Genistein protects against amyloid-beta-induced toxicity in SH-SY5Y cells by regulation of Akt and Tau phosphorylation.

Alzheimer's disease is a neurodegenerative disorder characterized by extracellular deposition of amyloid-ß (Aß) peptide and hyperphosphorylation of Tau protein, which ultimately leads to the formation of intracellular neurofibrillary tangles and cell death. Increasing evidence indicates that genistein, a soy isoflavone, has neuroprotective effects against Aß-induced toxicity. However, the molecular mechanisms involved in its neuroprotection are not well understood. In this study, we have established a neuronal damage model using retinoic-acid differentiated SH-SY5Y cells treated with different concentrations of Aß25-35 to investigate the effect of genistein against Aß-induced cell death and the possible involvement of protein kinase B (PKB, also termed Akt), glycogen synthase kinase 3ß (GSK-3ß), and Tau as an underlying mechanism to this neuroprotection. Differentiated SH-SY5Y cells were pre-treated for 24 hr with genistein (1 and 10 nM) and exposed to Aß25-35 (25 µM), and we found that genistein partially inhibited Aß induced cell death, primarily apoptosis. Furthermore, the protective effect of genistein was associated with the inhibition of Aß-induced Akt inactivation and Tau hyperphosphorylation. These findings reinforce the neuroprotective effects of genistein against Aß toxicity and provide evidence that its mechanism may involve regulation of Akt and Tau proteins.

Redispersible Spray-Dried Powder Containing Nanoencapsulated Curcumin: the Drying Process Does Not Affect Neuroprotection In vitro.

A redispersible spray-dried formulation containing curcumin-loaded, lipid-core nanocapsules (LNC-C) was developed for oral administration. The neuroprotective activity of curcumin after the spray-drying process was evaluated in vitro. The spray-dried powder (SD-LNC-C) was produced using a drying adjuvant composed of a blend of maltodextrin and L-leucine (90:10 w/w). Acceptable process yield (~ 70%) and drug content (6.5 ± 0.2 mg g-1) were obtained. SD-LNC-C was formed by smooth, spherical-shaped particles, and confocal Raman analysis indicated the distribution of the LNC-C on the surface of the leucine/maltodextrin agglomerates. The surface of the agglomerates was formed by a combination of LNC-C and adjuvants, and laser diffraction showed that SD-LNC-C had adequate aqueous redispersion, with no loss of controlled drug release behaviour of LNC-C. The in vitro curcumin activity against the lipopolysaccharide (LPS)-induced proinflammatory response in organotypic hippocampal slice cultures was evaluated. Both formulations (LNC-C and SD-LNC-C) reduced TNF-α to similar levels. Therefore, neuroprotection of curcumin in vitro may be improved by nanoencapsulation followed by spray-drying, with no loss of this superior performance. Hence, the redispersible spray-dried powder proposed here represents a suitable approach for the development of innovative nanomedicines containing curcumin for the prevention/treatment of neurodegenerative diseases.

Free and nanoencapsulated curcumin suppress ß-amyloid-induced cognitive impairments in rats: involvement of BDNF and Akt/GSK-3ß signaling pathway.

Alzheimer's disease (AD), a neurodegenerative disorder exhibiting progressive loss of memory and cognitive functions, is characterized by the presence of neuritic plaques composed of neurofibrillary tangles and ß-amyloid (Aß) peptide. Drug delivery to the brain still remains highly challenging for the treatment of AD. Several studies have been shown that curcumin is associated with anti-amyloidogenic properties, but therapeutic application of its beneficial effects is limited. Here we investigated possible mechanisms involved in curcumin protection against Aß(1-42)-induced cognitive impairment and, due to its poor bioavailability, we developed curcumin-loaded lipid-core nanocapsules in an attempt to improve the neuroprotective effect of this polyphenol. Animals received a single intracerebroventricular injection of Aß(1-42) and they were administered either free curcumin or curcumin-loaded lipid-core nanocapsules (Cur-LNC) intraperitoneally for 10days. Aß(1-42)-infused animals showed a significant impairment on learning-memory ability, which was paralleled by a significant decrease in hippocampal synaptophysin levels. Furthermore, animals exhibited activated astrocytes and microglial cells, as well as disturbance in BDNF expression and Akt/GSK-3ß signaling pathway, beyond tau hyperphosphorylation. Our findings demonstrate that administration of curcumin was effective in preventing behavioral impairments, neuroinflammation, tau hyperphosphorylation as well as cell signaling disturbances triggered by Aß in vivo. Of high interest, Cur-LNC in a dose 20-fold lower presented similar neuroprotective results compared to the effective dose of free curcumin. Considered overall, the data suggest that curcumin is a potential therapeutic agent for neurocognition and nanoencapsulation of curcumin in LNC might constitute a promising therapeutic alternative in the treatment of neurodegenerative diseases such as AD.

