RESUMEN
In the last several years a large number of studies have demonstrated the neurobiological and clinical continuum between depression and Alzheimer's disease (AD). Depression is a risk factor for the development of AD, and the presence of depressive symptoms significantly increases the conversion of Mild Cognitive Impairment (MCI) into AD. Common pathophysiological events have been identified in depression and AD, including neuroinflammation with an aberrant Tumor Necrosis Factor-α (TNF-α) signaling, and an impairment of Brain-Derived Neurotrophic Factor (BDNF) and Transforming-Growth-Factor-ß1 (TGF-ß1) signaling. TGF-ß1 is an anti-inflammatory cytokine that exerts neuroprotective effects against amyloid-ß (Aß)-induced neurodegeneration, and it has a key role in memory formation and synaptic plasticity. TGF-ß1 plasma levels are reduced in major depressed patients (MDD), correlate with depression severity, and significantly contribute to treatment resistance in MDD. The deficit of Smad-dependent TGF-ß1 signaling is also an early event in AD pathogenesis, which contributes to inflammaging and cognitive decline in AD. A long-term treatment with antidepressants such as selective-serotonin-reuptake inhibitors (SSRIs) is known to reduce the risk of AD in patients with depression and, SSRIs, such as fluoxetine, increase the release of TGF-ß1 from astrocytes and exert relevant neuroprotective effects in experimental models of AD. We propose the TGF-ß1 signaling pathway as a common pharmacological target in depression and AD, and discuss the potential rescue of TGF-ß1 signaling by antidepressants as a way to prevent the transition from depression to AD.
Asunto(s)
Enfermedad de Alzheimer/tratamiento farmacológico , Depresión/tratamiento farmacológico , Factor de Crecimiento Transformador beta1/metabolismo , Enfermedad de Alzheimer/epidemiología , Enfermedad de Alzheimer/metabolismo , Animales , Disfunción Cognitiva/metabolismo , Depresión/epidemiología , Depresión/metabolismo , Humanos , Transducción de SeñalRESUMEN
We synthesized a new ratiometric fluorescent Cu2+ probe, bearing a morpholine moiety for selective binding to lysosomes and two picolylamine arms for the specific chelation of divalent copper ions. The probe capability to detect lysosomal Cu2+ was demonstrated in human differentiated neuroblastoma cells by confocal microscopy.
Asunto(s)
Quelantes/química , Cobre/química , Colorantes Fluorescentes/química , Lisosomas/química , Neuroblastoma/patología , Imagen Óptica , Diferenciación Celular , Quelantes/síntesis química , Colorantes Fluorescentes/síntesis química , Humanos , Microscopía Confocal , Morfolinas/química , Picolinas/químicaRESUMEN
The inhibition of amyloid formation is a promising therapeutic approach for the treatment of neurodegenerative diseases. Peptide-based inhibitors, which have been widely investigated, are generally derived from original amyloid sequences. Most interestingly, trehalose, a nonreducing disaccharide of α-glucose, is effective in preventing the aggregation of numerous proteins. We have determined that the development of hybrid compounds could provide new molecules with improved properties that might synergically increase the potency of their single moieties. In this work, the ability of Ac-LPFFD-Th, a C-terminally trehalose-conjugated derivative, to slow down the Aß aggregation process was investigated by means of different biophysical techniques, including thioflavinâ T fluorescence, dynamic light scattering, ESI-MS, and NMR spectroscopy. Moreover, we demonstrate that Ac-LPFFD-Th modifies the aggregation features of Aß and protects neurons from Aßâ oligomers' toxic insult.
Asunto(s)
Péptidos beta-Amiloides/antagonistas & inhibidores , Peptidomiméticos/farmacología , Trehalosa/farmacología , Péptidos beta-Amiloides/química , Animales , Supervivencia Celular/efectos de los fármacos , Células Cultivadas , Estructura Molecular , Neuronas/citología , Neuronas/efectos de los fármacos , Peptidomiméticos/química , Ratas , Trehalosa/químicaRESUMEN
The ectopic re-activation of cell cycle in neurons is an early event in the pathogenesis of Alzheimer's disease (AD), which could lead to synaptic failure and ensuing cognitive deficits before frank neuronal death. Cytostatic drugs that act as cyclin-dependent kinase (CDK) inhibitors have been poorly investigated in animal models of AD. In the present study, we examined the effects of flavopiridol, an inhibitor of CDKs currently used as antineoplastic drug, against cell cycle reactivation and memory loss induced by intracerebroventricular injection of Aß1-42 oligomers in CD1 mice. Cycling neurons, scored as NeuN-positive cells expressing cyclin A, were found both in the frontal cortex and in the hippocampus of Aß-injected mice, paralleling memory deficits. Starting from three days after Aß injection, flavopiridol (0.5, 1 and 3mg/kg) was intraperitoneally injected daily, for eleven days. Here we show that a treatment with flavopiridol (0.5 and 1mg/kg) was able to rescue the loss of memory induced by Aß1-42, and to prevent the occurrence of ectopic cell-cycle events in the mouse frontal cortex and hippocampus. This is the first evidence that a cytostatic drug can prevent cognitive deficits in a non-transgenic animal model of AD.
