RyR-mediated calcium release in hippocampal health and disease.

Hippocampal synaptic plasticity is widely considered the cellular basis of learning and spatial memory processes. This article highlights the central role of Ca2+ release from the endoplasmic reticulum (ER) in hippocampal synaptic plasticity and hippocampus-dependent memory in health and disease. The key participation of ryanodine receptor (RyR) channels, which are the principal Ca2+ release channels expressed in the hippocampus, in these processes is emphasized. It is proposed that the increased neuronal oxidative tone displayed by hippocampal neurons during aging or Alzheimer's disease (AD) leads to excessive activation of RyR-mediated Ca2+ release, a process that is highly redox-sensitive, and that this abnormal response contributes to and aggravates these deleterious conditions.

Amyloid ß-Oligomers Inhibit the Nuclear Ca2+ Signals and the Neuroprotective Gene Expression Induced by Gabazine in Hippocampal Neurons.

Hippocampal neuronal activity generates dendritic and somatic Ca2+ signals, which, depending on stimulus intensity, rapidly propagate to the nucleus and induce the expression of transcription factors and genes with crucial roles in cognitive functions. Soluble amyloid-beta oligomers (AßOs), the main synaptotoxins engaged in the pathogenesis of Alzheimer's disease, generate aberrant Ca2+ signals in primary hippocampal neurons, increase their oxidative tone and disrupt structural plasticity. Here, we explored the effects of sub-lethal AßOs concentrations on activity-generated nuclear Ca2+ signals and on the Ca2+-dependent expression of neuroprotective genes. To induce neuronal activity, neuron-enriched primary hippocampal cultures were treated with the GABAA receptor blocker gabazine (GBZ), and nuclear Ca2+ signals were measured in AßOs-treated or control neurons transfected with a genetically encoded nuclear Ca2+ sensor. Incubation (6 h) with AßOs significantly reduced the nuclear Ca2+ signals and the enhanced phosphorylation of cyclic AMP response element-binding protein (CREB) induced by GBZ. Likewise, incubation (6 h) with AßOs significantly reduced the GBZ-induced increases in the mRNA levels of neuronal Per-Arnt-Sim domain protein 4 (Npas4), brain-derived neurotrophic factor (BDNF), ryanodine receptor type-2 (RyR2), and the antioxidant enzyme NADPH-quinone oxidoreductase (Nqo1). Based on these findings we propose that AßOs, by inhibiting the generation of activity-induced nuclear Ca2+ signals, disrupt key neuroprotective gene expression pathways required for hippocampal-dependent learning and memory processes.

Long-term potentiation and spatial memory training stimulate the hippocampal expression of RyR2 calcium release channels.

Introduction: Neuronal Ca2+ signals generated through the activation of Ca2+-induced Ca2+ release in response to activity-generated Ca2+ influx play a significant role in hippocampal synaptic plasticity, spatial learning, and memory. We and others have previously reported that diverse stimulation protocols, or different memory-inducing procedures, enhance the expression of endoplasmic reticulum-resident Ca2+ release channels in rat primary hippocampal neuronal cells or hippocampal tissue. Methods and Results: Here, we report that induction of long-term potentiation (LTP) by Theta burst stimulation protocols of the CA3-CA1 hippocampal synapse increased the mRNA and protein levels of type-2 Ryanodine Receptor (RyR2) Ca2+ release channels in rat hippocampal slices. Suppression of RyR channel activity (1 h preincubation with 20 µM ryanodine) abolished both LTP induction and the enhanced expression of these channels; it also promoted an increase in the surface expression of the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunits GluR1 and GluR2 and caused a moderate but significant reduction of dendritic spine density. In addition, training rats in the Morris water maze induced memory consolidation, which lasted for several days after the end of the training period, accompanied by an increase in the mRNA levels and the protein content of the RyR2 channel isoform. Discussion: We confirm in this work that LTP induction by TBS protocols requires functional RyR channels. We propose that the increments in the protein content of RyR2 Ca2+ release channels, induced by LTP or spatial memory training, play a significant role in hippocampal synaptic plasticity and spatial memory consolidation.

