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1.
Nat Med ; 7(1): 17-8, 2001 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-11135603

RESUMO

The clot-busting drug tissue plasminogen activator (tPA) is currently the only FDA-approved therapy for acute stroke. However, increasing evidence suggests that tPA can also contribute to excitotoxic neuronal damage in animal models of stroke.


Assuntos
Neurônios/efeitos dos fármacos , Acidente Vascular Cerebral/tratamento farmacológico , Ativador de Plasminogênio Tecidual/uso terapêutico , Animais , Modelos Animais de Doenças , Neurônios/patologia , Receptores de N-Metil-D-Aspartato/metabolismo , Acidente Vascular Cerebral/patologia
2.
Nat Med ; 4(2): 228-31, 1998 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-9461198

RESUMO

Intravenous tissue plasminogen activator (tPA) is used to treat acute stroke because of its thrombolytic activity and its ability to restore circulation to the brain. However, this protease also promotes neurodegeneration after intracerebral injection of excitotoxins such as glutamate, and neuronal damage after a cerebral infarct is thought to be mediated by excitotoxins. To investigate the effects of tPA on cerebral viability during ischemia/reperfusion, we occluded the middle cerebral artery in wild-type and tPA-deficient mice with an intravascular filament. This procedure allowed us to examine the role of tPA in ischemia, independent of its effect as a thrombolytic agent. tPA-deficient mice exhibited approximately 50% smaller cerebral infarcts than wild-type mice. Intravenous injection of tPA into tPA-/- or wild-type mice produced larger infarcts, indicating that tPA can increase stroke-induced injury. Since tPA promotes desirable (thrombolytic) as well as undesirable (neurotoxic) outcomes during stroke, future therapies should be aimed at countering the excitotoxic damage of tPA to afford even better neuroprotection after an acute cerebral infarct.


Assuntos
Isquemia Encefálica/patologia , Neurônios/efeitos dos fármacos , Ativador de Plasminogênio Tecidual/deficiência , Ativador de Plasminogênio Tecidual/farmacologia , Animais , Antígenos de Diferenciação/metabolismo , Encéfalo/efeitos dos fármacos , Encéfalo/patologia , Isquemia Encefálica/tratamento farmacológico , Circulação Cerebrovascular , Modelos Animais de Doenças , Hipocampo/efeitos dos fármacos , Hipocampo/patologia , Imuno-Histoquímica , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos , Microglia/efeitos dos fármacos , Ativador de Plasminogênio Tecidual/metabolismo
3.
J Exp Med ; 181(2): 735-45, 1995 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-7530762

RESUMO

Mononuclear phagocytes (monocytes, macrophages, and dendritic cells) play major roles in human immunodeficiency virus (HIV) persistence and disease pathogenesis. Macrophage antigen presentation and effector cell functions are impaired by HIV-1 infection. Abnormalities of macrophage effector cell function in bone marrow, lung, and brain likely result as a direct consequence of cellular activation and HIV replication. To further elucidate the extent of macrophage dysfunction in HIV-1 disease, a critical activation-specific regulatory molecule, nitric oxide (NO.), which may contribute to diverse pathology, was studied. Little, if any, NO. is produced by uninfected human monocytes. In contrast, infection with HIV-1 increases NO. production to modest, but significant levels (2-5 microM). Monocyte activation (with lipopolysaccharide, tumor necrosis factor alpha, or through interactions with astroglial cells) further enhances NO. production in HIV-infected cells, whereas its levels are diminished by interleukin 4. These results suggest a possible role for NO. in HIV-associated pathology where virus-infected macrophages are found. In support of this hypothesis, RNA encoding the inducible NO synthase (iNOS) was detected in postmortem brain tissue from one pediatric AIDS patient with advanced HIV encephalitis. Corresponding iNOS mRNA was not detected in brain tissue from five AIDS patients who died with less significant brain disease. These results demonstrate that HIV-1 can influence the expression of NOS in both cultured human monocytes and brain tissue. This newly described feature of HIV-macrophage interactions suggests previously unappreciated mechanisms of tissue pathology that result from productive viral replication.