Alterations on Na⁺,K⁺-ATPase and acetylcholinesterase activities induced by amyloid-ß peptide in rat brain and GM1 ganglioside neuroprotective action.

Alzheimer's disease (AD) is a neurodegenerative disorder whose pathogenesis involves production and aggregation of amyloid-ß peptide (Aß). Aß-induced toxicity is believed to involve alterations on as Na(+),K(+)-ATPase and acetylcholinesterase (AChE) activities, prior to neuronal death. Drugs able to prevent or to reverse these biochemical changes promote neuroprotection. GM1 is a ganglioside proposed to have neuroprotective roles in AD models, through mechanisms not yet fully understood. Therefore, this study aimed to investigate the effect of Aß1-42 infusion and GM1 treatment on recognition memory and on Na(+),K(+)-ATPase and AChE activities, as well as, on antioxidant defense in the brain cortex and the hippocampus. For these purposes, Wistar rats received i.c.v. infusion of fibrilar Aß1-42 (2 nmol) and/or GM1 (0.30 mg/kg). Behavioral and biochemical analyses were conducted 1 month after the infusion procedures. Our results showed that GM1 treatment prevented Aß-induced cognitive deficit, corroborating its neuroprotective function. Aß impaired Na(+),K(+)-ATPase and increase AChE activities in hippocampus and cortex, respectively. GM1, in turn, has partially prevented Aß-induced alteration on Na(+),K(+)-ATPase, though with no impact on AChE activity. Aß caused a decrease in antioxidant defense, specifically in hippocampus, an effect that was prevented by GM1 treatment. GM1, both in cortex and hippocampus, was able to increase antioxidant scavenge capacity. Our results suggest that Aß-triggered cognitive deficit involves region-specific alterations on Na(+),K(+)-ATPase and AChE activities, and that GM1 neuroprotection involves modulation of Na(+),K(+)-ATPase, maybe by its antioxidant properties. Although extrapolation from animal findings is difficult, it is conceivable that GM1 could play an important role in AD treatment.

Amburana cearensis seed extract stimulates astrocyte glutamate homeostatic mechanisms in hippocampal brain slices and protects oligodendrocytes against ischemia.

BACKGROUND: Stroke is a leading cause of death and disability worldwide. A major factor in brain damage following ischemia is excitotoxicity caused by elevated levels of the neurotransmitter glutamate. In the brain, glutamate homeostasis is a primary function of astrocytes. Amburana cearensis has long been used in folk medicine and seed extract obtained with dichloromethane (EDAC) have previously been shown to exhibit cytoprotective activity in vitro. The aim of the present study was to analyse the activity of EDAC in hippocampal brain slices. METHODS: We prepared a dichloromethane extract (EDAC) from A. cearensis seeds and characterized the chemical constituents by 1H and 13C-NMR. Hippocampal slices from P6-8 or P90 Wistar rats were used for cell viability assay or glutamate uptake test. Hippocampal slices from P10-12 transgenic mice SOX10-EGFP and GFAP-EGFP and immunofluorescence for GS, GLAST and GLT1 were used to study oligodendrocytes and astrocytes. RESULTS: Astrocytes play a critical role in glutamate homeostasis and we provide immunohistochemical evidence that in excitotoxicity EDAC increased expression of glutamate transporters and glutamine synthetase, which is essential for detoxifying glutamate. Next, we directly examined astrocytes using transgenic mice in which glial fibrillary acidic protein (GFAP) drives expression of enhanced green fluorescence protein (EGFP) and show that glutamate excitotoxicity caused a decrease in GFAP-EGFP and that EDAC protected against this loss. This was examined further in the oxygen-glucose deprivation (OGD) model of ischemia, where EDAC caused an increase in astrocytic process branching, resulting in an increase in GFAP-EGFP. Using SOX10-EGFP reporter mice, we show that the acute response of oligodendrocytes to OGD in hippocampal slices is a marked loss of their processes and EDAC protected oligodendrocytes against this damage. CONCLUSION: This study provides evidence that EDAC is cytoprotective against ischemia and glutamate excitotoxicity by modulating astrocyte responses and stimulating their glutamate homeostatic mechanisms.