Ryanodine Receptor Mediated Calcium Release Contributes to Ferroptosis Induced in Primary Hippocampal Neurons by GPX4 Inhibition.

Ferroptosis, a newly described form of regulated cell death, is characterized by the iron-dependent accumulation of lipid peroxides, glutathione depletion, mitochondrial alterations, and enhanced lipoxygenase activity. Inhibition of glutathione peroxidase 4 (GPX4), a key intracellular antioxidant regulator, promotes ferroptosis in different cell types. Scant information is available on GPX4-induced ferroptosis in hippocampal neurons. Moreover, the role of calcium (Ca2+) signaling in ferroptosis remains elusive. Here, we report that RSL3, a selective inhibitor of GPX4, caused dendritic damage, lipid peroxidation, and induced cell death in rat primary hippocampal neurons. Previous incubation with the ferroptosis inhibitors deferoxamine or ferrostatin-1 reduced these effects. Likewise, preincubation with micromolar concentrations of ryanodine, which prevent Ca2+ release mediated by Ryanodine Receptor (RyR) channels, partially protected against RSL3-induced cell death. Incubation with RSL3 for 24 h suppressed the cytoplasmic Ca2+ concentration increase induced by the RyR agonist caffeine or by the SERCA inhibitor thapsigargin and reduced hippocampal RyR2 protein content. The present results add to the current understanding of ferroptosis-induced neuronal cell death in the hippocampus and provide new information both on the role of RyR-mediated Ca2+ signals on this process and on the effects of GPX4 inhibition on endoplasmic reticulum calcium content.

Visual-spatial processing impairment in the occipital-frontal connectivity network at early stages of Alzheimer's disease.

Introduction: Alzheimer's disease (AD) is the leading cause of dementia worldwide, but its pathophysiological phenomena are not fully elucidated. Many neurophysiological markers have been suggested to identify early cognitive impairments of AD. However, the diagnosis of this disease remains a challenge for specialists. In the present cross-sectional study, our objective was to evaluate the manifestations and mechanisms underlying visual-spatial deficits at the early stages of AD. Methods: We combined behavioral, electroencephalography (EEG), and eye movement recordings during the performance of a spatial navigation task (a virtual version of the Morris Water Maze adapted to humans). Participants (69-88 years old) with amnesic mild cognitive impairment-Clinical Dementia Rating scale (aMCI-CDR 0.5) were selected as probable early AD (eAD) by a neurologist specialized in dementia. All patients included in this study were evaluated at the CDR 0.5 stage but progressed to probable AD during clinical follow-up. An equal number of matching healthy controls (HCs) were evaluated while performing the navigation task. Data were collected at the Department of Neurology of the Clinical Hospital of the Universidad de Chile and the Department of Neuroscience of the Faculty of Universidad de Chile. Results: Participants with aMCI preceding AD (eAD) showed impaired spatial learning and their visual exploration differed from the control group. eAD group did not clearly prefer regions of interest that could guide solving the task, while controls did. The eAD group showed decreased visual occipital evoked potentials associated with eye fixations, recorded at occipital electrodes. They also showed an alteration of the spatial spread of activity to parietal and frontal regions at the end of the task. The control group presented marked occipital activity in the beta band (15-20 Hz) at early visual processing time. The eAD group showed a reduction in beta band functional connectivity in the prefrontal cortices reflecting poor planning of navigation strategies. Discussion: We found that EEG signals combined with visual-spatial navigation analysis, yielded early and specific features that may underlie the basis for understanding the loss of functional connectivity in AD. Still, our results are clinically promising for early diagnosis required to improve quality of life and decrease healthcare costs.

Periodontal bacteria in the brain-Implication for Alzheimer's disease: A systematic review.