Assuntos
Aminoácido Oxirredutases/metabolismo , Encefalopatias/enzimologia , HIV-1/fisiologia , Monócitos/virologia , Aminoácido Oxirredutases/genética , Astrócitos/virologia , Sequência de Bases , Encefalopatias/virologia , Células Cultivadas , Primers do DNA , Encefalite/enzimologia , Encefalite/virologia , Indução Enzimática , Humanos , Dados de Sequência Molecular , Monócitos/enzimologia , Óxido Nítrico/biossíntese , Óxido Nítrico Sintase , RNA/metabolismo , Células Tumorais Cultivadas
4.
Science ; 239(4845): 1293-6, 1988 Mar 11.
Artigo em Inglês | MEDLINE | ID: mdl-3344435

RESUMO

Functional nicotinic cholinergic receptors are found on mammalian retinal ganglion cell neurons in culture. The neurotransmitter acetylcholine (ACh) can be detected in the medium of many of these retinal cultures, after release presumably from the choline acetyltransferase-positive amacrine cells. The postsynaptic effect of endogenous or applied ACh on the ganglion cells can be blocked with specific nicotinic antagonists. Here it is shown that within 24 hours of producing such a pharmacologic blockade, the retinal ganglion cells begin to sprout or regenerate neuronal processes. Thus, the growth-enhancing effect of nicotinic antagonists may be due to the removal of inhibition to growth by tonic levels of ACh present in the culture medium. Since there is a spontaneous leak of ACh in the intact retina, the effects of nicotinic cholinergic drugs on process outgrowth in culture may reflect a normal control mechanism for growth or regeneration of retinal ganglion cell processes that is exerted by ACh in vivo.


Assuntos
Atropina/farmacologia , Mecamilamina/farmacologia , Receptores Nicotínicos/fisiologia , Retina/citologia , Células Ganglionares da Retina/citologia , Tubocurarina/farmacologia , Animais , Células Cultivadas , Picrotoxina/farmacologia , Ratos , Receptores Nicotínicos/efeitos dos fármacos , Células Ganglionares da Retina/efeitos dos fármacos
5.
Science ; 248(4953): 364-7, 1990 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-2326646

RESUMO

Coat protein gp120 from the human immunodeficiency virus type-1 (HIV-1) increased intracellular free calcium and injured rodent retinal ganglion cells and hippocampal neurons in culture. Highly purified recombinant gp120 envelope protein produced these effects in a dose-dependent fashion at picomolar concentrations. Immunoprecipitation with antibody to gp120, but not with control immunoglobulin-containing serum, depleted solutions of the viral envelope protein and also prevented both the rise in intracellular calcium and neuronal toxicity. The gp120-induced increase in intracellular calcium was abrogated by transiently lowering extracellular calcium or by adding the dihydropyridine calcium channel antagonist nimodipine (100 nM). Calcium channel antagonists also prevented gp120-induced neuronal injury. In addition, intracellular stores appeared to contribute substantially to the increase in calcium elicited by gp120. Since increases in intracellular calcium have been associated with neurotoxicity, it is possible that an injurious effect of gp120 on neurons might be related to this mechanism and that treatment with calcium channel antagonists may prove useful in mitigating HIV-1-related neuronal injury.


Assuntos
Bloqueadores dos Canais de Cálcio/farmacologia , Cálcio/metabolismo , Proteína gp120 do Envelope de HIV/fisiologia , HIV-1/análise , Neurônios/efeitos dos fármacos , Animais , Células Cultivadas , Proteína gp120 do Envelope de HIV/administração & dosagem , Proteína gp120 do Envelope de HIV/antagonistas & inibidores , Hipocampo/citologia , Neurônios/metabolismo , Nimodipina/farmacologia , Ratos , Proteínas Recombinantes/farmacologia , Células Ganglionares da Retina/efeitos dos fármacos , Células Ganglionares da Retina/metabolismo
6.
Science ; 224(4646): 303-6, 1984 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-6143400

RESUMO

Ganglion cells were dissociated from postnatal rat retinas, identified by specific fluorescent labels, and maintained in culture on a variety of substrates. Regeneration of processes by retinal ganglion cells was enhanced when the cells were plated on glass coated with a monoclonal antibody against the Thy-1 determinant. Plain glass and glass coated with polylysine, collagen, fibronectin, or other monoclonal antibodies supported the growth of neural processes, but were less effective than antibody to Thy-1.