Resveratrol prevents CA1 neurons against ischemic injury by parallel modulation of both GSK-3ß and CREB through PI3-K/Akt pathways.

Accumulating evidence indicates that resveratrol potently protects against cerebral ischemia damage due to its oxygen free radicals scavenging and antioxidant properties. However, cellular mechanisms that may underlie the neuroprotective effects of resveratrol in brain ischemia are not fully understood yet. This study aimed to investigate the potential association between the neuroprotective effect of resveratrol and the apoptosis/survival signaling pathways, in particular the glycogen synthase kinase 3 (GSK-3ß) and cAMP response element-binding protein (CREB) through phosphatidylinositol 3-kinase (PI3-K)-dependent pathway. An experimental model of global cerebral ischemia was induced in rats by the four-vessel occlusion method for 10 min and followed by different periods of reperfusion. Nissl staining indicated extensive neuronal death at 7 days after ischemia/reperfusion. Administration of resveratrol by i.p. injections (30 mg/kg) for 7 days before ischemia significantly attenuated neuronal death. Both GSK-3ß and CREB appear to play a critical role in resveratrol neuroprotection through the PI3-K/Akt pathway, as resveratrol pretreatment increased the phosphorylation of Akt, GSK-3ß and CREB in 1 h in the CA1 hippocampus after ischemia/reperfusion. Furthermore, administration of LY294002, an inhibitor of PI3-K, compromised the neuroprotective effect of resveratrol and decreased the level of p-Akt, p-GSK-3ß and p-CREB after ischemic injury. Taken together, the results suggest that resveratrol protects against delayed neuronal death in the hippocampal CA1 by maintaining the pro-survival states of Akt, GSK-3ß and CREB pathways. These data suggest that the neuroprotective effect of resveratrol may be mediated through activation of the PI3-K/Akt signaling pathway, subsequently downregulating expression of GSK-3ß and CREB, thereby leading to prevention of neuronal death after brain ischemia in rats.

Genistein prevents the decrease in ganglioside levels induced by amyloid-beta in the frontal cortex of rats.

OBJECTIVES: In this study, an in vivo model of Aß toxicity was used to investigate the effects of this peptide and the treatment with genistein on the lipid composition (gangliosides, phospholipids and cholesterol) in the frontal cortex of rats. METHODS: Male Wistar rats received bilateral intracerebroventricular infusions of Aß1-42 (2 nmol) and genistein 10 mg/kg orally for 10 days. Frontal cortex was homogenized with chloroform:methanol for lipid extraction and ganglioside, phospholipid and cholesterol levels were evaluated. RESULTS: The Aß-infused animals showed a significant decrease in ganglioside concentration and relative reduction of GD1b and GQ1b species. Treatment with genistein prevented the decrease in ganglioside levels. Phospholipid and cholesterol contents did not show significant differences. DISCUSSION: Considering the roles of gangliosides on neuronal function, findings described here can contribute to the knowledge of the potential neuroprotective mechanisms of genistein against Aß-induced alterations in the frontal cortex of rats and provide a novel view in the multifaceted scenario associated with its beneficial effects.

Chronic stress and lithium treatments alter hippocampal glutamate uptake and release in the rat and potentiate necrotic cellular death after oxygen and glucose deprivation.

