Maternal Resveratrol Supplementation Prevents Cognitive Decline in Senescent Mice Offspring.

A variety of environmental factors contribute significantly to age-related cognitive decline and memory impairment in Alzheimer's Disease (AD) and other neurodegenerative diseases. Nutrition can alter epigenetics, improving health outcomes, which can be transmitted across generations; this process is called epigenetic inheritance. We investigate the beneficial effects of maternal resveratrol supplementation in the direct exposed F1 generation and the transgenerational F2 generation. The offspring was generated from females Senescence Accelerated Mouse-Prone (SAMP8) fed a resveratrol-enriched diet for two months prior to mating. Object novel recognition and Morris Water Maze (MWM) demonstrated improvements in cognition in the 6-month-old F1 and F2 generations from resveratrol fed mothers. A significant increase in global DNA methylation with a decrease in hydroxymethylation in F1 and F2 were found. Accordingly, Dnmt3a/b and Tet2 gene expression changed. Methylation levels of Nrf2 and NF-kß genes promoters raised in offspring, inducing changes in target genes expression, as well as hydrogen peroxide levels. Offspring that resulted from a resveratrol fed mother showed increase AMPKα activation, mTOR inhibition, and an increase in Pgc-1α gene expression and Beclin-1 protein levels. Endoplasmic reticulum stress sensors were found changed both in F1 and F2 generations. Overall, our results demonstrated that maternal resveratrol supplementation could prevent cognitive impairment in the SAMP8 mice offspring through epigenetic changes and cell signaling pathways.

Heme Oxygenase-1 and Brain Oxysterols Metabolism Are Linked to Egr-1 Expression in Aged Mice Cortex, but Not in Hippocampus.

Throughout life, stress stimuli act upon the brain leading to morphological and functional changes in advanced age, when it is likely to develop neurodegenerative disorders. There is an increasing need to unveil the molecular mechanisms underlying aging, in a world where populations are getting older. Egr-1 (early growth response 1), a transcriptional factor involved in cell survival, proliferation and differentiation - with a role also in memory, cognition and synaptic plasticity, can be implicated in the molecular mechanism of the aging process. Moreover, Heme Oxygenase-1a (HO), a 32 kDa heat-shock protein that converts heme to iron, carbon monoxide and biliverdin, is a key enzyme with neuroprotective properties. Several in vitro and in vivo studies reported that HO-1 could regulate the metabolism of oxysterols, oxidation products of cholesterol that include markers of oxidative stress. Recently, a link between Egr-1 and HO-1 has been demonstrated in mouse lung cells exposed to cigarette smoke. In view of these data, we wanted to investigate whether Egr-1 can be implicated also in the oxysterol metabolism during brain aging. Our results show that Egr-1 expression is differently expressed in the cortex and hippocampus of old mice, as well as the oxysterol profile between these two brain areas. In particular, we show that the cortex experiences in an age-dependent fashion increasing levels of the Egr-1 protein, and that these correlate with the level of HO-1 expression and oxysterol abundance. Such a situation was not observed in the hippocampus. These results are further strenghtened by our observations made with Egr-1 KO mice, confirming our hypothesis concerning the influence of Egr-1 on oxysterol production and accumulation via regulation of the expression of HO-1 in the cortex, but not the hippocampus, of old mice. It is important to notice that most of the oxysterols involved in this process are those usually stimulated by oxidative stress, which would then represent the triggering factor for this mechanism.

Downregulation of canonical Wnt signaling in hippocampus of SAMP8 mice.

In the adult brain, canonical Wnt (Wnt/ß-catenin) signaling modulates neuronal function, hippocampal neurogenesis, and synaptic plasticity. Indeed, growing evidence suggests that downregulation of Wnt signaling could be involved in the cognitive decline associated with aging and also with the physiopathology of Alzheimer's disease (AD). However, the molecular basis remains unknown. At present, SAMP8 is an experimental model that has been proposed for studying age-related neurodegenerative changes associated with aging and the pathogenesis of AD. Here, we examined Wnt signaling in the hippocampus of SAMP8 mice at 9 and 12 months of age, as well as in its control-strain SAMR1 mice. Our results showed increased Dickkopf-1 protein levels in SAMP8 with age, in addition to GSK-3 α/ß activation and hyperphosphorylated tau. Consequently, higher ß-catenin phosphorylation at Ser(33,37) and Thr(41), which promotes its degradation, along with a decrease in active ß-catenin (ABC) in the nucleus, were observed in SAMP8, mainly at the age of 12 months. Moreover, nuclear levels of Dvl3 were lower in 9- and 12-month-old SAMP8 mice. Related to these findings, SAMP8 showed an increase in neuronal loss in the hippocampus that was associated with lower protein levels of the antiapoptotic protein and the Wnt target gene, Bcl-2, in addition to an increase in the proapototic protein Bax. Our results suggest a relationship between age-related downregulation of canonical Wnt signaling and neuronal loss observed in the hippocampus of SAMP8 mice. Thus, enhancing Wnt signaling may represent a novel neuroprotective strategy aimed at counteracting the cognitive decline that is associated not only with aging but also with AD.