Assuntos
Anticorpos Monoclonais/fisiologia , Antígenos de Superfície/imunologia , Isoanticorpos/fisiologia , Regeneração Nervosa , Retina/fisiologia , Células Ganglionares da Retina/fisiologia , Animais , Adesão Celular , Células Cultivadas , Polilisina/farmacologia , Ratos , Retina/citologia , Antígenos Thy-1
7.
Science ; 172(3988): 1128-32, 1971 Jun 11.
Artigo em Inglês | MEDLINE | ID: mdl-17839819

RESUMO

Salt used for deicing the streets near Rochester, New York, has increased the chloride concentration in Irondequoit Bay at least fivefold during the past two decades. During the winter of 1969-70 the quantity and salinity of the dense runoff that accumulated on the bottom of the bay was sufficient to prevent complete vertical mixing of the bay during the spring. Comparison with 1939 conditions indicates that the period of summer stratification has been prolonged a month by the density gradient imposed by the salt runoff.

8.
Neuron ; 5(3): 373-81, 1990 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-2169272

RESUMO

Afferent influences on natural cell death were modeled in retinal cultures derived from neonatal rats. Tetrodotoxin (TTX) blockade of electrical activity produced a significant reduction in surviving retinal ganglion cell (RGC) neurons during a critical period of development, similar in magnitude to the reduction observed during natural cell death in the intact retina at a similar developmental stage. The addition of vasoactive intestinal peptide (VIP) protected the RGCs from the lethal action of TTX. This effect was specific, since the related peptides PHI-27 and secretin produced no significant increase in RGC survival. Radioimmunoassay of cyclic nucleotides showed that TTX decreased culture levels of cAMP and that this trend was reversed by VIP. Decreases in RGC survival associated with TTX electrical blockade were prevented by 8-bromo:cAMP or forskolin. Furthermore, VIP10-28, the C-terminal fragment that inhibits VIP stimulation of adenylate cyclase, reduced the number of surviving RGCs. Thus, our results suggest that VIP, acting by increasing cAMP, has a neurotrophic effect on electrically blocked RGCs and may be an endogenous factor modulating normal cell death in the retina.


Assuntos
AMP Cíclico/metabolismo , Retina/efeitos dos fármacos , Células Ganglionares da Retina/efeitos dos fármacos , Tetrodotoxina/farmacologia , Peptídeo Intestinal Vasoativo/farmacologia , 8-Bromo Monofosfato de Adenosina Cíclica/farmacologia , Animais , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Colforsina/farmacologia , GMP Cíclico/análogos & derivados , GMP Cíclico/farmacologia , Secretina/farmacologia , Tetrodotoxina/antagonistas & inibidores
9.
Neuron ; 23(1): 171-80, 1999 May.
Artigo em Inglês | MEDLINE | ID: mdl-10402203

RESUMO

Zinc (Zn2+) inhibition of N-methyl-D-aspartate receptor (NMDAR) activity involves both voltage-independent and voltage-dependent components. Recombinant NR1/NR2A and NR1/NR2B receptors exhibit similar voltage-dependent block, but voltage-independent Zn2+ inhibition occurs with much higher affinity for NR1/NR2A than NR1/NR2B receptors (nanomolar versus micromolar IC50, respectively). Here, we show that two neighboring histidine residues on NR2A represent the critical determinant (termed the "short spacer") for high-affinity, voltage-independent Zn2+ inhibition using the Xenopus oocyte expression system and site-directed mutagenesis. Mutation of either one of these two histidine residues (H42 and H44) in the extracellular N-terminal domain of NR2A shifted the IC50 for high-affinity Zn2+ inhibition approximately 200-fold without affecting the EC50 of the coagonists NMDA and glycine. We suggest that the mechanism of high-affinity Zn2+ inhibition on the NMDAR involves enhancement of proton inhibition.