Asunto(s)
Péptidos beta-Amiloides/efectos adversos , Antineoplásicos/farmacología , Flavonoides/farmacología , Trastornos de la Memoria/inducido químicamente , Trastornos de la Memoria/tratamiento farmacológico , Memoria/efectos de los fármacos , Fragmentos de Péptidos/efectos adversos , Piperidinas/farmacología , Enfermedad de Alzheimer/inducido químicamente , Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/metabolismo , Animales , Trastornos del Conocimiento/inducido químicamente , Trastornos del Conocimiento/tratamiento farmacológico , Trastornos del Conocimiento/metabolismo , Quinasas Ciclina-Dependientes/metabolismo , Modelos Animales de Enfermedad , Lóbulo Frontal/efectos de los fármacos , Lóbulo Frontal/metabolismo , Hipocampo/efectos de los fármacos , Hipocampo/metabolismo , Masculino , Trastornos de la Memoria/etiología , Trastornos de la Memoria/metabolismo , Ratones , Neuronas/efectos de los fármacos , Neuronas/metabolismoRESUMEN
Cell-cycle reactivation is a core feature of degenerating neurons in Alzheimer's disease (AD) and Parkinson's disease (PD). A variety of stressors, including ß-amyloid (Aß) in the case of AD, can force neurons to leave quiescence and to initiate an ectopic DNA replication process, leading to neuronal death rather than division. As the primary polymerase (pol) involved in neuronal DNA replication, DNA pol-ß contributes to neuronal death, and DNA pol-ß inhibitors may prove to be effective neuroprotective agents. Currently, specific and highly active DNA pol-ß inhibitors are lacking. Nine putative DNA pol-ß inhibitors were identified in silico by querying the ZINC database, containing more than 35 million purchasable compounds. Following pharmacological evaluation, only 5-methoxyflavone (1) was validated as an inhibitor of DNA pol-ß activity. Cultured primary neurons are a useful model to investigate the neuroprotective effects of potential DNA pol-ß inhibitors, since these neurons undergo DNA replication and death when treated with Aß. Consistent with the inhibition of DNA pol-ß, 5-methoxyflavone (1) reduced the number of S-phase neurons and the ensuing apoptotic death triggered by Aß. 5-Methoxyflavone (1) is the first flavonoid compound able to halt neurodegeneration via a definite molecular mechanism rather than through general antioxidant and anti-inflammatory properties.
Asunto(s)
ADN Polimerasa beta/antagonistas & inhibidores , Flavonas/farmacología , Neuronas/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Enfermedad de Alzheimer/patología , Ciclo Celular/efectos de los fármacos , Muerte Celular/efectos de los fármacos , División Celular/efectos de los fármacos , Replicación del ADN/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Flavonas/química , Humanos , Estructura Molecular , Neuronas/metabolismo , Fármacos Neuroprotectores/química , Enfermedad de Parkinson/patologíaRESUMEN
Several studies have highlighted the presence of nitration damage following neuroinflammation in Alzheimer's disease (AD). Accordingly, post-transcriptional modifications of ß-amyloid (Aß), including peptide nitration, have been explored as a marker of the disease. However, the implications of Aß nitration in terms of aggregation propensity and neurotoxicity are still debated. Here, we show new data obtained using a photoactivatable peroxynitrite generator (BPT-NO) to overcome the limitations associated with chemical nitration methods. We found that the photoactivation of BPT-NO with the highly biocompatible red light selectively induces the nitration of tyrosine 10 of freshly solubilized full-length Aß1-42. Photonitrated Aß1-42 was, therefore, investigated for aggregation states and functions. It resulted that photonitrated Aß1-42 did not aggregate into small oligomers but rather self-assembled into large amorphous aggregates. When tested on neuronal-like SH-SY5Y cells and microglial C57BL/6 BV2 cells, photonitrated Aß1-42 showed to be free of neurotoxicity and able to induce phagocytic microglia cells. We propose that light-controlled nitration of the multiple forms in which Aß occurs (i.e., monomers, oligomers, fibrils) could be a tool to assess in real-time the impact of tyrosine nitration on the amyloidogenic and toxic properties of Aß1-42.
