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1.
Proc Natl Acad Sci U S A ; 108(16): 6650-5, 2011 Apr 19.
Artículo en Inglés | MEDLINE | ID: mdl-21464302

RESUMEN

Alcohol's deleterious effects on memory are well known. Acute alcohol-induced memory loss is thought to occur via inhibition of NMDA receptor (NMDAR)-dependent long-term potentiation in the hippocampus. We reported previously that ethanol inhibition of NMDAR function and long-term potentiation is correlated with a reduction in the phosphorylation of Tyr(1472) on the NR2B subunit and ethanol's inhibition of the NMDAR field excitatory postsynaptic potential was attenuated by a broad spectrum tyrosine phosphatase inhibitor. These data suggested that ethanol's inhibitory effect may involve protein tyrosine phosphatases. Here we demonstrate that the loss of striatal-enriched protein tyrosine phosphatase (STEP) renders NMDAR function, phosphorylation, and long-term potentiation, as well as fear conditioning, less sensitive to ethanol inhibition. Moreover, the ethanol inhibition was "rescued" when the active STEP protein was reintroduced into the cells. Taken together, our data suggest that STEP contributes to ethanol inhibition of NMDAR function via dephosphorylation of tyrosine sites on NR2B receptors and lend support to the hypothesis that STEP may be required for ethanol's amnesic effects.


Asunto(s)
Depresores del Sistema Nervioso Central/efectos adversos , Etanol/efectos adversos , Miedo/efectos de los fármacos , Aprendizaje/efectos de los fármacos , Potenciación a Largo Plazo/efectos de los fármacos , Proteínas Tirosina Fosfatasas no Receptoras/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Potenciales Sinápticos/efectos de los fármacos , Amnesia/inducido químicamente , Amnesia/enzimología , Amnesia/genética , Animales , Depresores del Sistema Nervioso Central/farmacología , Etanol/farmacología , Humanos , Potenciación a Largo Plazo/genética , Ratones , Ratones Noqueados , Fosforilación/efectos de los fármacos , Fosforilación/genética , Proteínas Tirosina Fosfatasas no Receptoras/genética , Receptores de N-Metil-D-Aspartato/genética , Potenciales Sinápticos/genética
2.
J Pharmacol Exp Ther ; 341(3): 611-6, 2012 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-22375069

RESUMEN

High abstinence rates characterize alcohol-dependent liver graft recipients. The immunosuppressants cyclosporine A (CsA) and tacrolimus (TRL) also inhibit calcineurin (CLN) in the brain. Previously, we found that CsA reduces alcohol consumption in C57BL/6J mice. The goals of the present study were: 1) to compare the ethanol preference effects of CsA against TRL, as well as sirolimus (SRL), an immunosuppressant without CLN inhibition and 2) to establish that reduction of alcohol consumption is not caused by caloric reinforcement from these ligands. C57BL/6J mice trained to imbibe ethanol consumed ethanol or sucrose in a modified limited-access drinking-in-the-dark paradigm; test groups received vehicle or doses of CsA (5-50 mg/kg), TRL (0.5-2.5 mg/kg), or SRL (1.0-5.0 mg/kg) for 5 consecutive days, 30 min before each 2-h limited-access session. Brain CsA, TRL, and SRL concentrations were measured. CsA (p < 0.001) and TRL (p < 0.01) each decreased ethanol consumption, whereas SRL showed no significant effects at any dose. Effective doses included CsA at 10 mg/kg and above and TRL at 2.5 mg/kg. CsA (50 mg/kg) did not reduce sucrose consumption. Both CsA and TRL reached significant brain concentrations compared with very low values of SRL. These data suggest that CsA and TRL may reduce alcohol preference through central CLN inhibition rather than by immunosuppression.


Asunto(s)
Consumo de Bebidas Alcohólicas/tratamiento farmacológico , Encéfalo/efectos de los fármacos , Calcineurina/efectos de los fármacos , Ciclosporina/farmacología , Inmunosupresores/farmacología , Sirolimus/farmacología , Tacrolimus/farmacología , Animales , Encéfalo/metabolismo , Relación Dosis-Respuesta a Droga , Interacciones Farmacológicas , Etanol , Ligandos , Masculino , Ratones , Ratones Endogámicos C57BL , Autoadministración , Sacarosa/administración & dosificación
3.
Mol Pharmacol ; 80(3): 529-37, 2011 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-21680777