This study was undertaken to evaluate the effects of chronic variate stress and lithium treatment on glutamatergic activity and neuronal vulnerability of rat hippocampus. Male Wistar rats were simultaneously treated with lithium and submitted to a chronic variate stress protocol during 40 days, and afterwards the hippocampal glutamatergic uptake and release, measured in slices and synaptosomes, were evaluated. We observed an increased synaptosomal [(3)H]glutamate uptake and an increase in [(3)H]glutamate stimulated release in hippocampus of lithium-treated rats. Chronic stress increased basal [(3)H]glutamate release by synaptosomes, and decreased [(3)H]glutamate uptake in hippocampal slices. When evaluating cellular vulnerability, both stress and lithium increased cellular death after oxygen and glucose deprivation (OGD). We suggest that the manipulation of glutamatergic activity induced by stress may be in part responsible for the neuroendangerment observed after stress exposure, and that, in spite of the described neuroprotective effects of lithium, it increased the neuronal vulnerability after OGD.

Genistein attenuates amyloid-beta-induced cognitive impairment in rats by modulation of hippocampal synaptotoxicity and hyperphosphorylation of Tau.

Alzheimer's disease is a progressive neurodegenerative disorder characterized by extracellular accumulation of amyloid-beta (Aß) peptide, which induces synaptic dysfunction, alteration of intracellular signaling pathways, hyperphosphorylation of the Tau protein, and cognitive impairment. Genistein, one of the major isoflavones present in soy and soy products, has been shown to modulate some of the pathogenic events associated with the neurodegeneration process. However, its underlying mechanisms remain to be clarified. Therefore, the objectives of the present study were to evaluate the ability of genistein to protect against Aß1-42-induced cognitive impairment in rats and to elucidate some of the possible mechanisms involved in its neuroprotective effects in the hippocampus. Male Wistar rats received bilateral intracerebroventricular infusions of Aß1-42 (2 nmol) and genistein 10 mg/kg orally for 10 days. The Aß-infused animals showed significant impairment of memory, which was accompanied by the following neurochemical alterations in the hippocampus: decreased levels of the synaptic proteins synaptophysin and postsynaptic density protein 95 (PSD-95), hyperphosphorylation of Tau with increased activation of glycogen synthase kinase-3ß and c-Jun N-terminal kinase, and inactivation of ERK. Treatment with genistein improved Aß-induced cognitive impairment by attenuation of synaptotoxicity, hyperphosphorylation of Tau, and inactivation of ERK. Furthermore, treatment with this soy isoflavone did not cause systemic toxicity. These findings provide further evidence of the neuroprotective effect of genistein in an in vivo model of Aß toxicity and, importantly, extend the current knowledge concerning the mechanisms associated with the neuroprotective effects of this compound in the hippocampus.

Novel arylidene malonate derivative, KM-34, showed neuroprotective effects on in vitro and in vivo models of ischemia/reperfusion.

Cerebral ischemia constitutes the most frequent type of cerebrovascular disease. The reduction of blood supply to the brain initiates the ischemic cascade starting from ionic imbalance to subsequent glutamate excitotoxicity, neuroinflammation and oxidative stress, eventually causing neuronal death. Previously, the authors have demonstrated the in vitro cytoprotective and antioxidant effects of a new arylidene malonate derivative, KM-34, against oxidizing agents like hydrogen peroxide, glutamate or Fe3+/ascorbate. Here, we examined for the first time the neuroprotective effect of KM-34 on ischemia/reperfusion models. In vitro, treatment with 10 and 50 µM KM-34 reduced the cellular death (propidium iodide incorporation) induced by oxygen glucose deprivation (OGD) in rat organotypic hippocampal slices cultures. In vivo, stroke was induced in male Wistar rats through middle cerebral artery occlusion (MCAO), followed by 23 h of reperfusion. KM-34 was orally administered 105 min after MCAO onset. We noticed that 1 mg/kg KM-34 reduced infarct volume and neurological score, and increased the latency to fall in the Hanging Wire test compared to vehicle-treated ischemic animals. While ischemic and sham-operated groups showed similar horizontal locomotor activity, vertical counts decreased after MCAO, suggesting that vertical movements are more sensitive to the ischemic injury. Treatment with KM-34 also alleviated the mitochondrial impairment (ROS generation, swelling and membrane potential dissipation) induced by transient MCAO but not significant alterations were found in oxidative stress parameters. Overall, the study provides preclinical evidences confirming the neuroprotective effects of a novel synthetic molecule and paved the way for future investigations regarding its therapeutic potential against brain ischemia/reperfusion injury.