Asunto(s)
Péptidos beta-Amiloides , Luz , Fragmentos de Péptidos , Tirosina , Péptidos beta-Amiloides/metabolismo , Tirosina/metabolismo , Tirosina/análogos & derivados , Tirosina/química , Fragmentos de Péptidos/metabolismo , Fragmentos de Péptidos/toxicidad , Humanos , Animales , Microglía/metabolismo , Microglía/efectos de los fármacos , Ácido Peroxinitroso/metabolismo , Ratones , Agregado de Proteínas/fisiología , Enfermedad de Alzheimer/metabolismo , Enfermedad de Alzheimer/patología , Neuronas/metabolismo , Neuronas/efectos de los fármacosRESUMEN
Fingolimod (FTY720), a novel drug approved for the treatment of relapsing-remitting multiple sclerosis, activates different sphingosine-1-phosphate receptor (S1PR) subtypes. Its primary mechanism of action is to reduce the egress of T lymphocytes from secondary lymphoid organs, thus restraining neuroinflammation and autoimmunity. However, recent evidence suggests that the action of FTY720 involves S1PRs expressed by cells resident in the CNS, including neurons. Here, we examined the effect of FTY720, its active metabolite, FTY720-P, and sphingosine-1-phosphate (S1P) on neuronal viability using a classical in vitro model of excitotoxic neuronal death. Mixed cultures of mouse cortical cells were challenged with toxic concentrations of N-methyl-d-aspartate (NMDA) for 10 min, and neuronal death was assessed 20 h later. FTY720, FTY720-P, and S1P were all neuroprotective when applied 18-20 h prior to the NMDA pulse. Neuroprotection was attenuated by pertussis toxin, and inhibited by the selective type-1 S1PR (S1P1R) antagonist, W146, and by inhibitors of the mitogen associated protein kinase (MAPK) and the phosphatidylinositol-3-kinase (PtdIns-3-K) pathways. Both FTY720 and FTY720-P retained their protective activity in pure cultures of mouse or rat cortical neurons. These data offer the first direct demonstration that FTY720 and its active metabolite protect neurons against excitotoxic death.
Asunto(s)
Inmunosupresores/farmacología , Neuronas/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Organofosfatos/farmacología , Glicoles de Propileno/farmacología , Esfingosina/análogos & derivados , Animales , Astrocitos/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Células Cultivadas , Femenino , Clorhidrato de Fingolimod , Lisofosfolípidos/farmacología , Ratones , Ratas , Esfingosina/farmacologíaRESUMEN
BACKGROUND: Ectopic cell cycle reactivation in neurons is associated with neuronal death in Alzheimer's disease. In cultured rodent neurons, synthetic ß-amyloid (Aß) reproduces the neuronal cell cycle re-entry observed in the Alzheimer's brain, and blockade of the cycle prevents Aß-induced neurodegeneration. DNA polymerase-ß, whose expression is induced by Aß, is responsible for the DNA replication process that ultimately leads to neuronal death, but the molecular mechanism(s) linking DNA replication to neuronal apoptosis are presently unknown. AIM: To explore the role of a conserved checkpoint pathway started by DNA replication stress, namely the ATM-ATR/Claspin/Chk-1 pathway, in switching the neuronal response from DNA replication to apoptosis. METHODS: Experiments were carried out in cultured rat cortical neurons challenged with toxic oligomers of Aß protein. RESULTS: Small inhibitory molecules of ATM/ATR kinase or Chk-1 amplified Aß-induced neuronal DNA replication and apoptosis, as they were permissive to the DNA polymerase-ß activity triggered by Aß oligomers. Claspin, i.e., the adaptor protein between ATM/ATR kinase and the downstream Chk-1, was present on DNA replication forks of neurons early after Aß challenge, and decreased at times coinciding with neuronal apoptosis. The caspase-3/7 inhibitor I maintained overtime the amount of Claspin loaded on DNA replication forks and, concomitantly, reduced neuronal apoptosis by holding neurons in the S phase. Moreover, a short phosphopeptide mimicking the Chk-1-binding motif of Claspin was able to prevent Aß-challenged neurons from entering apoptosis. CONCLUSION: We speculate that, in the Alzheimer's brain, Claspin degradation by intervening factors may precipitate the death of neurons engaged into DNA replication.