RESUMEN

The hippocampal N-methyl-D-aspartate receptor (NMDAR) activity plays important roles in cognition and is a major substrate for ethanol-induced memory dysfunction. This receptor is a glutamate-gated ion channel, which is composed of NR1 and NR2 subunits in various brain areas. Although homomeric NR1 subunits form an active ion channel that conducts Na⁺ and Ca²âº currents, the incorporation of NR2 subunits allows this channel to be modulated by the Src family of kinases, phosphatases, and by simple molecules such as ethanol. We have found that short-term ethanol application inhibits the NMDAR activity via striatal enriched protein tyrosine phosphatase (STEP)-regulated mechanisms. The genetic deletion of the active form of STEP, STEP61, leads to marked attenuation of ethanol inhibition of NMDAR currents. In addition, STEP61 negatively regulates Fyn and p38 mitogen-activated protein kinase (MAPK), and these proteins are members of the NMDAR super molecular complex. Here we demonstrate, using whole-cell electrophysiological recording, Western blot analysis, and pharmacological manipulations, that neurons exposed to a 3-h, 45 mM ethanol treatment develop an adaptive attenuation of short-term ethanol inhibition of NMDAR currents in brain slices. Our results suggest that this adaptation of NMDAR responses is associated with a partial inactivation of STEP61, an activation of p38 MAPK, and a requirement for NR2B activity. Together, these data indicate that altered STEP61 and p38 MAPK signaling contribute to the modulation of ethanol inhibition of NMDARs in brain neurons.


Asunto(s)
Adaptación Fisiológica , Cuerpo Estriado/efectos de los fármacos , Etanol/farmacología , Proteínas Tirosina Fosfatasas/metabolismo , Receptores de N-Metil-D-Aspartato/efectos de los fármacos , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo , Animales , Cuerpo Estriado/enzimología , Cuerpo Estriado/metabolismo , Activación Enzimática , Masculino , Ratas , Ratas Sprague-Dawley , Receptores de N-Metil-D-Aspartato/fisiología
4.
J Neurosci ; 29(29): 9330-43, 2009 Jul 22.
Artículo en Inglés | MEDLINE | ID: mdl-19625523

RESUMEN

NMDA receptor (NMDAR)-mediated excitotoxicity plays an important role in several CNS disorders, including epilepsy, stroke, and ischemia. Here we demonstrate the involvement of striatal-enriched protein tyrosine phosphatase (STEP) in this critical process. STEP(61) is an alternatively spliced member of the family that is present in postsynaptic terminals. In an apparent paradox, STEP(61) regulates extracellular signal-regulated kinase 1/2 (ERK1/2) and p38, two proteins with opposing functions; activated p38 promotes cell death, whereas activated ERK1/2 promotes cell survival. We found that synaptic stimulation of NMDARs promoted STEP(61) ubiquitination and degradation, concomitant with ERK1/2 activation. In contrast, extrasynaptic stimulation of NMDARs invoked calpain-mediated proteolysis of STEP(61), producing the truncated cleavage product STEP(33) and activation of p38. The calpain cleavage site on STEP was mapped to the kinase interacting motif, a domain required for substrate binding. As a result, STEP(33) neither interacts with nor dephosphorylates STEP substrates. A synthetic peptide spanning the calpain cleavage site efficiently reduced STEP(61) degradation and attenuated p38 activation and cell death in slice models. Furthermore, this peptide was neuroprotective when neurons were subjected to excitotoxicity or cortical slices were exposed to ischemic conditions. These findings suggest a novel mechanism by which differential NMDAR stimulation regulates STEP(61) to promote either ERK1/2 or p38 activation and identifies calpain cleavage of STEP(61) as a valid target for the development of neuroprotective therapy.


Asunto(s)
Calpaína/metabolismo , Proteínas Tirosina Fosfatasas no Receptoras/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Empalme Alternativo , Animales , Encéfalo/fisiología , Encéfalo/fisiopatología , Muerte Celular/fisiología , Células Cultivadas , Quinasa 5 Dependiente de la Ciclina/metabolismo , Endocitosis/fisiología , Ácido Glutámico/toxicidad , Técnicas In Vitro , Ratones , Ratones Noqueados , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Neuronas/fisiología , Proteínas Tirosina Fosfatasas no Receptoras/genética , Ratas , Ratas Sprague-Dawley , Sinapsis/fisiología , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo
5.
J Neurochem ; 115(5): 1112-22, 2010 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-20831600