In-depth quantitative proteomic characterization of organotypic hippocampal slice culture reveals sex-specific differences in biochemical pathways.

Sex differences in the brain of mammals range from neuroarchitecture through cognition to cellular metabolism. The hippocampus, a structure mostly associated with learning and memory, presents high vulnerability to neurodegeneration and aging. Therefore, we explored basal sex-related differences in the proteome of organotypic hippocampal slice culture, a major in vitro model for studying the cellular and molecular mechanisms related to neurodegenerative disorders. Results suggest a greater prevalence of astrocytic metabolism in females and significant neuronal metabolism in males. The preference for glucose use in glycolysis, pentose phosphate pathway and glycogen metabolism in females and high abundance of mitochondrial respiration subunits in males support this idea. An overall upregulation of lipid metabolism was observed in females. Upregulation of proteins responsible for neuronal glutamate and GABA synthesis, along with synaptic associated proteins, were observed in males. In general, the significant spectrum of pathways known to predominate in neurons or astrocytes, together with the well-known neuronal and glial markers observed, revealed sex-specific metabolic differences in the hippocampus. TEM qualitative analysis might indicate a greater presence of mitochondria at CA1 synapses in females. These findings are crucial to a better understanding of how sex chromosomes can influence the physiology of cultured hippocampal slices and allow us to gain insights into distinct responses of males and females on neurological diseases that present a sex-biased incidence.

Na+, K+ ATPase activity is reduced in amygdala of rats with chronic stress-induced anxiety-like behavior.

In this study, we examined the effects of two chronic stress regimens upon anxiety-like behavior, Na(+), K(+)-ATPase activity and immunocontent, and oxidative stress parameters (antioxidant enzymes and reactive oxygen species production) in the amygdala. Male rats were subjected to chronic unpredictable and to chronic restraint stress for 40 days. Subsequently, anxiety-like behavior was examined. Both stressed groups presented increased anxiety-like behavior. Reduced amygdalal Na(+), K(+)-ATPase activity in the synaptic plasma membranes was also observed, without alterations in the amygdala immunocontent. In addition, when analyzing oxidative stress parameters, only superoxide dismutase activity was decreased in the amygdala of animals subjected to unpredictable stress. We conclude that both models of chronic stress lead to anxiety-like behavior and decreased amygdalal Na(+), K(+)-ATPase activity, which appears not to be related to oxidative imbalance. The relationship between this decreased activity and anxiety-like behavior remains to be studied.

Amyloid-beta neurotoxicity in organotypic culture is attenuated by melatonin: involvement of GSK-3beta, tau and neuroinflammation.

Alzheimer's disease (AD) is a progressive neurodegenerative disorder marked by accumulation of extracellular deposits of amyloid-beta (Abeta) peptide in brain regions that are important for memory and cognition. The buildup of Abeta aggregates in the AD is followed by the formation of intracellular neurofibrillary tangles and activation of neuroinflammatory reactions. The present study investigated whether melatonin possesses a neuroprotective effect against Abeta-induced toxicity. For this purpose, organotypic hippocampal slices were cultured and exposed to 25 microm of Abeta(25-35) in the absence or in the presence of melatonin (25, 50, or 100 microm). In addition, the authors have investigated the involvement of GSK-3beta, tau protein, astroglial, and microglial activation, and cytokine levels in the melatonin protection against Abeta-induced neurotoxicity. Melatonin prevented the cell damage in hippocampus induced by the exposure to Abeta(25-35). In addition, melatonin significantly reduced the activation of GSK-3beta, the phosphorylation of tau protein, the glial activation and the Abeta-induced increase of TNF-alpha and IL-6 levels. On the basis of these findings, we speculate that melatonin may provide an effective therapeutic strategy for AD, by attenuating Abeta-induced phosphorylation of tau protein, and preventing GSK-3beta activation and neuroinflammation.

Protective effect of maternal exercise against amyloid-ß neurotoxicity in the male rat offspring's cerebellum.