Asunto(s)
Enfermedad de Alzheimer , Péptidos beta-Amiloides , Ratas , Animales , Péptidos beta-Amiloides/toxicidad , Replicación del ADN , Muerte Celular , Apoptosis/fisiología , Neuronas/fisiología , ADN Polimerasa Dirigida por ADNRESUMEN
BACKGROUND: The effects of Alzheimer's disease (AD) pathology on the experience of pain are poorly understood. OBJECTIVE: To understand the pathophysiological mechanisms underlying pain sensory transmission in the transgenic mouse model of AD, CRND8. METHODS: We explored AD-related pathology in the spinal cord and dorsal root ganglia of 18-week-old female CRND8 mice. We assessed nociceptive responses to both acute heat stimuli and persistent inflammatory pain in CRND8 mice and non-transgenic (non-Tg) littermates. In addition, we searched for differences in biochemical correlates of inflammatory pain between CRND8 and non-Tg mice. Finally, we investigated the excitability of dorsal horn noc iceptive neurons in spinal cord slices from CRND8 and non-Tg mice. RESULTS: We demonstrated the presence of intracellular AD-like pathology in the spinal cord and in the dorsal root ganglia nociceptive sensory neurons of CRND8 mice. We found that CRND8 mice had a reduced susceptibility to acute noxious heat stimuli and an increased sensitivity to tonic inflammatory pain. Tonic inflammatory pain correlated with a lack of induction of pro-opiomelanocortin in the spinal cord of CRND8 mice as compared to non-Tg mice. Electrophysiological recording in acute spinal cord slice preparations indicated an increased probability of glutamate release at the membrane of dorsal horn nociceptive neurons in CRND8 mice. CONCLUSION: This study suggests that an increased thermal tolerance and a facilitation of nociception by peripheral inflammation can coexist in AD.
Asunto(s)
Enfermedad de Alzheimer , Hiperalgesia , Ratones , Femenino , Animales , Enfermedad de Alzheimer/complicaciones , Calor , Dolor/etiología , Ratones TransgénicosRESUMEN
ß-Amyloid protein (Aß) is thought to be responsible for neuronal apoptosis in Alzheimer's disease (AD). Paradoxically, Aß can also promote neurogenesis, both in vitro and in vivo, by inducing neural progenitor cells (NPCs) to differentiate into neurons. However, the mechanisms of Aß-induced neurogenesis are unknown. Here we examined the role of DNA polymerase-ß (DNA pol-ß), a DNA repair enzyme that is required for proper neurogenesis during brain development and is also responsible for Aß-induced neuronal apoptosis. In neurospheres obtained from the adult mouse subventricular zone (SVZ), the knockdown of DNA pol-ß or its pharmacological blockade showed that the enzyme functioned both to repress proliferation of early nestin(+) progenitor cells and to promote the maturation of TuJ-1(+) neuronal cells. In neurospheres challenged with oligomers of synthetic Aß(42) , the expression levels of DNA pol-ß were rapidly increased. DNA pol-ß knockdown prevented the Aß(42) -promoted differentiation of nestin(+) progenitor cells into nestin(+) /Dlx-2(+) neuroblasts. Moreover, when neurospheres were seeded to allow full differentiation of their elements, blockade of DNA pol-ß prevented Aß(42) -induced differentiation of progenitors into MAP-2(+) neurons. Thus, our data demonstrate that Aß(42) arrests the proliferation of a subpopulation of nestin(+) cells via the induction of DNA pol-ß, thereby allowing for their differentiation toward the neuronal lineage. Our findings reveal a novel role of DNA pol-ß in Aß(42) -induced neurogenesis and identify DNA pol-ß as a key mechanistic link between the neurogenic effect of Aß(42) on NPCs and the proapoptotic effect of Aß(42) on mature neurons.