Assuntos
Histidina/fisiologia , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Zinco/farmacologia , Animais , Dietil Pirocarbonato/farmacologia , Feminino , Histidina/efeitos dos fármacos , Histidina/genética , Mutação/fisiologia , Oócitos , Técnicas de Patch-Clamp , Isoformas de Proteínas/antagonistas & inibidores , Receptores de N-Metil-D-Aspartato/metabolismo , Xenopus laevis , Zinco/antagonistas & inibidores
10.
Neuron ; 2(3): 1257-63, 1989 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-2696504

RESUMO

Electrophysiological responses to the glutamate analog N-methyl-D-aspartate (NMDA) measured in three different central neuronal preparations are subject to a novel modulatory mechanism: they are substantially potentiated after exposure to the disulfide reducing agent dithiothreitol, while oxidation with 5-5-dithiobis-2-nitrobenzoic acid decreases the magnitude of the response. Modification of the NMDA response by either oxidation or reduction does not appear to affect the pharmacological properties of the receptor-channel complex. Since we observe that the redox state of the native receptor-channel complex varies widely among neurons, an in vivo mechanism that can strongly regulate NMDA-activated functions by either reduction or oxidation may exist. In addition, these results suggest that it may be possible to design specific redox agents for characterizing the NMDA receptor-channel complex.


Assuntos
Ácido Aspártico/análogos & derivados , Córtex Cerebral/fisiologia , Neurônios/fisiologia , Retina/fisiologia , Células Ganglionares da Retina/fisiologia , Animais , Ácido Aspártico/farmacologia , Células Cultivadas , Galinhas , Ácido Ditionitrobenzoico/farmacologia , Ditiotreitol/farmacologia , Feto , Potenciais da Membrana/efeitos dos fármacos , N-Metilaspartato , Neurônios/efeitos dos fármacos , Oxirredução , Ratos , Células Ganglionares da Retina/efeitos dos fármacos , Zinco/farmacologia
11.
Neuron ; 7(1): 111-8, 1991 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-1676893

RESUMO

Exposure of rat retinal cultures to HIV-1 coat protein gp120 for several minutes increases [Ca2+]i in approximately half of the ganglion cells; this effect is associated with delayed-onset neuronal injury, similar to that previously reported in NMDA receptor-mediated neurotoxicity. Here we show that NMDA antagonists can prevent both the rise in [Ca2+]i and subsequent neuronal damage engendered by 20 pM gp120. However, whole-cell patch-clamp recordings demonstrate that gp120 does not directly evoke an NMDA-like response or enhance glutamate/NMDA-activated currents. Moreover, complete protection from gp120-induced [Ca2+]i increases and neurotoxicity is afforded by incubation with glutamate-pyruvate transaminase, which breaks down endogenous glutamate as verified by HPLC. Since, under standard conditions in these cultures, neither glutamate nor a low picomolar concentration of gp120 is deleterious on its own, our results suggest that their neurotoxicity is synergistic.


Assuntos
Proteína gp120 do Envelope de HIV/farmacologia , Neurotoxinas/farmacologia , Receptores de N-Metil-D-Aspartato/fisiologia , Células Ganglionares da Retina/efeitos dos fármacos , 2-Amino-5-fosfonovalerato/farmacologia , Alanina Transaminase/farmacocinética , Animais , Cálcio/metabolismo , Maleato de Dizocilpina/farmacologia , Glutamatos/metabolismo , Ácido Glutâmico , Proteína gp120 do Envelope de HIV/antagonistas & inibidores , N-Metilaspartato/antagonistas & inibidores , Concentração Osmolar , Células Ganglionares da Retina/metabolismo
12.
Neuron ; 13(4): 929-36, 1994 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-7524561