Mavoglurant as a treatment for Parkinson's disease.

INTRODUCTION: A major unresolved issue in the Parkinson's disease (PD) treatment is the development of l-DOPA-induced dyskinesias (LIDs) as a side effect of chronic L-DOPA administration. Currently, LIDs are managed in part by reducing the L-DOPA dose or by the administration of amantadine. However, this treatment is only partially effective. A potential strategy, currently under investigation, is the coadministration of metabotropic glutamate receptor 5 (mGluR5) negative allosteric modulators (NAMs) and L-DOPA; a treatment that results in the improvement of dyskinesia symptoms and that permits reductions in l-DOPA dosage frequency. AREAS COVERED: The authors examine the role of mGluR5 in the pathophysiology of PD and the potential use of mGluR5 NAM as an adjuvant therapy together with a primary treatment with L-DOPA. Specifically, the authors look at the mavoglurant therapy and the evidence presented through preclinical and clinical trials. EXPERT OPINION: Interaction between mGluR5 NAM and L-DOPA is an area of interest in PD research as concomitant treatment results in the improvement of LID symptoms in humans, thus enhancing the patient's quality of life. However, few months ago, Novartis decided to discontinue clinical trials of mavoglurant for the treatment of LID, due to the lack of efficacy demonstrated in trials NCT01385592 and NCT01491529, although no safety concerns were involved in this decision. Nevertheless, the potential application of mGluR5 antagonists as neuroprotective agents must be considered and further studies are warranted to better investigate their potential.

Antiapoptotic drugs: a therapautic strategy for the prevention of neurodegenerative diseases.

The purpose of this review is to discuss potential pathways involved in the pathogenesis of neurodegenerative diseases, highlighting current pharmacological drug targets in neuronal apoptosis prevention. The incidence of these disorders is expected to rise in the coming years and so finding effective treatments represents a significant challenge for medicine. Alzheimer's disease and Parkinson's disease were both described almost a century ago and are the most important neurodegenerative disorders in the developed world. However, the molecular mechanisms that lead to the development of the neuronal pathology in both diseases are unclear. For this reason, despite substantial research in the area, an effective treatment for these diseases does not yet exist. In the present study we discuss in depth the pathways involved in apoptosis and neuronal death in neurodegenerative diseases. We also examine drugs that may have a neuroprotective effect. Inhibition of apoptosis mediated by oxidative stress generation and mitochondrial alteration or by the blockade of NMDA receptors could constitute a suitable therapeutic strategy for Alzheimer's disease. A multiple therapy with antioxidants, cell cycle inhibitors, GSK3ß inhibitors, and STATINS could, in the future, represent a suitable strategy for delaying the progression of neurodegenerative diseases. This research contributes to the development of new methods in the field of apoptosis inhibitors that could provide the future tools for the treatment of Alzheimer's and Parkinson's disease, as well as other neurodegenerative diseases.

Enhanced tau phosphorylation in the hippocampus of mice treated with 3,4-methylenedioxymethamphetamine ("Ecstasy").

3,4-Methylenedioxymethamphetamine (MDMA) ("Ecstasy") produces neurotoxic effects, which result into an impairment of learning and memory and other neurological dysfunctions. We examined whether MDMA induces increases in tau protein phosphorylation, which are typically associated with Alzheimer's disease and other chronic neurodegenerative disorders. We injected mice with MDMA at cumulative doses of 10-50 mg/kg intraperitoneally, which are approximately equivalent to doses generally consumed by humans. MDMA enhanced the formation of reactive oxygen species and induced reactive gliosis in the hippocampus, without histological evidence of neuronal loss. An acute or 6 d treatment with MDMA increased tau protein phosphorylation in the hippocampus, revealed by both anti-phospho(Ser(404))-tau and paired helical filament-1 antibodies. This increase was restricted to the CA2/CA3 subfields and lasted 1 and 7 d after acute and repeated MDMA treatment, respectively. Tau protein was phosphorylated as a result of two nonredundant mechanisms: (1) inhibition of the canonical Wnt (wingless-type MMTV integration site family) pathway, with ensuing activation of glycogen synthase kinase-3beta; and (2) activation of type-5 cyclin-dependent kinase (Cdk5). MDMA induced the expression of the Wnt antagonist, Dickkopf-1, and the expression of the Cdk5-activating protein, p25. In addition, the increase in tau phosphorylation was attenuated by strategies that rescued the Wnt pathway or inhibited Cdk5. Finally, an impairment in hippocampus-dependent spatial learning was induced by doses of MDMA that increased tau phosphorylation, although the impairment outlasted this biochemical event. We conclude that tau hyperphosphorylation in the hippocampus may contribute to the impairment of learning and memory associated with MDMA abuse.

