issues, we have created knock-in mice in which the pH-sensitive green fluorescent protein (GFP) and the Myc epitope were introduced to the extracellular domain of the mature receptor α2 subunit (pHα2). Using immunoaffinity purification and mass spectroscopy, we identified a stable complex of 174 proteins that were associated with pHα2, including other GABAAR subunits, and previously identified receptor-associated proteins such as gephyrin and collybistin. 149 of these proteins were novel GABAAR binding partners and included G-protein-coupled receptors and ion channel subunits, proteins that regulate trafficking and degradation, regulators of protein phosphorylation, GTPases, and a number of proteins that regulate their activity. Notably, members of the postsynaptic density family of proteins that are critical components of excitatory synapses were not associated with GABAARs. Crucially, we demonstrated for a subset of these novel proteins (including cullin1, ephexin, potassium channel tetramerization domain containing protein 12, mitofusin2, metabotropic glutamate receptor 5, p21-activated kinase 7, and Ras-related protein 5A) bind directly to the intracellular domains of GABAARs, validating our proteomic analysis. Thus, our experiments illustrate the complexity of the GABAAR proteome and enhance our understanding of the mechanisms neurons use to construct inhibitory synapses.


Assuntos
Proteínas de Fluorescência Verde/metabolismo , Proteoma/metabolismo , Proteômica/métodos , Receptores de GABA-A/metabolismo , Sinapses/metabolismo , Animais , Western Blotting , Fenômenos Eletrofisiológicos , Proteínas de Fluorescência Verde/genética , Células HEK293 , Hipocampo/metabolismo , Hipocampo/fisiologia , Humanos , Concentração de Íons de Hidrogênio , Potenciais Pós-Sinápticos Inibidores , Espectrometria de Massas , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neurônios/metabolismo , Neurônios/fisiologia , Proteoma/genética , Receptores de GABA-A/genética , Sinapses/fisiologia
11.
Front Cell Neurosci ; 10: 40, 2016.
Artigo em Inglês | MEDLINE | ID: mdl-26973459

RESUMO

The GABAergic neuroactive steroid (3α,5α)-3-hydroxy-pregnan-20-one (3α,5α-THP, allopregnanolone) is decreased in various brain regions of C57BL/6J mice following exposure to an acute stressor or chronic intermittent ethanol (CIE) exposure and withdrawal. It is well established that there are complex interactions between stress and ethanol drinking, with mixed literature regarding the effects of stress on ethanol intake. However, there is little research examining how chronic ethanol exposure alters stress responses. The present work examined the impact of CIE exposure and withdrawal on changes in brain levels of 3α,5α-THP, as well as hormonal and behavioral responses to forced swim stress (FSS). Adult male C57BL/6J mice were exposed to four cycles of CIE to induce ethanol dependence. Following 8 h or 72 h withdrawal, mice were subjected to FSS for 10 min, and 50 min later brains were collected for immunohistochemical analysis of cellular 3α,5α-THP. Behavioral and circulating corticosterone responses to FSS were quantified. Following 8 h withdrawal, ethanol exposure potentiated the corticosterone response to FSS. Following 72 h withdrawal, this difference was no longer observed. Following 8 h withdrawal, stress-exposed mice showed no differences in immobility, swimming or struggling behavior. However, following 72 h withdrawal, ethanol-exposed mice showed less immobility and greater swimming behavior compared to air-exposed mice. Interestingly, cellular 3α,5α-THP levels were increased in the lateral amygdala 8 h and 72 h post-withdrawal in stressed ethanol-exposed mice compared to ethanol-exposed/non-stressed mice. In the paraventricular nucleus of the hypothalamus, stress exposure decreased 3α,5α-THP levels compared to controls following 72 h withdrawal, but no differences were observed 8 h post-withdrawal. There were no differences in cellular 3α,5α-THP levels in the nucleus accumbens shell at either withdrawal time point. These data suggest that there are different mechanisms mediating hormonal, behavioral, and brain responses to stress following CIE exposure. The lateral amygdala appears to be an extremely sensitive brain region exhibiting changes in cellular 3α,5α-THP levels following CIE and exposure to swim stress. It is likely that these changes in cellular 3α,5α-THP levels in the lateral amygdala contribute to the behavioral effects observed following 72 h withdrawal.