The Developmental Origins of Health and Disease (DOHaD) states that intrauterine maternal environment influences postnatal life by programming offspring's metabolism. Intrauterine milieu induced by exercise during pregnancy promotes long-lasting benefits to the offspring's health and seems to offer some resistance against chronic diseases in adult life. Alzheimer's disease is a public health concern with limited treatment options. In the present study, we assessed the potential of maternal exercise during pregnancy in long-term programming of young adult male rat offspring's cerebellar metabolism in conferring neuroprotection against amyloid-ß (Aß) neurotoxicity. Female Wistar rats were submitted to a swimming protocol 1 week prior mating and throughout pregnancy (five sessions/a week lasting 30 min). Aß oligomers were infused bilaterally in the brain ventricles of 60-day-old male offspring. Fourteen days after surgery, we measured parameters related to redox state, mitochondrial function, and the immunocontent of proteins related to synaptic function. We found that maternal exercise during pregnancy attenuated several parameters in the offspring's male rat cerebellum, such as the reactive species rise, the increase of inducible nitric oxide synthase immunocontent and tau phosphorylation induced by Aß oligomers, increased mitochondrial fission indicated by dynamin-related protein 1 (DRP1), and protein oxidation identified by carbonylation. Strikingly, we find that maternal exercise promotes changes in the rat offspring's cerebellum that are still evident in young adult life. These favorable neurochemical changes in offspring's cerebellum induced by maternal exercise may contribute to a protective phenotype against Aß-induced neurotoxicity in young adult male rat offspring.

Resveratrol and quercetin cooperate to induce senescence-like growth arrest in C6 rat glioma cells.

Glioma is the most frequent and malignant primary human brain tumor with dismal prognosis despite multimodal therapy. Resveratrol and quercetin, two structurally related and naturally occurring polyphenols, are proposed to have anticancer effects. We report here that resveratrol and quercetin decreased the cell number in four glioma cell lines but not in rat astrocytes. Low doses of resveratrol (10 microM) or quercetin (25 microM) separately had no effect on apoptosis induction, but had a strong effect on caspase 3/7 activation when administered together. Western blot analyses showed that resveratrol (10 microM) and quercetin (25 microM) caused a reduction in phosphorylation of Akt, but this reduction was not sufficient by itself to mediate the effects of these polyphenols. Most important, resveratrol and quercetin chronically administered presented a strong synergism in inducing senescence-like growth arrest. These results suggest that the combination of polyphenols can potentialize their antitumoral activity, thereby reducing the therapeutic concentration needed for glioma treatment.

Boldine: a potential new antiproliferative drug against glioma cell lines.

Malignant gliomas are the most common and devastating primary tumors of the central nervous system. Currently no efficient treatment is available. This study evaluated the effect and underlying mechanisms of boldine, an aporphine alkaloid of Peumus boldus, on glioma proliferation and cell death. Boldine decreased the cell number of U138-MG, U87-MG and C6 glioma lines at concentrations of 80, 250 and 500 muM. We observed that cell death caused by boldine was cell-type specific and dose-dependent. Exposure to boldine for 24 h did not activate key mediators of apoptosis. However, it induced alterations in the cell cycle suggesting a G(2)/M arrest in U138-MG cells. Boldine had no toxic effect on non-tumor cells when used at the same concentrations as those used on tumor cells. Based on these results, we speculate that boldine may be a promising compound for evaluation as an anti-cancer agent.

Conditioned medium from mesenchymal stem cells induces cell death in organotypic cultures of rat hippocampus and aggravates lesion in a model of oxygen and glucose deprivation.

Cell therapy using bone marrow-derived mesenchymal stem cells (MSC) seems to be a new alternative for the treatment of neurological diseases, including stroke. In order to investigate the response of hippocampal tissue to factors secreted by MSC and if these factors are neuroprotective in a model of oxygen and glucose deprivation (OGD), we used organotypic hippocampal cultures exposed to conditioned medium from bone marrow-derived MSC. Our results suggest that the conditioned medium obtained from these cells aggravates lesion caused by OGD. In addition, the presence of the conditioned medium alone was toxic mainly to cells in the CA1, CA2 and CA3 areas of the hippocampal organotypic culture even in basal conditions. GABA stimulation and NMDA and AMPA receptors antagonists were able to reduce propidium iodide staining, suggesting that the cell death induced by the toxic factors secreted by MSC could involve these receptors.

A comparative study of beta-amyloid peptides Abeta1-42 and Abeta25-35 toxicity in organotypic hippocampal slice cultures.