Inhibition of ataxia telangiectasia-p53-E2F-1 pathway in neurons as a target for the prevention of neuronal apoptosis.

Over the last few decades, understanding of the mechanisms involved in the process of neuronal cell death has grown. Recent findings have established that DNA damage, and specifically ataxia telangiectasia mutated protein (ATM), is key to the cascade of regulation of neuronal apoptosis. Another characteristic common to all neurodegenerative diseases is oxidative stress. Likewise, a common feature in the brain of patients with neurodegenerative diseases such as Alzheimer's and Parkinson's diseases and other neurological disorders is the expression of proteins involved in cell-cycle control. In the process of re-entry in the cell cycle, an additional component, transcription factor E2F-1, also involved in the regulation of apoptosis, is expressed. Finally, in this complex puzzle, mitochondrial activation with the release of proteins and the activation of cystein proteases, specifically caspase-3, is prominent in the last step of neuronal apoptosis. This review focuses on the role of ATM activation and its re-entry into the cell cycle in neurons as a potential target for the prevention of neuronal apoptosis. We suggest the mechanisms by which ATM and E2F-1 orchestrate the apoptotic process. Among them, p53 could be a common point on this apoptotic route. Finally, we put forward drugs that are now being studied experimentally, such as p53 inhibitors, ATM inhibitors and cyclin-dependent kinase (CDKs) inhibitors, for the treatment of neurodegenerative diseases.

Increased permeability of blood-brain barrier on the hippocampus of a murine model of senescence.

SAMP8 mice show several indicative characteristics of accelerated aging and have been used to study the physiological and physiopathological processes that take place during senescence. There is some controversy about the presence of a functional blood-brain barrier (BBB) disturbance on these animals, which could be related to the oxidative stress or the amyloidosis present in their brain. In order to elucidate BBB status in the hippocampus of SAMP8 mice, in this study we have determined the extravasation from brain microvessels of endogenous IgG in SAMP8 mice aged 3, 7 and 12 months and in age-matched control SAMR1 mice. Immunohistochemistry, confocal microscopy and an imaging methodology specially designed to quantify IgG extravasation have been used. The choroid plexus was analyzed as a control for positive extravasation in SAMP8 and SAMR1 mice and, as expected, in all studied ages high IgG immunoreactivity was observed in both strains. We have found significantly higher levels of IgG extravasation in the hippocampus of 12-month-old SAMP8 mice compared to SAMR1 mice, indicating an increased permeability of BBB in aged senescence-accelerated mice.

The role of CDK5/P25 formation/inhibition in neurodegeneration.

Cdk5 is an atypical cyclin-dependent kinase localized in the brain, and its activity is dependent upon binding to p35/p39. In addition, while cdk5 has important physiological functions related to brain development, the breakdown of cdk5/p35 into cdk5/p25 increases its kinase activity and neurotoxicity. Interestingly, in recent years increased cdk5/p25 expression has been demonstrated in the brains of patients with Alzheimer's and Parkinson's diseases. Experimental studies performed in neuronal cell cultures indicate that cdk5/p25 plays a prominent role in apoptosis. Moreover, an apoptotic pathway, via an intracellular calcium increase following calpain activation and cdk5/p25 formation, has been postulated. Cdk5/p25 subsequently phosphorylates the nuclear transcription factor myocyte enhancer factor (MEF2), thereby inhibiting its prosurvival activity. However, cdk5/p25 could phosphorylate other substrates such as tau and p53, as well as the retinoblastoma protein pRb. All these data lend credence to the hypothesis that cdk5/p25 acts as a master regulator of neuronal cell death. In addition, cdk5/p25 might also interact with other pathways such as glycogen synthetase kinase 3beta (GSK3beta) and c-JUN kinase. Drugs like roscovitine, flavopiridol, calpain inhibitors, kenpaullone and induribins, which inhibit cdk5/p25 formation, constitute potential drugs for the treatment of neurological disorders. Furthermore, the dual inhibitory effect of some of these drugs on cdk5 and GSK3beta could be beneficial.

Changes in oxidative stress parameters and neurodegeneration markers in the brain of the senescence-accelerated mice SAMP-8.