RESUMO

Modulation of NMDA-mediated responses by oxidizing and reducing reagents has been described in a variety of neuronal preparations. Here, we report that NMDA-gated currents of oocytes expressing heteromeric NMDA receptors are also modulated by sulfhydryl redox reagents. Each cysteine residue in the NMDAR1 (NR1) subunit and each conserved NMDAR2 (NR2) cysteine residue in a prototypical subunit (NR2B) was tested for its role in redox modulation. We have identified 2 cysteines in the NR1 subunit that are required for redox modulation of NMDA-gated currents in oocytes expressing NR1-NR2B, NR1-NR2C, or NR1-NR2D receptors. Mutation of these same 2 cysteines also eliminated potentiation by spermine and shifted the IC50 for H+ inhibition and the EC50 for NMDA. Redox modulation of heteromeric NR1-NR2A receptors appeared to be different from that of the other heteromeric receptors, indicating the presence of one or more unique redox modulatory sites on NR1-NR2A receptors.


Assuntos
Cisteína/química , Receptores de N-Metil-D-Aspartato/química , Animais , Cisteína/genética , Ditiotreitol/farmacologia , Eletrofisiologia , Feminino , Ativação do Canal Iônico/efeitos dos fármacos , Canais Iônicos/fisiologia , Dados de Sequência Molecular , Mutagênese Sítio-Dirigida , N-Metilaspartato/farmacologia , Oócitos/metabolismo , Oxirredução , Técnicas de Patch-Clamp , Ratos , Receptores de N-Metil-D-Aspartato/genética , Receptores de N-Metil-D-Aspartato/fisiologia , Proteínas Recombinantes , Espermina/farmacologia , Relação Estrutura-Atividade , Xenopus
13.
Neuron ; 8(6): 1087-99, 1992 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-1376999

RESUMO

Nitric oxide (NO) is an important messenger both systemically and in the CNS. In digital Ca2+ imaging and patch-clamp experiments, clinically available nitroso compounds that generate NO are shown to inhibit responses mediated by the NMDA subtype of the glutamate receptor on rat cortical neurons in vitro. A mechanism of action for this effect was investigated by using the specific NO-generating agent S-nitrosocysteine. We propose that free sulfhydryl groups on the NMDA receptor-channel complex react to form one or more S-nitrosothiols in the presence of NO. If vicinal thiol groups react in this manner, they can form a disulfide bond(s), which is thought to constitute the redox modulatory site of the receptor, resulting in a relatively persistent blockade of NMDA responses. These reactions with NO can afford protection from NMDA receptor-mediated neurotoxicity. Our results demonstrate a new pathway for NO regulation of physiological function that is not via cGMP, but instead involves reactions with membrane-bound thiol groups on the NMDA receptor-channel complex.


Assuntos
Canais Iônicos/metabolismo , Óxido Nítrico/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Alquilantes/farmacologia , Animais , Cálcio/metabolismo , Eletrofisiologia , Membranas Intracelulares/metabolismo , Ácido Caínico/farmacologia , N-Metilaspartato/farmacologia , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Compostos Nitrosos/farmacologia , Oxirredução , Potássio/farmacologia
14.
Neuron ; 15(4): 961-73, 1995 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-7576644

RESUMO

During ischemic brain injury, glutamate accumulation leads to overstimulation of postsynaptic glutamate receptors with intracellular Ca2+ overload and neuronal cell death. Here we show that glutamate can induce either early necrosis or delayed apoptosis in cultures of cerebellar granule cells. During and shortly after exposure to glutamate, a subpopulation of neurons died by necrosis. In these cells, mitochondrial membrane potential collapsed, nuclei swelled, and intracellular debris were scattered in the incubation medium. Neurons surviving the early necrotic phase recovered mitochondrial potential and energy levels. Later, they underwent apoptosis, as shown by the formation of apoptotic nuclei and by chromatin degradation into high and low molecular weight fragments. These results suggest that mitochondrial function is a critical factor that determines the mode of neuronal death in excitotoxicity.