Periodontitis is a chronic non-communicable disease caused by a dysbiotic microbiota. Pathogens can spread to the bloodstream, colonize other tissues or organs, and favor the onset of other pathologies, such as Alzheimer's disease (AD). Pathogens could permanently or transiently colonize the brain and induce an immune response. Thus, we analyzed the evidence combining oral bacteria's detection in the brain, both in animals and humans affected with AD. This systematic review was carried out following the PRISMA guideline. Studies that detected oral bacteria at the brain level were selected. The search was carried out in the Medline, Latindex, SciELO, and Cochrane Library databases. SYRCLE tool and Newcastle-Ottawa Scale were used for the risk of bias assessment. 23 studies were selected according to the eligibility criteria. Infection with oral pathogens in animals was related to developing neuropathological characteristics of AD and bacteria detection in the brain. In patients with AD, oral bacteria were detected in brain tissues, and increased levels of pro-inflammatory cytokines were also detected. There is evidence of a microbiological susceptibility to develop AD when the most dysbiosis-associated oral bacteria are present. The presence of bacteria in the brain is related to AD's pathological characteristics, suggesting an etiological oral-brain axis.

Hippocampal dendritic spines express the RyR3 but not the RyR2 ryanodine receptor isoform.

The hippocampus is a brain region implicated in synaptic plasticity and memory formation; both processes require neuronal Ca2+ signals generated by Ca2+ entry via plasma membrane Ca2+ channels and Ca2+ release from the endoplasmic reticulum (ER). Through Ca2+-induced Ca2+ release, the ER-resident ryanodine receptor (RyR) Ca2+ channels amplify and propagate Ca2+ entry signals, leading to activation of cytoplasmic and nuclear Ca2+-dependent signaling pathways required for synaptic plasticity and memory processes. Earlier reports have shown that mice and rat hippocampus expresses mainly the RyR2 isoform, with lower expression levels of the RyR3 isoform and almost undetectable levels of the RyR1 isoform; both the RyR2 and RyR3 isoforms have central roles in synaptic plasticity and hippocampal-dependent memory processes. Here, we describe that dendritic spines of rat primary hippocampal neurons express the RyR3 channel isoform, which is also expressed in the neuronal body and neurites. In contrast, the RyR2 isoform, which is widely expressed in the neuronal body and neurites of primary hippocampal neurons, is absent from the dendritic spines. We propose that this asymmetric distribution is of relevance for hippocampal neuronal function. We suggest that the RyR3 isoform amplifies activity-generated Ca2+ entry signals at postsynaptic dendritic spines, from where they propagate to the dendrite and activate primarily RyR2-mediated Ca2+ release, leading to Ca2+ signal propagation into the soma and the nucleus where they activate the expression of genes that mediate synaptic plasticity and memory.

Gingival crevicular fluid as Biomarker's source for alzheimer's disease / Fluido crevicular gingival como biomarcador de enfermedad de Alzheimer

Abstract Periodontitis is a low-grade inflammatory disease caused by a subgingival dysbiotic microbiota. Multiple studies have determined the higher prevalence of tooth loss and poor oral hygiene in patients with Alzheimer's disease (AD). However, the periodontal diagnosis, periodontal bacteria or mediators has not been measured to date. Aim: To determine the periodontal status, the pro-inflammatory mediators, Porphyromonas gingivalis load, and Apoliporpotein E (ApoE) in patients with AD. A complete dental examination was performed on 30 patients, and cognitive status was determined by the Montreal Cognitive Assessment (MoCA). Subgingival microbiota and GCF samples were then taken from all patients from the deepest sites. Total DNA was isolated from the microbiota samples for the quantification of the 16S ribosomal subunit. Pro-inflammatory mediators and ApoE were quantified from the gingival crevicular fluid (GCF). Patients with AD had periodontitis stage III-IV in 80%, a higher concentration of pro-inflammatory and ApoE mediators, and a higher P. gingivalis load compared to healthy subjects. The pro-inflammatory mediators, P. gingivalis load had a negative correlation with the MoCA test scores. Finally, a ROC curve was performed to assess the specificity and sensitivity of ApoE levels, detecting an area of 84.9%. In AD patients, we found a more severe periodontitis, a higher levels of pro-inflammatory mediators, and higher bacterial load. In addition, there is an increase in ApoE that allows to clearly determine patients with health, periodontitis and periodontitis and AD.