The senescence-accelerated strains of mice (SAMP) are well-characterized animal models of senescence. Senescence may be related to enhanced production or defective control of reactive oxygen species, which lead to neuronal damage. Therefore, the activity of various oxidative-stress related enzymes was determined in the cortex of 5 months-old senescence-accelerated mice prone-8 (SAMP-8) of both sexes and compared with senescence-accelerated mice-resistant-1 (SAMR-1). Glutathione reductase and peroxidase activities in SAMP-8 male mice were lower than in male SAMR-1, and a decreased catalase activity was found in both male and female SAMP-8 mice, which correlates with the lower catalase expression found by Western blotting. Nissl staining showed marked loss of neuronal cells in the cerebral cortex of five month-old SAMP-8 mice. SAMP-8 mice also had marked astrogliosis and microgliosis. We also found an increase in caspase-3 and calpain activity in the cortex. In addition, we observed morphological changes in the immunostaining of tau protein in SAMP-8, indicative of a loss of their structural function. Altogether, these results show that, at as early as 5 months of age, SAMP-8 mice have cytological and molecular alterations indicative of neurodegeneration in the cerebral cortex and suggestive of altered control of the production of oxidative species and hyper-activation of calcium-dependent enzymes.

Inhibition of multiple pathways accounts for the antiapoptotic effects of flavopiridol on potassium withdrawal-induced apoptosis in neurons.

Serum and potassium (S/K) deprivation is a well-known apoptotic model in cerebellar granule neurons (CGNs), used to study the efficacy of potential neuroprotective drugs. The objective of this study was to determine the pathways involved in the neuroprotective role of flavopiridol, a pan-inhibitor of cyclin-dependent kinases (CDKs), upon S/K withdrawal-induced apoptosis in CGNs. Cell death in primary cultures of rat CGNs was accompanied by chromatin condensation and activation of caspases-3, -6, and -9. Caspase-3 activity was also evaluated by cleavage of 120-kDa alpha-spectrin. Flavopiridol (1 microM) prevented caspase activation and abolished apoptotic features mediated by S/K withdrawal. Re-entry in the cell cycle is also involved in apoptotic neuronal cell death. Flavopiridol (1 microM) inhibited DNA synthesis as measured by BrdU incorporation, thus enhancing proliferating cell nuclear antigen expression. Serum/potassium (S/K) deprivation induced apoptotic cell death mediated by the activation of several kinases such as glycogen synthase kinase-3beta and CDK5, as well as the breakdown of p35 in the neurotoxic fragment p25; inactivation of myocyte enhancer factor-2 (MEF2) was also found. Pretreatment with flavopiridol prevented these biochemical and molecular alterations. Taken together, these findings suggest an apoptotic route in CGNs after S/K withdrawal mediated by the activation of several kinases involved in cell cycle deregulation and MEF2 inactivation. We propose that the antiapoptotic properties of flavopiridol are mediated through kinase pathway inhibition.

p21(WAF1/Cip1) is not involved in kainic acid-induced apoptosis in murine cerebellar granule cells.

Kainic acid (KA) treatment induced neuronal death and apoptosis in murine cerebellar granule cells (CGNs) cultures from both wild-type and knockout p21(-/-) mice. There was not statistically significant difference in the percentage of neuronal apoptosis among strains. KA-induced neurotoxicity was prevented in the presence of NBQX (20 microM) and GYKI 52446 (20 microM), but not by z-VAD-fmk, suggesting that caspases are not involved in the apoptotic process. Data suggest that p21(WAF/Cip) was unable to modulate KA-induced apoptosis in murine CGNs.

Cyclosporin A enhances colchicine-induced apoptosis in rat cerebellar granule neurons.

1. Cyclosporin A (CsA, 1-50 microM), an immunosuppressive drug with known neurotoxic effects, did not decrease the viability of primary cultures of rat cerebellar granule neurons (CGN) or induce apoptotic features. However, CsA specifically enhanced the cytotoxicity and apoptosis induced by colchicine (1 microM). 2. Flavopiridol, an inhibitor of cyclin-dependent kinases (CDKs), prevented the neurotoxic effects of colchicine plus CsA. At 0.1-5 microM, it also showed antiapoptotic effects, as revealed by propidium iodide staining, flow cytometry and counting of cell nuclei. 3. Roscovitine (25-50 microM), a selective cdk1, 2 and 5 inhibitor, showed an antiapoptotic effect against colchicine- and colchicine plus CsA-induced apoptosis. 4. CsA increased the expression of cdk5 and cdk5/p25 mediated by colchicine, a CDK involved in neuronal apoptosis. After treatment of CGN with colchicine plus CsA, the changes in the p25/p35 ratio pointed to cdk5 activation. 5. Immunohistochemical results showed a nuclear localization of cdk5 after neurotoxic treatment, which was prevented by cdk inhibitors. Thus, we propose a new mechanism of modulation of CsA neurotoxicity mediated by cdk5.