Asunto(s)
Péptidos beta-Amiloides/metabolismo , Ventrículos Cerebrales/metabolismo , ADN Polimerasa beta/metabolismo , Neurogénesis/fisiología , Neuronas/metabolismo , Péptidos beta-Amiloides/farmacología , Animales , Puntos de Control del Ciclo Celular/efectos de los fármacos , Puntos de Control del Ciclo Celular/fisiología , Proliferación Celular/efectos de los fármacos , Ventrículos Cerebrales/citología , ADN Polimerasa beta/farmacología , Ratones , Neurogénesis/efectos de los fármacos , Neuronas/citología , Neuronas/efectos de los fármacosRESUMEN
Alzheimer's disease (AD) is a neurodegenerative disorder that affects about 35 million people worldwide. Current drugs for AD only treat the symptoms and do not interfere with the underlying pathogenic mechanisms of the disease. AD is characterized by the presence of ß-amyloid (Aß) plaques, neurofibrillary tangles, and neuronal loss. Identification of the molecular determinants underlying Aß-induced neurodegeneration is an essential step for the development of disease-modifying drugs. Recently, an impairment of the transforming growth factor-ß1 (TGF-ß1) signaling pathway has been demonstrated to be specific to the AD brain and, particularly, to the early phase of the disease. TGF-ß1 is a neurotrophic factor responsible for the initiation and maintenance of neuronal differentiation and synaptic plasticity. The deficiency of TGF-ß1 signaling is associated with Aß pathology and neurofibrillary tangle formation in AD animal models. Reduced TGF-ß1 signaling seems to contribute both to microglial activation and to ectopic cell-cycle re-activation in neurons, two events that contribute to neurodegeneration in the AD brain. The neuroprotective features of TGF-ß1 indicate the advantage of rescuing TGF-ß1 signaling as a means to slow down the neurodegenerative process in AD.
Asunto(s)
Enfermedad de Alzheimer/tratamiento farmacológico , Enfermedad de Alzheimer/patología , Enfermedad de Alzheimer/fisiopatología , Fármacos Neuroprotectores/uso terapéutico , Transducción de Señal/fisiología , Factor de Crecimiento Transformador beta1/metabolismo , Animales , Humanos , Fármacos Neuroprotectores/metabolismo , Proteínas Smad/metabolismoRESUMEN
Dual orthosteric agonists of metabotropic glutamate 2 (mGlu2) and mGlu3 receptors are being developed as novel antipsychotic agents devoid of the adverse effects of conventional antipsychotics. Therefore, these drugs could be helpful for the treatment of psychotic symptoms associated with Alzheimer's disease (AD). In experimental animals, the antipsychotic activity of mGlu2/3 receptor agonists is largely mediated by the activation of mGlu2 receptors and is mimicked by selective positive allosteric modulators (PAMs) of mGlu2 receptors. We investigated the distinct influence of mGlu2 and mGlu3 receptors in mixed and pure neuronal cultures exposed to synthetic ß-amyloid protein (Aß) to model neurodegeneration occurring in AD. The mGlu2 receptor PAM, N-4'-cyano-biphenyl-3-yl)-N-(3-pyridinylmethyl)-ethanesulfonamide hydrochloride (LY566332), devoid of toxicity per se, amplified Aß-induced neurodegeneration, and this effect was prevented by the mGlu2/3 receptor antagonist (2S,1'S,2'S)-2-(9-xanthylmethyl)-2-(2'-carboxycyclopropyl)glycine (LY341495). LY566332 potentiated Aß toxicity regardless of the presence of glial mGlu3 receptors, but it was inactive when neurons lacked mGlu2 receptors. The dual mGlu2/3 receptor agonist, (-)-2-oxa-4-aminobicyclo[3.1.0]exhane-4,6-dicarboxylic acid (LY379268), was neuroprotective in mixed cultures via a paracrine mechanism mediated by transforming growth factor-ß1. LY379268 lost its protective activity in neurons grown with astrocytes lacking mGlu3 receptors, indicating that protection against Aß neurotoxicity was mediated entirely by glial mGlu3 receptors. The selective noncompetitive mGlu3 receptor antagonist, (3S)-1-(5-bromopyrimidin-2-yl)-N-(2,4-dichlorobenzyl)pyrrolidin-3-amine methanesulfonate hydrate (LY2389575), amplified Aß toxicity on its own, and, interestingly, unmasked a neurotoxic activity of LY379268, which probably was mediated by the activation of mGlu2 receptors. These data indicate that selective potentiation of mGlu2 receptors enhances neuronal vulnerability to Aß, whereas dual activation of mGlu2 and mGlu3 receptors is protective against Aß-induced toxicity.