Resumen La periodontitis es una enfermedad crónica no transmisible que se caracteriza por generar una inflamación sistémica de bajo grado causada por una microbiota disbiótica subgingival. Múltiples estudios han determinado la mayor prevalencia de pérdida de dientes y mala higiene bucal en pacientes con enfermedad de Alzheimer (EA). Sin embargo, el diagnóstico periodontal, bacterias periodontales o mediadores pro-inflamatorio no se ha medido hasta la fecha. Determinar el estado periodontal, los mediadores pro-inflamatorios, la carga de Porphyromonas gingivalis y la apoliporpoteína E (ApoE) en pacientes con EA. Se realizó un examen odontológico completo en 30 pacientes y el estado cognitivo se determinó mediante la Evaluación Cognitiva de Montreal (MoCA). Luego, se tomaron muestras de microbiota subgingival y FCG de todos los pacientes de los sitios más profundos. Se aisló el DNA total de las muestras de microbiota para la cuantificación de la subunidad ribosómica 16S. Los mediadores pro-inflamatorios y la ApoE se cuantificaron a partir del líquido crevicular gingival (GCF). Los pacientes con EA tenían periodontitis en estadio III-IV en 80%, una mayor concentración de mediadores pro-inflamatorios y ApoE, y una mayor carga de P. gingivalis en comparación con los sujetos sanos. Los mediadores pro-inflamatorios y la carga de P. gingivalis tuvieron una correlación negativa con las puntuaciones de la prueba MoCA. Finalmente, se realizó una curva ROC para evaluar la especificidad y sensibilidad de los niveles de ApoE, detectando un área del 84,9%. En los pacientes con EA encontramos una periodontitis más severa, mayores niveles de mediadores pro-inflamatorios y mayor carga bacteriana. Además, un aumento de ApoE que permite determinar claramente a los pacientes con salud, periodontitis y periodontitis y EA.

RyR-mediated Ca2+ release elicited by neuronal activity induces nuclear Ca2+ signals, CREB phosphorylation, and Npas4/RyR2 expression.

The expression of several hippocampal genes implicated in learning and memory processes requires that Ca2+ signals generated in dendritic spines, dendrites, or the soma in response to neuronal stimulation reach the nucleus. The diffusion of Ca2+ in the cytoplasm is highly restricted, so neurons must use other mechanisms to propagate Ca2+ signals to the nucleus. Here, we present evidence showing that Ca2+ release mediated by the ryanodine receptor (RyR) channel type-2 isoform (RyR2) contributes to the generation of nuclear Ca2+ signals induced by gabazine (GBZ) addition, glutamate uncaging in the dendrites, or high-frequency field stimulation of primary hippocampal neurons. Additionally, GBZ treatment significantly increased cyclic adenosine monophosphate response element binding protein (CREB) phosphorylation-a key event in synaptic plasticity and hippocampal memory-and enhanced the expression of Neuronal Per Arnt Sim domain protein 4 (Npas4) and RyR2, two central regulators of these processes. Suppression of RyR-mediated Ca2+ release with ryanodine significantly reduced the increase in CREB phosphorylation and the enhanced Npas4 and RyR2 expression induced by GBZ. We propose that RyR-mediated Ca2+ release induced by neuronal activity, through its contribution to the sequential generation of nuclear Ca2+ signals, CREB phosphorylation, Npas4, and RyR2 up-regulation, plays a central role in hippocampal synaptic plasticity and memory processes.

The calcium-iron connection in ferroptosis-mediated neuronal death.

Iron, through its participation in oxidation/reduction processes, is essential for the physiological function of biological systems. In the brain, iron is involved in the development of normal cognitive functions, and its lack during development causes irreversible cognitive damage. Yet, deregulation of iron homeostasis provokes neuronal damage and death. Ferroptosis, a newly described iron-dependent cell death pathway, differs at the morphological, biochemical, and genetic levels from other cell death types. Ferroptosis is characterized by iron-mediated lipid peroxidation, depletion of the endogenous antioxidant glutathione and altered mitochondrial morphology. Although iron promotes the emergence of Ca2+ signals via activation of redox-sensitive Ca2+ channels, the role of Ca2+ signaling in ferroptosis has not been established. The early dysregulation of the cellular redox state observed in ferroptosis is likely to disturb Ca2+ homeostasis and signaling, facilitating ferroptotic neuronal death. This review presents an overview of the role of iron and ferroptosis in neuronal function, emphasizing the possible involvement of Ca2+ signaling in these processes. We propose, accordingly, that the iron-ferroptosis-Ca2+ association orchestrates the progression of cognitive dysfunctions and memory loss that occurs in neurodegenerative diseases. Therefore, to prevent iron dyshomeostasis and ferroptosis, we suggest the use of drugs that target the abnormal Ca2+ signaling caused by excessive iron levels as therapy for neurological disorders.