12.
J Neurosci ; 35(21): 8291-6, 2015 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-26019342

RESUMO

GABA(A) receptors form Cl(-) permeable channels that mediate the majority of fast synaptic inhibition in the brain. The K(+)/Cl(-) cotransporter KCC2 is the main mechanism by which neurons establish low intracellular Cl(-) levels, which is thought to enable GABAergic inhibitory control of neuronal activity. However, the widely used KCC2 inhibitor furosemide is nonselective with antiseizure efficacy in slices and in vivo, leading to a conflicting scheme of how KCC2 influences GABAergic control of neuronal synchronization. Here we used the selective KCC2 inhibitor VU0463271 [N-cyclopropyl-N-(4-methyl-2-thiazolyl)-2-[(6-phenyl-3-pyridazinyl)thio]acetamide] to investigate the influence of KCC2 function. Application of VU0463271 caused a reversible depolarizing shift in E(GABA) values and increased spiking of cultured hippocampal neurons. Application of VU0463271 to mouse hippocampal slices under low-Mg(2+) conditions induced unremitting recurrent epileptiform discharges. Finally, microinfusion of VU0463271 alone directly into the mouse dorsal hippocampus rapidly caused epileptiform discharges. Our findings indicated that KCC2 function was a critical inhibitory factor ex vivo and in vivo.


Assuntos
Hipocampo/fisiologia , Inibidores de Simportadores de Cloreto de Sódio e Potássio/farmacologia , Simportadores/antagonistas & inibidores , Simportadores/fisiologia , Animais , Animais Recém-Nascidos , Células Cultivadas , Células HEK293 , Hipocampo/efeitos dos fármacos , Humanos , Potenciais da Membrana/efeitos dos fármacos , Potenciais da Membrana/fisiologia , Camundongos , Camundongos Endogâmicos C57BL , Técnicas de Cultura de Órgãos , Ratos , Ratos Sprague-Dawley , Transmissão Sináptica/efeitos dos fármacos , Transmissão Sináptica/fisiologia , Cotransportadores de K e Cl-
13.
Alcohol Clin Exp Res ; 38(10): 2561-71, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25293837

RESUMO

BACKGROUND: The GABAergic neuroactive steroid (3α,5α)-3-hydroxy-pregnan-20-one (3α,5α-THP; allopregnanolone) has been studied during withdrawal from ethanol (EtOH) in humans, rats, and mice. Serum 3α,5α-THP levels decreased, and brain levels were not altered following acute EtOH administration (2 g/kg) in male C57BL/6J mice; however, the effects of chronic intermittent ethanol (CIE) exposure on 3α,5α-THP levels have not been examined. Given that CIE exposure changes subsequent voluntary EtOH drinking in a time-dependent fashion following repeated cycles of EtOH exposure, we conducted a time-course analysis of CIE effects on 3α,5α-THP levels in specific brain regions known to influence drinking behavior. METHODS: Adult male C57BL/6J mice were exposed to 4 cycles of CIE to induce EtOH dependence. All mice were sacrificed and perfused at 1 of 2 time points, 8 or 72 hours following the final exposure cycle. Free-floating brain sections (40 µm; 3 to 5 sections/region/animal) were immunostained and analyzed to determine relative levels of cellular 3α,5α-THP. RESULTS: Withdrawal from CIE exposure produced time-dependent and region-specific effects on immunohistochemical detection of 3α,5α-THP levels across cortical and limbic brain regions. A transient reduction in 3α,5α-THP immunoreactivity was observed in the central nucleus of the amygdala 8 hours after withdrawal from CIE (-31.4 ± 9.3%). Decreases in 3α,5α-THP immunoreactivity were observed 72 hours following withdrawal in the medial prefrontal cortex (-25.0 ± 9.3%), nucleus accumbens core (-29.9 ± 6.6%), and dorsolateral striatum (-18.5 ± 6.0%), while an increase was observed in the CA3 pyramidal cell layer of the hippocampus (+42.8 ± 19.5%). Sustained reductions in 3α,5α-THP immunoreactivity were observed at both time points in the lateral amygdala (8 hours -28.3 ± 12.8%; 72 hours -27.5 ± 12.4%) and in the ventral tegmental area (8 hours -26.5 ± 9.9%; 72 hours -31.6 ± 13.8%). CONCLUSIONS: These data suggest that specific neuroadaptations in 3α,5α-THP levels may be present in regions of brain that mediate anxiety, stress, and reinforcement relevant to EtOH dependence. The changes that occur at different time points likely modulate neurocircuitry involved in EtOH withdrawal as well as the elevated drinking observed after CIE exposure.


Assuntos
Núcleo Central da Amígdala/metabolismo , Etanol/administração & dosagem , Etanol/farmacologia , Núcleo Accumbens/metabolismo , Córtex Pré-Frontal/metabolismo , Pregnanolona/metabolismo , Suspensão de Tratamento , Alcoolismo/fisiopatologia , Animais , Ansiedade/fisiopatologia , Corpo Estriado/metabolismo , Relação Dose-Resposta a Droga , Hipocampo/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Modelos Animais , Estresse Psicológico/fisiopatologia , Fatores de Tempo
14.
Psychopharmacology (Berl) ; 231(17): 3281-92, 2014 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-24744202