Asunto(s)
Enfermedad de Alzheimer/tratamiento farmacológico , Péptidos beta-Amiloides/efectos de los fármacos , Antipsicóticos/farmacología , Fármacos Neuroprotectores/farmacología , Trastornos Psicóticos/tratamiento farmacológico , Receptores de Glutamato Metabotrópico/efectos de los fármacos , Enfermedad de Alzheimer/complicaciones , Enfermedad de Alzheimer/metabolismo , Aminoácidos/farmacología , Péptidos beta-Amiloides/metabolismo , Animales , Astrocitos/efectos de los fármacos , Astrocitos/metabolismo , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Células Cultivadas , Ratones , Ratones Noqueados , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Trastornos Psicóticos/etiología , Trastornos Psicóticos/metabolismo , Piridinas/farmacología , Ratas , Ratas Sprague-Dawley , Receptores de Glutamato Metabotrópico/metabolismo , Receptores de Glutamato Metabotrópico/fisiología , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Sulfonamidas/farmacología , Factor de Crecimiento Transformador beta1/farmacología , Xantenos/farmacologíaRESUMEN
Group II metabotropic glutamate receptors (mGluRs) couple to the inhibitory G-protein Gi. The group II mGluRs include two subtypes, mGlu2 and mGlu3, and their pharmacological activation produces analgesic effects in inflammatory and neuropathic pain states. However, the specific contribution of each one of the two subtypes has not been clarified due to the lack of selective orthosteric ligands that can discriminate between mGlu2 and mGlu3 subtypes.In this study we used mGlu2 or mGlu3 knock-out mice to dissect the specific role for these two receptors in the endogenous control of inflammatory pain and their specific contribution to the analgesic activity of mixed mGlu2/3 receptor agonists.Our results showed that mGlu2â»(/)â» mice display a significantly greater pain response compared to their wild type littermates. Interestingly the increased pain sensitivity in mGlu2â»(/)â» mice occurred only in the second phase of the formalin test. No differences were observed in the first phase. In contrast, mGlu3â»(/)â» mice did not significantly differ from their wild type littermates in either phase of the formalin test.When systemically injected, a single administration of the mGlu2/3 agonist, LY379268 (3 mg/kg, ip), showed a significant reduction of both phases in wild-type mice and in mGlu3â»(/)â» but not in mGlu2â»(/)â» mice. However tolerance to the analgesic effect of LY379268 (3 mg/kg, ip) in mGlu3â»(/)â» mice developed following 5 consecutive days of injection.Taken together, these results demonstrate that: (i) mGlu2 receptors play a predominant role over mGlu3 receptors in the control of inflammatory pain in mice; (ii) the analgesic activity of mixed mGlu2/3 agonists is entirely mediated by the activation of the mGlu2 subtype and (iii) the development of tolerance to the analgesic effect of mGlu2/3 agonists develops despite the lack of mGlu3 receptors.
Asunto(s)
Aminoácidos/farmacología , Analgésicos/farmacología , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Inflamación/patología , Dolor/patología , Receptores de Glutamato Metabotrópico/agonistas , Aminoácidos/administración & dosificación , Animales , Compuestos Bicíclicos Heterocíclicos con Puentes/administración & dosificación , Relación Dosis-Respuesta a Droga , Inflamación/complicaciones , Inflamación/fisiopatología , Ratones , Ratones Endogámicos C57BL , Actividad Motora/efectos de los fármacos , Dolor/complicaciones , Dolor/fisiopatología , Dimensión del Dolor , Receptores de Glutamato Metabotrópico/deficiencia , Receptores de Glutamato Metabotrópico/metabolismo , Prueba de Desempeño de Rotación con Aceleración ConstanteRESUMEN
BACKGROUND: Kisspeptin is a neuropeptide known for its role in the hypothalamic regulation of the reproductive axis. Following the recent description of kisspeptin and its 7-TM receptor, GPR54, in the dorsal root ganglia and dorsal horns of the spinal cord, we examined the role of kisspeptin in the regulation of pain sensitivity in mice. RESULTS: Immunofluorescent staining in the mouse skin showed the presence of GPR54 receptors in PGP9.5-positive sensory fibers. Intraplantar injection of kisspeptin (1 or 3 nmol/5 µl) induced a small nocifensive response in naive mice, and lowered thermal pain threshold in the hot plate test. Both intraplantar and intrathecal (0.5 or 1 nmol/3 µl) injection of kisspeptin caused hyperalgesia in the first and second phases of the formalin test, whereas the GPR54 antagonist, p234 (0.1 or 1 nmol), caused a robust analgesia. Intraplantar injection of kisspeptin combined with formalin enhanced TRPV1 phosphorylation at Ser800 at the injection site, and increased ERK1/2 phosphorylation in the ipsilateral dorsal horn as compared to naive mice and mice treated with formalin alone. CONCLUSION: These data demonstrate for the first time that kisspeptin regulates pain sensitivity in rodents and suggest that peripheral GPR54 receptors could be targeted by novel drugs in the treatment of inflammatory pain.