RESUMEN

Alcohol abuse leads to tolerance, dependence, and memory impairments that involve excitatory glutamatergic NMDA synaptic transmission. The NMDA receptor (NMDAR) is known to undergo activity-dependent adaptive functional changes. Since we observed that acute ethanol inhibition of the NMDAR was regulated by protein tyrosine phosphorylation, we investigated the role of protein tyrosine kinases and phosphatases on the NMDAR functions by chronic ethanol treatment. We carried out whole-cell recording, western blotting, and behavioral righting reflex measurements to assess the impact of chronic ethanol treatment on NMDAR function. Our results indicated that these receptors became resistant to the acute ethanol inhibition following chronic ethanol consumption. This resistance occurred without an increase in the NMDAR subunit expression but was associated with decreases in the levels of phospho-Y-1472 NR2B, increases in the levels of STEP33, increases in phospho-p38 mitogen-activated protein kinase (pp38 MAPK), and acquisition of tolerance to the sedative effects of ethanol. These data suggested that altered protein tyrosine phosphorylation of the NMDAR subunits significantly contributes to functional changes of this receptor by chronic ethanol ingestion. Therefore, preservation of the integrity of tyrosine phosphorylation mechanisms of the NMDAR may be important in controlling the progression of alcohol tolerance and dependence.


Asunto(s)
Depresores del Sistema Nervioso Central/farmacología , Etanol/farmacología , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Receptores de N-Metil-D-Aspartato/metabolismo , 2-Amino-5-fosfonovalerato/farmacología , Animales , Bencilaminas/farmacología , Bicuculina/análogos & derivados , Bicuculina/farmacología , Depresores del Sistema Nervioso Central/administración & dosificación , Etanol/administración & dosificación , Antagonistas de Aminoácidos Excitadores/farmacología , Potenciales Postsinápticos Excitadores/fisiología , Conducta Alimentaria/efectos de los fármacos , Antagonistas de Receptores de GABA-A/farmacología , Hipocampo/citología , Técnicas In Vitro , Masculino , Técnicas de Placa-Clamp/métodos , Ácidos Fosfínicos/farmacología , Fosforilación/efectos de los fármacos , Células Piramidales/efectos de los fármacos , Células Piramidales/fisiología , Ratas , Ratas Sprague-Dawley , Estadística como Asunto , Factores de Tiempo , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismo
6.
ILAR J ; 53(1): 4-13, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-23520595

RESUMEN

Substance addiction is a maladaptive behavior characterized by compulsive and uncontrolled self-administration of a substance (drug). Years of research indicate that addictive behavior is the result of complex interactions between the drug, the user, and the environment in which the drug is used; therefore, addiction cannot simply be attributed to the neurobiological actions of a drug. However, despite the obvious complexity of addictive behavior, animal models have both advanced understanding of addiction and contributed importantly to the development of medications to treat this disease. We briefly review recent animal models used to study drug addiction and the contribution of data generated by these animal models for the clinical treatment of addictive disorders.


Asunto(s)
Conducta Adictiva/fisiopatología , Animales , Humanos , Modelos Animales , Trastornos Relacionados con Sustancias/fisiopatología
7.
PLoS One ; 7(5): e37677, 2012.
Artículo en Inglés | MEDLINE | ID: mdl-22662188

RESUMEN

Every year, nearly 200,000 patients undergo radiation for brain tumors. For both patients and caregivers the most distressing adverse effect is impaired cognition. Efforts to protect against this debilitating effect have suffered from inadequate understanding of the cellular mechanisms of radiation damage. In the past it was accepted that radiation-induced normal tissue injury resulted from a progressive reduction in the survival of clonogenic cells. Moreover, because radiation-induced brain dysfunction is believed to evolve over months to years, most studies have focused on late changes in brain parenchyma. However, clinically, acute changes in cognition are also observed. Because neurons are fully differentiated post-mitotic cells, little information exists on the acute effects of radiation on synaptic function. The purpose of our study was to assess the potential acute effects of radiation on neuronal function utilizing ex vivo hippocampal brain slices. The cellular localization and functional status of excitatory and inhibitory neurotransmitter receptors was identified by immunoblotting. Electrophysiological recordings were obtained both for populations of neuronal cells and individual neurons. In the dentate gyrus region of isolated ex vivo slices, radiation led to early decreases in tyrosine phosphorylation and removal of excitatory N-methyl-D-aspartate receptors (NMDARs) from the cell surface while simultaneously increasing the surface expression of inhibitory gamma-aminobutyric acid receptors (GABA(A)Rs). These alterations in cellular localization corresponded with altered synaptic responses and inhibition of long-term potentiation. The non-competitive NMDAR antagonist memantine blocked these radiation-induced alterations in cellular distribution. These findings demonstrate acute effects of radiation on neuronal cells within isolated brain slices and open new avenues for study.