Assuntos
Apoptose/efeitos dos fármacos , Cerebelo/patologia , Ácido Glutâmico/farmacologia , Mitocôndrias/fisiologia , Neurônios/efeitos dos fármacos , Animais , Cromatina/efeitos dos fármacos , Cromatina/ultraestrutura , DNA/metabolismo , Potenciais da Membrana/efeitos dos fármacos , Peso Molecular , Necrose/induzido quimicamente , Neurônios/fisiologia , Neurônios/ultraestrutura , Ratos , Ratos Sprague-Dawley
15.
Neuron ; 24(2): 461-9, 1999 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-10571239

RESUMO

Recent evidence indicates that the NO-related species, nitroxyl anion (NO), is produced in physiological systems by several redox metal-containing proteins, including hemoglobin, nitric oxide synthase (NOS), superoxide dismutase, and S-nitrosothiols (SNOs), which have recently been identified in brain. However, the chemical biology of NO- remains largely unknown. Here, we show that NO- -unlike NO*, but reminiscent of NO+ transfer (or S-nitrosylation)- -reacts mainly with Cys-399 in the NR2A subunit of the N-methyl-D-aspartate (NMDA) receptor to curtail excessive Ca2+ influx and thus provide neuroprotection from excitotoxic insults. This effect of NO- closely resembles that of NOS, which also downregulates NMDA receptor activity under similar conditions in culture.


Assuntos
Neurotoxinas/metabolismo , Óxidos de Nitrogênio/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Animais , Ânions/metabolismo , Células Cultivadas , Córtex Cerebral/citologia , Feminino , N-Metilaspartato/farmacologia , Neurônios/metabolismo , Óxido Nítrico/farmacologia , Doadores de Óxido Nítrico/farmacologia , Óxido Nítrico Sintase/metabolismo , Oócitos , Ratos , Ratos Sprague-Dawley , Receptores de N-Metil-D-Aspartato/química , Receptores de N-Metil-D-Aspartato/efeitos dos fármacos , Receptores de N-Metil-D-Aspartato/fisiologia , Compostos de Sulfidrila/química , Compostos de Sulfidrila/metabolismo , Xenopus laevis
16.
Cell Death Differ ; 14(7): 1305-14, 2007 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-17431424

RESUMO

Although activation of glutamate receptors is essential for normal brain function, excessive activity leads to a form of neurotoxicity known as excitotoxicity. Key mediators of excitotoxic damage include overactivation of N-methyl-D-aspartate (NMDA) receptors, resulting in excessive Ca(2+) influx with production of free radicals and other injurious pathways. Overproduction of free radical nitric oxide (NO) contributes to acute and chronic neurodegenerative disorders. NO can react with cysteine thiol groups to form S-nitrosothiols and thus change protein function. S-nitrosylation can result in neuroprotective or neurodestructive consequences depending on the protein involved. Many neurodegenerative diseases manifest conformational changes in proteins that result in misfolding and aggregation. Our recent studies have linked nitrosative stress to protein misfolding and neuronal cell death. Molecular chaperones - such as protein-disulfide isomerase, glucose-regulated protein 78, and heat-shock proteins - can provide neuroprotection by facilitating proper protein folding. Here, we review the effect of S-nitrosylation on protein function under excitotoxic conditions, and present evidence that NO contributes to degenerative conditions by S-nitrosylating-specific chaperones that would otherwise prevent accumulation of misfolded proteins and neuronal cell death. In contrast, we also review therapeutics that can abrogate excitotoxic damage by preventing excessive NMDA receptor activity, in part via S-nitrosylation of this receptor to curtail excessive activity.


Assuntos
Encéfalo/metabolismo , Citoproteção/fisiologia , Doenças Neurodegenerativas/tratamento farmacológico , Doenças Neurodegenerativas/metabolismo , Óxido Nítrico/metabolismo , Dobramento de Proteína , Animais , Encéfalo/efeitos dos fármacos , Encéfalo/fisiopatologia , Citoproteção/efeitos dos fármacos , Antagonistas de Aminoácidos Excitatórios/farmacologia , Antagonistas de Aminoácidos Excitatórios/uso terapêutico , Humanos , Chaperonas Moleculares/metabolismo , Doenças Neurodegenerativas/fisiopatologia , Compostos de Nitrogênio/metabolismo , Receptores de N-Metil-D-Aspartato/antagonistas & inibidores , Receptores de N-Metil-D-Aspartato/metabolismo
17.
Cell Death Differ ; 14(2): 296-305, 2007 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-16841089