Asunto(s)
Hiperalgesia/metabolismo , Kisspeptinas/metabolismo , Neuropéptidos/metabolismo , Animales , Técnica del Anticuerpo Fluorescente Indirecta , Hiperalgesia/tratamiento farmacológico , Hiperalgesia/fisiopatología , Masculino , Ratones , Ratones Endogámicos , Umbral del Dolor , Receptores Acoplados a Proteínas G/metabolismo , Receptores de Kisspeptina-1RESUMEN
Anabolic-androgenic steroid (AAS) abuse is associated with multiple neurobehavioral disturbances. The sites of action and the neurobiological sequels of AAS abuse are unclear at present. We investigated whether two different AASs, nandrolone and methandrostenolone, could affect neuronal survival in culture. The endogenous androgenic steroid testosterone was used for comparison. Both testosterone and nandrolone were neurotoxic at micromolar concentrations, and their effects were prevented by blockade of androgen receptors (ARs) with flutamide. Neuronal toxicity developed only over a 48-hr exposure to the steroids. The cell-impermeable analogues testosterone-BSA and nandrolone-BSA, which preferentially target membrane-associated ARs, were also neurotoxic in a time-dependent and flutamide-sensitive manner. Testosterone-BSA and nandrolone-BSA were more potent than their parent compounds, suggesting that membrane-associated ARs were the relevant sites for the neurotoxic actions of the steroids. Unlike testosterone and nandrolone, toxicity by methandrostenolone and methandrostenolone-BSA was insensitive to flutamide, but it was prevented by the glucocorticoid receptor (GR) antagonist RU-486. Methandrostenolone-BSA was more potent than the parent compound, suggesting that its toxicity relied on the preferential activation of putative membrane-associated GRs. Consistently with the evidence that membrane-associated GRs can mediate rapid effects, a brief challenge with methandrostenolone-BSA was able to promote neuronal toxicity. Activation of putative membrane steroid receptors by nontoxic (nanomolar) concentrations of either nandrolone-BSA or methandrostenolone-BSA became sufficient to increase neuronal susceptibility to the apoptotic stimulus provided by ß-amyloid (the main culprit of AD). We speculate that AAS abuse might facilitate the onset or progression of neurodegenerative diseases not usually linked to drug abuse.
Asunto(s)
Anabolizantes/toxicidad , Metandrostenolona/toxicidad , Nandrolona/toxicidad , Neuronas/efectos de los fármacos , Síndromes de Neurotoxicidad/metabolismo , Andrógenos/toxicidad , Animales , Astrocitos/efectos de los fármacos , Astrocitos/metabolismo , Astrocitos/patología , Western Blotting , Muerte Celular/efectos de los fármacos , Células Cultivadas , Técnicas de Cocultivo , Técnica del Anticuerpo Fluorescente , Microscopía Confocal , Neuronas/metabolismo , Neuronas/patología , Síndromes de Neurotoxicidad/patología , Ratas , Ratas Sprague-Dawley , Receptores Androgénicos/metabolismo , Receptores de Glucocorticoides/metabolismo , Albúmina Sérica Bovina/farmacologíaRESUMEN
Research on cerebral glucose metabolism has shown that the aging brain experiences a fall of aerobic glycolysis, and that the age-related loss of aerobic glycolysis may accelerate Alzheimer's disease pathology. In the healthy brain, aerobic glycolysis, namely the use of glucose outside oxidative phosphorylation, may cover energy demand and increase neuronal resilience to stressors at once. Currently, the drivers of aerobic glycolysis in neurons are unknown. We previously demonstrated that synthetic monomers of ß-amyloid protein (Aß) enhance glucose uptake in neurons, and that endogenous Aß is required for depolarization-induced glucose uptake in cultured neurons. In this work, we show that cultured cortical neurons increased aerobic glycolysis in response to the inhibition of oxidative phosphorylation by oligomycin or to a kainate pulse. Such an increase was prevented by blocking the endogenous Aß tone and re-established by the exogenous addition of synthetic Aß monomers. The activity of mitochondria-bound hexokinase-1 appeared to be necessary for monomers-stimulated aerobic glycolysis during oxidative phosphorylation blockade or kainate excitation. Our data suggest that, through Aß release, neurons coordinate glucose uptake with aerobic glycolysis in response to metabolic stressors. The implications of this new finding are that the age-related drop in aerobic glycolysis and the susceptibility to Alzheimer's disease could be linked to factors interfering with release and functions of Aß monomers.