Oral-Gut-Brain Axis in Experimental Models of Periodontitis: Associating Gut Dysbiosis With Neurodegenerative Diseases.

Periodontitis is considered a non-communicable chronic disease caused by a dysbiotic microbiota, which generates a low-grade systemic inflammation that chronically damages the organism. Several studies have associated periodontitis with other chronic non-communicable diseases, such as cardiovascular or neurodegenerative diseases. Besides, the oral bacteria considered a keystone pathogen, Porphyromonas gingivalis, has been detected in the hippocampus and brain cortex. Likewise, gut microbiota dysbiosis triggers a low-grade systemic inflammation, which also favors the risk for both cardiovascular and neurodegenerative diseases. Recently, the existence of an axis of Oral-Gut communication has been proposed, whose possible involvement in the development of neurodegenerative diseases has not been uncovered yet. The present review aims to compile evidence that the dysbiosis of the oral microbiota triggers changes in the gut microbiota, which creates a higher predisposition for the development of neuroinflammatory or neurodegenerative diseases.The Oral-Gut-Brain axis could be defined based on anatomical communications, where the mouth and the intestine are in constant communication. The oral-brain axis is mainly established from the trigeminal nerve and the gut-brain axis from the vagus nerve. The oral-gut communication is defined from an anatomical relation and the constant swallowing of oral bacteria. The gut-brain communication is more complex and due to bacteria-cells, immune and nervous system interactions. Thus, the gut-brain and oral-brain axis are in a bi-directional relationship. Through the qualitative analysis of the selected papers, we conclude that experimental periodontitis could produce both neurodegenerative pathologies and intestinal dysbiosis, and that periodontitis is likely to induce both conditions simultaneously. The severity of the neurodegenerative disease could depend, at least in part, on the effects of periodontitis in the gut microbiota, which could strengthen the immune response and create an injurious inflammatory and dysbiotic cycle. Thus, dementias would have their onset in dysbiotic phenomena that affect the oral cavity or the intestine. The selected studies allow us to speculate that oral-gut-brain communication exists, and bacteria probably get to the brain via trigeminal and vagus nerves.

Alzheimer's Disease-Like Pathology Triggered by Porphyromonas gingivalis in Wild Type Rats Is Serotype Dependent.

Periodontal disease is a disease of tooth-supporting tissues. It is a chronic disease with inflammatory nature and infectious etiology produced by a dysbiotic subgingival microbiota that colonizes the gingivodental sulcus. Among several periodontal bacteria, Porphyromonas gingivalis (P. gingivalis) highlights as a keystone pathogen. Previous reports have implied that chronic inflammatory response and measurable bone resorption are observed in young mice, even after a short period of periodontal infection with P. gingivalis, which has been considered as a suitable model of experimental periodontitis. Also, encapsulated P. gingivalis strains are more virulent than capsular-defective mutants, causing an increased immune response, augmented osteoclastic activity, and accrued alveolar bone resorption in these rodent experimental models of periodontitis. Recently, P. gingivalis has been associated with Alzheimer's disease (AD) pathogenesis, either by worsening brain pathology in AD-transgenic mice or by inducing memory impairment and age-dependent neuroinflammation middle-aged wild type animals. We hypothesized here that the more virulent encapsulated P. gingivalis strains could trigger the appearance of brain AD-markers, neuroinflammation, and cognitive decline even in young rats subjected to a short periodontal infection exposure, due to their higher capacity of activating brain inflammatory responses. To test this hypothesis, we periodontally inoculated 4-week-old male Sprague-Dawley rats with K1, K2, or K4 P. gingivalis serotypes and the K1-isogenic non-encapsulated mutant (GPA), used as a control. 45-days after periodontal inoculations with P. gingivalis serotypes, rat´s spatial memory was evaluated for six consecutive days in the Oasis maze task. Following functional testing, the animals were sacrificed, and various tissues were removed to analyze alveolar bone resorption, cytokine production, and detect AD-specific biomarkers. Strikingly, only K1 or K2 P. gingivalis-infected rats displayed memory deficits, increased alveolar bone resorption, pro-inflammatory cytokine production, changes in astrocytic morphology, increased Aß1-42 levels, and Tau hyperphosphorylation in the hippocampus. None of these effects were observed in rats infected with the non-encapsulated bacterial strains. Based on these results, we propose that the bacterial virulence factors constituted by capsular polysaccharides play a central role in activating innate immunity and inflammation in the AD-like pathology triggered by P. gingivalis in young rats subjected to an acute experimental infection episode.