RESUMO

The chemokine receptors CCR5 and CXCR4 serve, in addition to CD4, as coreceptors for human immunodeficiency virus-1 (HIV-1), and infection with HIV-1 can cause dementia. In brain-derived cells, HIV-1 envelope glycoprotein gp120 initiates a signaling cascade that involves p38 mitogen-activated protein kinase and leads to neuronal cell death. Using mixed neuronal/glial cultures from rats and mice genetically deficient in one or both HIV coreceptors, we show here that CCR5, CXCR4 or both can mediate HIV/gp120 neurotoxicity depending on the viral strain. Paradoxically, we also found evidence for a CCR5-mediated neuroprotective pathway. We identify protein kinase Akt/PKB as an essential component of this pathway, which can be triggered by the CCR5 agonists macrophage inflammatory protein-1beta and regulated-and-normal-T-cell-expressed-and-secreted. Moreover, these CCR5 ligands prevent neuronal cell death induced by stromal cell-derived factor-1, a CXCR4 agonist. Both neurons and glia coexpress CXCR4 and CCR5. Ca2+ imaging experiments demonstrate that engagement of CCR5 prevents CXCR4-triggered increases in intracellular free Ca2+. This finding suggests that CCR5 ligands can protect neurons at least, in part, by modulating CXCR4-mediated toxicity through heterologous desensitization.


Assuntos
HIV-1/imunologia , Neurônios/citologia , Neurônios/patologia , Receptores CCR5/metabolismo , Receptores CXCR4/metabolismo , Animais , Morte Celular/efeitos dos fármacos , Quimiocina CXCL12 , Citocinas/toxicidade , Proteína gp120 do Envelope de HIV/toxicidade , Infecções por HIV/virologia , Imidazóis/farmacologia , Ligantes , Camundongos , Camundongos Endogâmicos C57BL , Modelos Imunológicos , N-Metilaspartato/toxicidade , Neuroglia/citologia , Neuroglia/efeitos dos fármacos , Neuroglia/patologia , Neurônios/virologia , Neurotoxinas/toxicidade , Piridinas/farmacologia , Ratos , Receptores CCR5/deficiência , Receptores CXCR4/deficiência , Proteínas Quinases p38 Ativadas por Mitógeno/antagonistas & inibidores
18.
Cell Death Differ ; 14(3): 462-71, 2007 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-17053808

RESUMO

Mitochondrial dysfunction is an underpinning event in many neurodegenerative disorders. Less clear, however, is how mitochondria become injured during neuronal demise. Nitric oxide (NO) evokes rapid mitochondrial fission in cortical neurons. Interestingly, proapoptotic Bax relocates from the cytoplasm into large foci on mitochondrial scission sites in response to nitrosative stress. Antiapoptotic Bcl-xL does not prevent mitochondrial fission despite its ability to block Bax puncta formation on mitochondria and to mitigate neuronal cell death. Mitofusin 1 (Mfn1) or dominant-negative dynamin-related protein 1(K38A) (Drp1(k38A)) inhibits mitochondrial fission and Bax accumulation on mitochondria induced by exposure to an NO donor. Although NO is known to cause a bioenergetic crisis, lowering ATP by glycolytic or mitochondrial inhibitors neither induces mitochondrial fission nor Bax foci formation on mitochondria. Taken together, these data indicate that the mitochondrial fission machinery acts upstream of the Bcl-2 family of proteins in neurons challenged with nitrosative stress.


Assuntos
Mitocôndrias/metabolismo , Neurônios/metabolismo , Óxido Nítrico/farmacologia , Proteína X Associada a bcl-2/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Morte Celular , Células Cultivadas , Córtex Cerebral/citologia , Córtex Cerebral/efeitos dos fármacos , Córtex Cerebral/metabolismo , Córtex Cerebral/fisiologia , Glicólise , Proteínas de Membrana/metabolismo , Proteínas de Membrana/fisiologia , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/fisiologia , Mitocôndrias/ultraestrutura , Proteínas Mitocondriais/metabolismo , Proteínas Mitocondriais/fisiologia , Modelos Biológicos , Neurônios/efeitos dos fármacos , Neurônios/fisiologia , Transporte Proteico , Ratos , Ratos Sprague-Dawley , Transfecção , Proteína bcl-X/metabolismo , Proteína bcl-X/fisiologia
19.
Nat Neurosci ; 3(1): 15-21, 2000 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-10607390