Accumulation of the neurotoxic amyloid beta-peptide (Abeta) in the brain is a hallmark of Alzheimer's disease (AD). Several synthetic Abeta peptides have been used to study the mechanisms of toxicity. Here, we sought to establish comparability between two commonly used Abeta peptides Abeta1-42 and Abeta25-35 on an in vitro model of Abeta toxicity. For this purpose we used organotypic slice cultures of rat hippocampus and observed that both Abeta peptides caused similar toxic effects regarding to propidium iodide uptake and caspase-3 activation. In addition, we also did not observe any effect of both peptides on Akt and PTEN phosphorylation; otherwise the phosphorylation of GSK-3beta was increased. Although further studies are necessary for understanding mechanisms underlying Abeta peptide toxicity, our results provide strong evidence that Abeta1-42 and the Abeta25-35 peptides induce neural injury in a similar pattern and that Abeta25-35 is a convenient tool for the investigation of neurotoxic mechanisms involved in AD.

Effects of chronic restraint stress and estradiol replacement on glutamate release and uptake in the spinal cord from ovariectomized female rats.

Glutamate is an excitatory neurotransmitter involved in neuronal plasticity and neurotoxicity. Chronic stress produces several physiological changes on the spinal cord, many of them presenting sex-specific differences, which probably involve glutamatergic system alterations. The aim of the present study was to verify possible effects of exposure to chronic restraint stress and 17beta-estradiol replacement on [3H]-glutamate release and uptake in spinal cord synaptosomes of ovariectomized (OVX) rats. Female rats were subjected to OVX, and half of the animals received estradiol replacement. Animals were subdivided in controls and chronically stressed. Restraint stress or estradiol had no effect on [3H]-glutamate release. The chronic restraint stress promoted a decrease and 17beta-estradiol induced an increase on [3H]-glutamate uptake, but the uptake observed in the restraint stress +17beta-estradiol group was similar to control. Furthermore, 17beta-estradiol treatment caused a significant increase in the immunocontent of the three glutamate transporters present in spinal cord. Restraint stress had no effect on the expression of these transporters, but prevented the 17beta-estradiol effect. We suggest that changes in the glutamatergic system are likely to take part in the mechanisms involved in spinal cord plasticity following repeated stress exposure, and that 17beta-estradiol levels may affect chronic stress effects in this structure.

Dual Effect of Doxazosin: Anticancer Activity on SH-SY5Y Neuroblastoma Cells and Neuroprotection on an In Vitro Model of Alzheimer's Disease.

Several studies have demonstrated the antitumor effect of doxazosin, an α1-adrenergic blocker, against glioma and breast, bladder and prostate cancers. Doxazosin is also being evaluated as a treatment for posttraumatic stress disorder (PTSD) and alcoholism, and α1-adrenergic blockers have been linked to neuroprotection in neurodegenerative disorders, such as Alzheimer's Disease (AD). Cancer and AD have an inverse relationship in many aspects, with several factors that contribute to apoptosis inhibition and proliferation being increased in cancers but decreased in AD. Neuroblastoma (NB) is a pediatric tumor derived from embryonic neural-crest cells, with an overall cure rate of 40%, despite aggressive treatment. Thus, due to the need of new therapeutic strategies against NB and neurodegenerative disorders and the inverse relationship between these diseases, we investigated whether doxazosin may serve as an antitumor and neuroprotective agent. We analyzed the drug's effects on undifferentiated and retinoic acid-differentiated SH-SY5Y human NB cells and on an in vitro model of organotypic hippocampal cultures exposed to amyloid-ß. Doxazosin showed antitumor effect on undifferentiated NB cells by induction of apoptosis, necrosis, cell cycle arrest and decrease of p-EGFRTyr1048 levels. On differentiated cells, doxazosin was less cytotoxic and increased p-EGFRTyr1048, p-AktSer473 and p-GSK-3ßSer9 levels. Moreover, the drug was able to protect hippocampal slices from amyloid-ß toxicity through prevention of GSK-3ß activation and of Tau hyperphosphorylation. Therefore, our results show that doxazosin has antitumor activity against undifferentiated NB and is neuroprotective on an in vitro model of Alzheimer's disease.