Nicotinamide, a Poly [ADP-Ribose] Polymerase 1 (PARP-1) Inhibitor, as an Adjunctive Therapy for the Treatment of Alzheimer's Disease.

Nicotinamide (vitamin B3) is a key component in the cellular production of Nicotinamide Adenine Dinucleotide (NAD) and has long been associated with neuronal development, survival and death. Numerous data suggest that nicotinamide may offer therapeutic benefits in neurodegenerative disorders, including Alzheimer's Disease (AD). Beyond its effect in NAD+ stores, nicotinamide is an inhibitor of Poly [ADP-ribose] polymerase 1 (PARP-1), an enzyme with multiple cellular functions, including regulation of cell death, energy/metabolism and inflammatory response. PARP-1 functions as a DNA repair enzyme but under intense DNA damage depletes the cell of NAD+ and ATP and leads to a non-apoptotic type of cell death called Parthanatos, which has been associated with the pathogenesis of neurodegenerative diseases. Moreover, NAD+ availability might potentially improve mitochondrial function, which is severely impaired in AD. PARP-1 inhibition may also exert a protective effect against neurodegeneration by its action to diminish neuroinflammation and microglial activation which are also implicated in the pathogenesis of AD. Here we discuss the evidence supporting the use of nicotinamide as adjunctive therapy for the treatment of early stages of AD based on the inhibitory effect of nicotinamide on PARP-1 activity. The data support evaluating nicotinamide as an adjunctive treatment for AD at early stages of the disease not only to increase NAD+ stores but as a PARP-1 inhibitor, raising the hypothesis that other PARP-1 inhibitors - drugs that are already approved for breast cancer treatment - might be explored for the treatment of AD.

Astaxanthin Counteracts Excitotoxicity and Reduces the Ensuing Increases in Calcium Levels and Mitochondrial Reactive Oxygen Species Generation.

Astaxanthin (ASX) is a carotenoid pigment with strong antioxidant properties. We have reported previously that ASX protects neurons from the noxious effects of amyloid-ß peptide oligomers, which promote excessive mitochondrial reactive oxygen species (mROS) production and induce a sustained increase in cytoplasmic Ca2+ concentration. These properties make ASX a promising therapeutic agent against pathological conditions that entail oxidative and Ca2+ dysregulation. Here, we studied whether ASX protects neurons from N-methyl-D-aspartate (NMDA)-induced excitotoxicity, a noxious process which decreases cellular viability, alters gene expression and promotes excessive mROS production. Incubation of the neuronal cell line SH-SY5Y with NMDA decreased cellular viability and increased mitochondrial superoxide production; pre-incubation with ASX prevented these effects. Additionally, incubation of SH-SY5Y cells with ASX effectively reduced the basal mROS production and prevented hydrogen peroxide-induced cell death. In primary hippocampal neurons, transfected with a genetically encoded cytoplasmic Ca2+ sensor, ASX also prevented the increase in intracellular Ca2+ concentration induced by NMDA. We suggest that, by preventing the noxious mROS and Ca2+ increases that occur under excitotoxic conditions, ASX could be useful as a therapeutic agent in neurodegenerative pathologies that involve alterations in Ca2+ homeostasis and ROS generation.