Asunto(s)
Envejecimiento/metabolismo , Péptidos beta-Amiloides/metabolismo , Glucosa/metabolismo , Neuronas/metabolismo , Enfermedad de Alzheimer/metabolismo , Animales , Encéfalo/metabolismo , Metabolismo Energético , Glucólisis , Fosforilación Oxidativa , RatasRESUMEN
The 42-aa-long beta-amyloid protein--Abeta(1-42)--is thought to play a central role in the pathogenesis of Alzheimer's disease (AD) (Walsh and Selkoe, 2007). Data from AD brain (Shankar et al., 2008), transgenic APP (amyloid precursor protein)-overexpressing mice (Lesné et al., 2006), and neuronal cultures treated with synthetic Abeta peptides (Lambert et al., 1998) indicate that self-association of Abeta(1-42) monomers into soluble oligomers is required for neurotoxicity. The function of monomeric Abeta(1-42) is unknown. The evidence that Abeta(1-42) is present in the brain and CSF of normal individuals suggests that the peptide is physiologically active (Shoji, 2002). Here we show that synthetic Abeta(1-42) monomers support the survival of developing neurons under conditions of trophic deprivation and protect mature neurons against excitotoxic death, a process that contributes to the overall neurodegeneration associated with AD. The neuroprotective action of Abeta(1-42) monomers was mediated by the activation of the PI-3-K (phosphatidylinositol-3-kinase) pathway, and involved the stimulation of IGF-1 (insulin-like growth factor-1) receptors and/or other receptors of the insulin superfamily. Interestingly, monomers of Abeta(1-42) carrying the Arctic mutation (E22G) associated with familiar AD (Nilsberth et al., 2001) were not neuroprotective. We suggest that pathological aggregation of Abeta(1-42) may also cause neurodegeneration by depriving neurons of the protective activity of Abeta(1-42) monomers. This "loss-of-function" hypothesis of neuronal death should be taken into consideration when designing therapies aimed at reducing Abeta burden.
Asunto(s)
Péptidos beta-Amiloides/farmacología , Neuronas/efectos de los fármacos , Fármacos Neuroprotectores/farmacología , Fragmentos de Péptidos/farmacología , Péptidos beta-Amiloides/química , Análisis de Varianza , Animales , Butadienos/farmacología , Muerte Celular/efectos de los fármacos , Células Cultivadas , Corteza Cerebral/citología , Relación Dosis-Respuesta a Droga , Embrión de Mamíferos , Inhibidores Enzimáticos/farmacología , Agonistas de Aminoácidos Excitadores/toxicidad , N-Metilaspartato/toxicidad , Nitrilos/farmacología , Fragmentos de Péptidos/química , Podofilotoxina/análogos & derivados , Podofilotoxina/farmacología , Ratas , Tirfostinos/farmacologíaRESUMEN
Metallothioneins (MTs) are metal-binding proteins that are overexpressed in various human cancers and are thought to be associated with resistance to cytotoxic drugs. The knowledge on MT expression, regulation, and function in human gliomas is limited. We found that MT3 mRNA was highly expressed in cell lines derived from grade IV gliomas (i.e., A172 and U87 cells), as compared to grade II astrocytoma cells (i.e., 1321N1). Different from 1321N1, U87 cells were partly resistant to the alkylating drug, temozolomide (TMZ) (100 µM for 96 h), which induced a massive accumulation of U87 into the S and G2 fractions of the cell cycle but not apoptotic death. Silencing of MT3 did not significantly affect U87 cell proliferation and survival, but it delayed G1/S transition and favored the occurrence of apoptosis in TMZ-treated cells. Accordingly, the combination of MT3 silencing and TMZ treatment increased the protein levels of checkpoint kinase-1, which was ultimately responsible for the lasting G1 arrest and death of double treated U87 cells.
RESUMEN
Estrogen receptor alpha (ERalpha) is over-expressed in reactive glia under conditions of neuronal damage. To elucidate the functional significance of ERalpha overexpression, an in vitro model of reactive astrocytes with enhanced expression of ERalpha was obtained by growth in G5 culture supplement. Exposure of cortical neurons to beta-amyloid in the presence of either conditioned medium from reactive astrocytes previously treated with 17beta-estradiol (17betaE2) or transferring of 17betaE2-pretreated astrocytes, caused a greater neuroprotective effect compared to the respective control conditions, although reactive glia resulted being per se neuroprotective. Blockade of ERalpha overexpression by the ER antagonist ICI182,780 was not successful as ICI182,780 behaved as an agonist. However, complete prevention of 17betaE2 effect by ICI182,780 produced an increased sensitivity of neurons to beta-amyloid toxicity. A similar effect was observed when ERalpha knock-down was induced by siRNA. It is suggested that increased ERalpha expression in reactive glia may have a role in limiting neuronal damage.