RESUMO

Several ion channels are thought to be directly modulated by nitric oxide (NO), but the molecular basis of this regulation is unclear. Here we show that the NMDA receptor (NMDAR)-associated ion channel was modulated not only by exogenous NO but also by endogenous NO. Site-directed mutagenesis identified a critical cysteine residue (Cys 399) on the NR2A subunit whose S-nitrosylation (NO+ transfer) under physiological conditions underlies this modulation. In cell systems expressing NMDARs with mutant NR2A subunits in which this single cysteine was replaced by an alanine, the effect of endogenous NO was lost. Thus endogenous S-nitrosylation can regulate ion channel activity.


Assuntos
Transporte de Íons/fisiologia , Óxido Nítrico/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , S-Nitrosotióis , Animais , Linhagem Celular , Cromatografia Líquida de Alta Pressão , Cisteína/análogos & derivados , Cisteína/metabolismo , Cisteína/farmacologia , Relação Dose-Resposta a Droga , Metanossulfonato de Etila/análogos & derivados , Metanossulfonato de Etila/farmacologia , Humanos , Indicadores e Reagentes/farmacologia , Transporte de Íons/efeitos dos fármacos , Mesilatos/farmacologia , Mutagênese Sítio-Dirigida , N-Metilaspartato/antagonistas & inibidores , N-Metilaspartato/farmacologia , Óxido Nítrico/farmacologia , Compostos Nitrosos/farmacologia , Oócitos/citologia , Técnicas de Patch-Clamp , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Receptores de N-Metil-D-Aspartato/genética , Transfecção , Xenopus laevis
20.
Mol Cell Biol ; 13(4): 2564-77, 1993 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-8455629

RESUMO

The myocyte enhancer-binding factor 2 (MEF2) site is an essential element of many muscle-specific enhancers and promoters that binds nuclear proteins from muscle and brain. Recently, we have cloned a family of MEF2 transcription factors produced by two genes that, at the mRNA level, are broadly expressed and produce tissue-specific isoforms by posttranscriptional processes (Y.-T. Yu, R. E. Breitbart, L. B. Smoot, Y. Lee, V. Mahdavi, and B. Nadal-Ginard, Genes Dev. 6:1783-1798, 1992). Here, we report the isolation and functional characterization of cDNA clones encoding four MEF2 factors derived from a separate gene that we have named hMEF2C. In contrast to those of the previously reported genes, the transcripts of the hMEF2C gene are restricted to skeletal muscle and brain. One of the alternate exons is exclusively present in brain transcripts. The products of this gene have DNA-binding and trans-activating activities indistinguishable from those of the previously reported MEF2 factors. The hMEF2C gene is induced late during myogenic differentiation, and its expression is limited to a subset of cortical neurons. The potential targets for this transcription factor in a subset of neurons are not known at this time. The strict tissue-specific pattern of expression of hMEF2C in comparison with the more ubiquitous expression of other MEF2 genes suggests a different mode of regulation and a potentially important role of hMEF2C factors in myogenesis and neurogenesis.


Assuntos
Encéfalo/fisiologia , Proteínas de Ligação a DNA/genética , Genes , Músculos/fisiologia , Fatores de Transcrição/genética , Processamento Alternativo , Sequência de Aminoácidos , Animais , Sequência de Bases , Células Cultivadas , Clonagem Molecular , Sequência Consenso , Expressão Gênica , Humanos , Técnicas Imunológicas , Técnicas In Vitro , Fatores de Transcrição MEF2 , Camundongos , Dados de Sequência Molecular , Fatores de Regulação Miogênica , Neurônios/fisiologia , Oligodesoxirribonucleotídeos/química , Reação em Cadeia da Polimerase , RNA Mensageiro/genética , Alinhamento de Sequência , Distribuição Tecidual , Transcrição Gênica
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