Asunto(s)
Neuronas/metabolismo , Neuronas/efectos de la radiación , Animales , Transporte Biológico/efectos de los fármacos , Caspasa 3/metabolismo , Activación Enzimática/efectos de la radiación , Potenciación a Largo Plazo/efectos de la radiación , Memantina/farmacología , N-Metilaspartato/metabolismo , Fosforilación/efectos de la radiación , Proteínas Tirosina Fosfatasas/metabolismo , Ratas , Ratas Sprague-Dawley , Receptores de GABA-A/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Transmisión Sináptica/efectos de la radiación
8.
Br J Pharmacol ; 162(6): 1351-63, 2011 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-21133888

RESUMEN

BACKGROUND AND PURPOSE: Tobacco and alcohol are often co-abused producing interactive effects in the brain. Although nicotine enhances memory while ethanol impairs it, variable cognitive changes have been reported from concomitant use. This study was designed to determine how nicotine and alcohol interact at synaptic sites to modulate neuronal processes. EXPERIMENTAL APPROACH: Acute effects of nicotine, ethanol, and both drugs on synaptic excitatory glutamatergic and inhibitory GABAergic transmission were measured using whole-cell recording in hippocampal CA1 pyramidal neurons from brain slices of mice on control or nicotine-containing diets. KEY RESULTS: Acute nicotine (50 nM) enhanced both GABAergic and glutamatergic synaptic transmission; potentiated GABA(A) receptor currents via activation of α7* and α4ß2* nAChRs, and increased N-methyl-D-aspartate (NMDA) and α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor currents through α7* receptors. While ethanol (80 mM) also increased GABA(A) currents, it inhibited NMDA currents. Although ethanol had no effect on AMPA currents, it blocked nicotine-induced increases in NMDA and AMPA currents. Following chronic nicotine treatment, acute nicotine or ethanol did not affect NMDA currents, while the effects of GABAergic responses were not altered. CONCLUSIONS AND IMPLICATIONS: Acute ethanol ingestion selectively attenuated nicotine enhancement of excitatory glutamatergic NMDA and AMPA receptor function, suggesting an overall reduction in excitatory output from the hippocampus. It also indicated that ethanol could decrease the beneficial effects of nicotine on memory performance. In addition, chronic nicotine treatment produced tolerance to the effects of nicotine and cross-tolerance to the effects of ethanol on glutamatergic activity, leading to a potential increase in the use of these drugs.


Asunto(s)
Región CA1 Hipocampal/efectos de los fármacos , Depresores del Sistema Nervioso Central/farmacología , Etanol/farmacología , Nicotina/farmacología , Agonistas Nicotínicos/farmacología , Células Piramidales/efectos de los fármacos , Receptores de GABA-A/metabolismo , Receptores de Glutamato/metabolismo , Animales , Región CA1 Hipocampal/metabolismo , Depresores del Sistema Nervioso Central/administración & dosificación , Depresores del Sistema Nervioso Central/metabolismo , Etanol/administración & dosificación , Etanol/metabolismo , Potenciales Postsinápticos Excitadores/efectos de los fármacos , Técnicas In Vitro , Potenciales Postsinápticos Inhibidores/efectos de los fármacos , Masculino , Ratones , Ratones Endogámicos C57BL , Nicotina/administración & dosificación , Nicotina/metabolismo , Agonistas Nicotínicos/administración & dosificación , Agonistas Nicotínicos/metabolismo , Técnicas de Placa-Clamp , Receptores AMPA/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Transmisión Sináptica/efectos de los fármacos , Factores de Tiempo
9.
J Pharmacol Exp Ther ; 312(3): 1082-9, 2005 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-15615867