RESUMO

RATIONALE: Stress activates the hypothalamic-pituitary-adrenal (HPA) axis, and GABAergic neuroactive steroids contribute to homeostatic regulation of this circuitry. Acute forced swim stress (FSS) increases plasma, cortical, and hypothalamic (3α,5α)-3-hydroxy-pregnan-20-one (3α,5α-THP) levels in rats. However, there have not been systemic investigations of acute stress on changes in plasma and brain levels of 3α,5α-THP in mouse models. OBJECTIVES: The present experiments aimed to assess circulating and local brain levels of 3α,5α-THP following acute FSS in C57BL/6J mice. METHODS: Mice were exposed to FSS (10 min), and 50 min later, blood and brains were collected. Circulating pregnenolone and 3α,5α-THP levels were assessed in serum. Free-floating brain sections (40 µm, four to five sections/region) were immunostained and analyzed in cortical and limbic brain structures. RESULTS: FSS decreased circulating 3α,5α-THP (-41.6 ± 10.4 %) and reduced 3α,5α-THP immunolabeling in the paraventricular nucleus of the hypothalamus (-15.2 ± 5.7 %), lateral amygdala (LA, -31.1 ± 13.4 %), and nucleus accumbens (NAcc) shell (-31.9 ± 14.6). Within the LA, vesicular glutamate transporter 1 (VGLUT1) and vesicular GABA transporter were localized in 3α,5α-THP-positively stained cells, while in the NAcc shell, only VGLUT1 was localized in 3α,5α-THP-positively stained cells, suggesting that both glutamatergic and GABAergic cells within the LA are 3α,5α-THP-positive, while in the NAcc shell, 3α,5α-THP only localizes to glutamatergic cells. CONCLUSIONS: The decrease in circulating and brain levels of 3α,5α-THP may be due to alterations in the biosynthesis/metabolism or changes in the regulation of the HPA axis following FSS. Changes in GABAergic neuroactive steroids in response to stress likely mediate functional adaptations in neuronal activity. This may provide a potential targeted therapeutic avenue to address maladaptive stress responsivity.


Assuntos
Sistema Límbico/metabolismo , Pregnanolona/metabolismo , Estresse Psicológico/metabolismo , Natação/psicologia , Glândulas Suprarrenais/enzimologia , Glândulas Suprarrenais/metabolismo , Animais , Química Encefálica/efeitos dos fármacos , Corticosterona/sangue , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Estresse Psicológico/psicologia , Proteína Vesicular 1 de Transporte de Glutamato/metabolismo , Proteínas Vesiculares de Transporte de Aminoácidos Inibidores/metabolismo
15.
Neuropsychopharmacology ; 39(8): 1978-87, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-24566803

RESUMO

The neuroactive steroid (3α,5α)-3-hydroxypregnan-20-one (3α,5α-THP or allopregnanolone) is a positive modulator of GABAA receptors synthesized in the brain, adrenal glands, and gonads. In rats, ethanol activates the hypothalamic-pituitary-adrenal axis and elevates 3α,5α-THP in plasma, cerebral cortex, and hippocampus. In vivo, these effects are dependent on both the pituitary and adrenal glands. In vitro, however, ethanol locally increases 3α,5α-THP in hippocampal slices, in the absence of adrenal influence. Therefore, it is not known whether ethanol can change local brain levels of 3α,5α-THP in vivo, independent of the adrenals. To directly address this controversy, we administered ethanol (2 g/kg) or saline to rats that underwent adrenalectomy (ADX) or received sham surgery and performed immunohistochemistry for 3α,5α-THP. In the medial prefrontal cortex (mPFC), ethanol increased 3α,5α-THP after sham surgery, compared with saline controls, with no ethanol-induced change in 3α,5α-THP following ADX. In subcortical regions, 3α,5α-THP was increased independent of adrenals in the CA1 pyramidal cell layer, dentate gyrus polymorphic layer, bed nucleus of the stria terminalis, and paraventricular nucleus of the hypothalamus. Furthermore, ethanol decreased 3α,5α-THP labeling in the nucleus accumbens shore and central nucleus of the amygdala, independent of the adrenal glands. These data indicate that ethanol dynamically regulates local 3α,5α-THP levels in several subcortical regions; however, the adrenal glands contribute to 3α,5α-THP elevations in the mPFC. Using double immunofluorescent labeling we determined that adrenal dependence of 3α,5α-THP induction by ethanol is not due to a lack of colocalization of 3α,5α-THP with the cholesterol transporters steroidogenic acute regulatory protein (StAR) or translocator protein (TSPO).


Assuntos
Encéfalo/efeitos dos fármacos , Encéfalo/metabolismo , Depressores do Sistema Nervoso Central/farmacologia , Etanol/farmacologia , Pregnanolona/metabolismo , Glândulas Suprarrenais/fisiologia , Adrenalectomia , Animais , Masculino , Córtex Pré-Frontal/metabolismo , Ratos , Ratos Wistar
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