Redox modifications in synaptic components as biomarkers of cognitive status, in brain aging and disease.

Aging is a natural process that includes several changes that gradually make organisms degenerate and die. Harman's theory proposes that aging is a consequence of the progressive accumulation of oxidative modifications mediated by reactive oxygen/nitrogen species, which plays an essential role in the development and progression of many neurodegenerative diseases. This review will focus on how abnormal redox modifications induced by age impair the functionality of neuronal redox-sensitive proteins involved in axonal elongation and guidance, synaptic plasticity, and intercellular communication. We will discuss post-transcriptional regulation of gene expression by microRNAs as a mechanism that controls the neuronal redox state. Finally, we will discuss how some brain-permeant antioxidants from the diet have a beneficial effect on cognition. Taken together, the evidence revised here indicates that oxidative-driven modifications of specific proteins and changes in microRNA expression may be useful biomarkers for aging and neurodegenerative diseases. Also, some specific antioxidant therapies have undoubtedly beneficial neuroprotective effects when administered in the correct doses, in the ideal formulation combination, and during the appropriate therapeutic window. The use of some antioxidants is, therefore, still poorly explored for the treatment of neurodegenerative diseases such as Alzheimer's disease.

Differential navigational strategies during spatial learning in a new modified version of the Oasis maze.

During spatial navigation, some typical parameters of learning have been observed, such as latency or path length. However, these parameters are sensitive to patterns of navigation and orientation that are not easily measurable. In the present study, we used a modified version of the Oasis maze and evaluated different parameters of learning, navigation, and orientation in different animal groups. Through a PCA (Principal component analysis) we found different factors such as learning, navigation, speediness, anxiety, orientation, path variability, and turning behavior. Each factor gathers different groups of behavioral variables. ANOVA analysis of those factors demonstrates that some of them are more strongly modulated by trial progression, while others by animal group differences, indicating that each group of variables is better reflecting one of these dimensions. To understand the nature of these navigation differences, we studied orientation strategies between animal conditions and across trials. We found that the main navigational strategy used by the animals consist of locating the target and directing their behaviors towards this area. When testing how this strategy changed after cognitive impairment or enhancement, we found that AßOs treated animals (Amyloid ß Oligomers, Alzheimer animal model) have strong orientation difficulties at locating the target at longer distances. While animals with learning enhancement (exercised rat) do not show changes in orientation behaviors. These analyses highlight that experimental manipulations affect learning, but also induced changes in the navigational strategies. We concluded that both dimensions can explain the differences observed in typical learning variables, such as latency or path length, motivating the development of new tools that asses this two-dimension as a separate but, interacting phenomenon.

N-Acetylcysteine Prevents the Spatial Memory Deficits and the Redox-Dependent RyR2 Decrease Displayed by an Alzheimer's Disease Rat Model.

We have previously reported that primary hippocampal neurons exposed to synaptotoxic amyloid beta oligomers (AßOs), which are likely causative agents of Alzheimer's disease (AD), exhibit abnormal Ca2+ signals, mitochondrial dysfunction and defective structural plasticity. Additionally, AßOs-exposed neurons exhibit a decrease in the protein content of type-2 ryanodine receptor (RyR2) Ca2+ channels, which exert critical roles in hippocampal synaptic plasticity and spatial memory processes. The antioxidant N-acetylcysteine (NAC) prevents these deleterious effects of AßOs in vitro. The main contribution of the present work is to show that AßOs injections directly into the hippocampus, by engaging oxidation-mediated reversible pathways significantly decreased RyR2 protein content but increased single RyR2 channel activation by Ca2+ and caused considerable spatial memory deficits. AßOs injections into the CA3 hippocampal region impaired rat performance in the Oasis maze spatial memory task, decreased hippocampal glutathione levels and overall content of plasticity-related proteins (c-Fos, Arc, and RyR2) and increased ERK1/2 phosphorylation. In contrast, in hippocampus-derived mitochondria-associated membranes (MAM) AßOs injections increased RyR2 levels. Rats fed with NAC for 3-weeks prior to AßOs injections displayed comparable redox potential, RyR2 and Arc protein contents, similar ERK1/2 phosphorylation and RyR2 single channel activation by Ca2+ as saline-injected (control) rats. NAC-fed rats subsequently injected with AßOs displayed the same behavior in the spatial memory task as control rats. Based on the present in vivo results, we propose that redox-sensitive neuronal RyR2 channels partake in the mechanism underlying AßOs-induced memory disruption in rodents.