RESUMEN

We tested the hypothesis that differential sensitivity to ethanol of synaptic GABA(A) somatic and dendritic inhibitory postsynaptic currents (IPSCs) in hippocampal CA1 pyramidal neurons could be due to differences in the extent of GABA(B) receptor activity at GABAergic synapses in these two hippocampal subfields. Our present results show that dendritic (distally evoked) GABA IPSCs contain a larger GABA(B) IPSC component of the total GABA IPSC than the somatic (proximally evoked) subfield. The inhibition of GABA(B) receptors by pretreatment of hippocampal slices with CGP-52432 [3[[(3,4-dichlorophenyl)methyl]amino]propyl](diethoxymethyl) phosphinic acid], a selective GABA(B) receptor antagonist, changes the basal ethanol-insensitive, distally evoked GABA(A) IPSCs to become more sensitive to ethanol. In addition, paired-pulse stimulation of the proximal and distal subfields of hippocampal pyramidal neurons shows that ethanol alone increases the probability of GABA release at proximal but not distal regions. Changes by ethanol on the probability of GABA release are only seen at distal locations during GABA(B) blockade. Finally, when the modulation of presynaptic GABA(B) receptors is minimized by the local application of 10 mM GABA directly onto somatic or dendritic GABAergic synaptic regions, postsynaptic GABA(B) receptors seem to exert significant negative (inhibiting) influence on the effects of ethanol on GABA(A) IPSCs in the distal subfields of CA1 pyramidal neurons. Together, our data suggest that differences in both presynaptic and postsynaptic GABA(B) receptor activity at these GABAergic synapses may modulate the differential ethanol sensitivity of proximal and distal GABA IPSCs(A) in hippocampal CA1 pyramidal neurons.


Asunto(s)
Etanol/farmacología , Hipocampo/efectos de los fármacos , Receptores de GABA-A/efectos de los fármacos , Receptores de GABA-B/efectos de los fármacos , Sinapsis/efectos de los fármacos , Animales , Bencilaminas/farmacología , Potenciales Evocados/efectos de los fármacos , Hipocampo/fisiología , Masculino , Inhibición Neural/efectos de los fármacos , Ácidos Fosfínicos/farmacología , Ratas , Ratas Sprague-Dawley , Sinapsis/fisiología
10.
Alcohol Clin Exp Res ; 28(9): 1277-83, 2004 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-15365296

RESUMEN

BACKGROUND: Ethanol enhances gamma-aminobutyric acid (GABA)A receptor-mediated responses in the brain, and this enhancement is greater in a mouse line behaviorally more sensitive to ethanol (long sleep) than in a line (short sleep) behaviorally less ethanol sensitive (assayed by loss of righting; sleep time). Quantitative trait locus (QTL) analysis of inbred long sleep (ILS) and inbred short sleep (ISS) phenotypes revealed four chromosomal regions (Lore1, Lore2, Lore4, and Lore5) that together account for approximately 50% of ethanol-induced sleep-time variance. Congenic strains were generated, each of which is homozygous for one of four ISS Lore QTLs on the ILS background. These congenic mouse strains are ideally suited for asking which QTL regions might correlate with other phenotypes that differ between ILS and ISS mice. Here we used the congenics to investigate altered GABAA responses to ethanol. METHODS: Evoked GABAA receptor-mediated inhibitory postsynaptic currents (IPSCs) were measured by whole-cell voltage-clamp recording procedures in CA1 pyramidal neurons in hippocampal brain slices. RESULTS: GABAA IPSC responses in hippocampal brain slices from ILS mice were significantly enhanced by 80 mM ethanol, whereas those from ISS mice were not affected. ILS.Lore2 and ILS.Lore5 congenic strains were significantly enhanced by 80 mM ethanol, similar to the background (control) ILS mice. However, ethanol had no significant effect on GABAA responses in ILS.Lore1 and ILS.Lore4 congenic mice, similar to the ISS mice, thus reflecting the influence of ISS alleles on the ILS phenotype. CONCLUSIONS: Our results suggest that alleles located in the Lore1 and Lore4 QTL regions confer ethanol sensitivity of GABAA receptor-mediated IPSCs. Thus, for these QTLs, GABAA IPSCs may represent an endophenotype of sedative/hypnotic sensitivity to ethanol. Although the Lore2 and Lore5 QTL regions have a significant effect on sleep time, they do not play a significant role in the differential ethanol enhancement of GABAA IPSCs between ILS and ISS mice.


Asunto(s)
Etanol/farmacología , Receptores de GABA-A/fisiología , Sinapsis/efectos de los fármacos , Transmisión Sináptica/efectos de los fármacos , Animales , Masculino , Ratones , Ratones Congénicos , Receptores de GABA-A/genética , Sinapsis/genética , Sinapsis/fisiología , Transmisión Sináptica/genética , Transmisión Sináptica/fisiología
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