The signaling pathways underlying BDNF-induced Nrf2 hippocampal nuclear translocation involve ROS, RyR-Mediated Ca2+ signals, ERK and PI3K.

The neurotrophin Brain-Derived Neurotrophic Factor (BDNF) induces complex neuronal signaling cascades that are critical for the cellular changes underlying synaptic plasticity. These pathways include activation of Ca2+ entry via N-methyl-D-aspartate receptors and sequential activation of nitric oxide synthase and NADPH oxidase, which via generation of reactive nitrogen/oxygen species stimulate Ca2+-induced Ca2+ release mediated by Ryanodine Receptor (RyR) channels. These sequential events underlie BDNF-induced spine remodeling and type-2 RyR up-regulation. In addition, BDNF induces the nuclear translocation of the transcription factor Nrf2, a master regulator of antioxidant protein expression that protects cells against the oxidative damage caused by injury and inflammation. To investigate the possible BDNF-induced signaling cascades that mediate Nrf2 nuclear translocation in primary hippocampal cultures, we tested here whether reactive oxygen species, RyR-mediated Ca2+ release, ERK or PI3K contribute to this response. We found that pre-incubation of cultures with inhibitory ryanodine to suppress RyR-mediated Ca2+ release, with the reducing agent N-acetylcysteine or with inhibitors of ERK or PI3K activity, prevented the nuclear translocation of Nrf2 induced by incubation for 6 h with BFNF. Based on these combined results, we propose that the key role played by BDNF as an inducer of neuronal antioxidant responses, characterized by BDNF-induced Nfr2 nuclear translocation, entails crosstalk between reactive oxygen species and RyR-mediated Ca2+ release, and the participation of ERK and PI3K activities.

Reactive oxygen species released from astrocytes treated with amyloid beta oligomers elicit neuronal calcium signals that decrease phospho-Ser727-STAT3 nuclear content.

The transcription factor STAT3 has a crucial role in the development and maintenance of the nervous system. In this work, we treated astrocytes with oligomers of the amyloid beta peptide (AßOs), which display potent synaptotoxic activity, and studied the effects of mediators released by AßOs-treated astrocytes on the nuclear location of neuronal serine-727-phosphorylated STAT3 (pSerSTAT3). Treatment of mixed neuron-astrocyte cultures with 0.5µMAßOs induced in neurons a significant decrease of nuclear pSerSTAT3, but not of phosphotyrosine-705 STAT3, the other form of STAT3 phosphorylation. This decrease did not occur in astrocyte-poor neuronal cultures revealing a pivotal role for astrocytes in this response. To test if mediators released by astrocytes in response to AßOs induce pSerSTAT3 nuclear depletion, we used conditioned medium derived from AßOs-treated astrocyte cultures. Treatment of astrocyte-poor neuronal cultures with this medium caused pSerSTAT3 nuclear depletion but did not modify overall STAT3 levels. Extracellular catalase prevented the pSerSTAT3 nuclear depletion caused by astrocyte-conditioned medium, indicating that reactive oxygen species (ROS) mediate this response. This conditioned medium also increased neuronal oxidative tone, leading to a ryanodine-sensitive intracellular calcium signal that proved to be essential for pSerSTAT3 nuclear depletion. In addition, this depletion decreased BCL2 and Survivin transcription and significantly increased BAX/BCL2 ratio. This is the first description that ROS generated by AßOs-treated astrocytes and neuronal calcium signals jointly regulate pSerSTAT3 nuclear distribution in neurons. We propose that astrocytes release ROS in response to AßOs, which by increasing neuronal oxidative tone, generate calcium signals that cause pSerSTAT3 nuclear depletion and loss of STAT3 protective transcriptional activity.