Altered Kv2.1 functioning promotes increased excitability in hippocampal neurons of an Alzheimer's disease mouse model.

Altered neuronal excitability is emerging as an important feature in Alzheimer's disease (AD). Kv2.1 potassium channels are important modulators of neuronal excitability and synaptic activity. We investigated Kv2.1 currents and its relation to the intrinsic synaptic activity of hippocampal neurons from 3xTg-AD (triple transgenic mouse model of Alzheimer's disease) mice, a widely employed preclinical AD model. Synaptic activity was also investigated by analyzing spontaneous [Ca(2+)]i spikes. Compared with wild-type (Non-Tg (non-transgenic mouse model)) cultures, 3xTg-AD neurons showed enhanced spike frequency and decreased intensity. Compared with Non-Tg cultures, 3xTg-AD hippocampal neurons revealed reduced Kv2.1-dependent Ik current densities as well as normalized conductances. 3xTg-AD cultures also exhibited an overall decrease in the number of functional Kv2.1 channels. Immunofluorescence assay revealed an increase in Kv2.1 channel oligomerization, a condition associated with blockade of channel function. In Non-Tg neurons, pharmacological blockade of Kv2.1 channels reproduced the altered pattern found in the 3xTg-AD cultures. Moreover, compared with untreated sister cultures, pharmacological inhibition of Kv2.1 in 3xTg-AD neurons did not produce any significant modification in Ik current densities. Reactive oxygen species (ROS) promote Kv2.1 oligomerization, thereby acting as negative modulator of the channel activity. Glutamate receptor activation produced higher ROS levels in hippocampal 3xTg-AD cultures compared with Non-Tg neurons. Antioxidant treatment with N-Acetyl-Cysteine was found to rescue Kv2.1-dependent currents and decreased spontaneous hyperexcitability in 3xTg-AD neurons. Analogous results regarding spontaneous synaptic activity were observed in neuronal cultures treated with the antioxidant 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox). Our study indicates that AD-related mutations may promote enhanced ROS generation, oxidative-dependent oligomerization, and loss of function of Kv2.1 channels. These processes can be part on the increased neuronal excitability of these neurons. These steps may set a deleterious vicious circle that eventually helps to promote excitotoxic damage found in the AD brain

Inhibition of de novo ceramide biosynthesis affects aging phenotype in an in vitro model of neuronal senescence.

Although aging is considered to be an unavoidable event, recent experimental evidence suggests that the process can be counteracted. Intracellular calcium (Ca2+i) dyshomeostasis, mitochondrial dysfunction, oxidative stress, and lipid dysregulation are critical factors that contribute to senescence-related processes. Ceramides, a pleiotropic class of sphingolipids, are important mediators of cellular senescence, but their role in neuronal aging is still largely unexplored. In this study, we investigated the effects of L-cycloserine (L-CS), an inhibitor of thede novoceramide biosynthesis, on the aging phenotype of cortical neurons cultured for 22 days, a setting employed as anin vitromodel of senescence. Our findings indicate that, compared to control cultures, 'aged' neurons display dysregulation of [Ca2+]ilevels, mitochondrial dysfunction, increased generation of reactive oxygen species (ROS), altered synaptic activity as well as the activation of neuronal death-related molecules. Treatment with L-CS positively affected the senescent phenotype, a result associated with recovery of neuronal [Ca2+]isignaling and reduction of mitochondrial dysfunction and ROS generation. The results suggest that thede novoceramide biosynthesis represents a critical intermediate in the molecular and functional cascade leading to neuronal senescence and identify ceramide biosynthesis inhibitors as promising pharmacological tools to decrease age-related neuronal dysfunctions

Analysis of calcium homeostasis in fresh lymphocytes from patients with sporadic Alzheimer's disease or mild cognitive impairment

AbstractAlzheimer's disease (AD) is the most widespread, age-related neurodegenerative disorder. Its two subtypes are sporadic AD (SAD) of unknown etiology and genetically encoded familial AD (FAD). The onset of AD is often preceded by mild cognitive impairment (MCI). Calcium dynamics were found to be dysregulated in FAD models, but little is known about the features of calcium dynamics in SAD. To explore calcium homeostasis during the early stages of SAD, we investigated store-operated calcium entry (SOCE) and inositol triphosphate receptor (IP3R)-mediated calcium release into the cytoplasm in unmodified B lymphocytes from MCI and SAD patients and compared them with non-demented subjects (NDS). Calcium levels in the endoplasmic reticulum and both the rising and falling SOCE slopes were very similar in all three groups. However, we found that SAD and MCI cells were more prone to IP3R activation than NDS cells, and increases in calcium levels in the cytoplasm were almost twice as frequent in SAD cells than in NDS cells. MCI cells and SAD cells exhibited an enhanced magnitude of calcium influx during SOCE. MCI cells but not SAD cells were characterized by higher basal cellular calcium levels than NDS cells. In summary, perturbed calcium homeostasis was observed in peripheral cells from MCI and SAD patients. Thus, lymphocytes obtained from MCI subjects may be promising in the early diagnosis of individuals who will eventually develop SAD. However, no conclusions are made regarding SAD due to the limited number patients. This article is part of a Special Issue entitled: 12th European Symposium on Calcium

When, where and how? Focus on neuronal calcium dysfunctions in Alzheimer's Disease.

Alzheimer\u2019s disease (AD), since its characterization as a precise form of dementia with its own pathological hallmarks, has captured scientists\u2019 attention because of its complexity. The last 30 years have been filled with discoveries regarding the elusive aetiology of this disease and, thanks to advances in molecular biology and live imaging techniques, we now know that an important role is played by calcium (Ca2+). Ca2+, as ubiquitous second messenger, regulates a vast variety of cellular processes, from neuronal excitation and communication, to muscle fibre contraction and hormone secretion, with its action spanning a temporal scale that goes from microseconds to hours. It is therefore very challenging to conceive a single hypothesis that can integrate the numerous findings on this issue with those coming from the classical fields of AD research such as amyloid-beta (A) and tau pathology. In this contribution, we will focus our attention on the Ca2+ hypothesis of AD, dissecting it, as much as possible, in its subcellular localization, where the Ca2+ signal meets its specificity. We will also follow the temporal evolution of the Ca2+ hypothesis, providing some of the most updated discoveries. Whenever possible, we will link the findings regarding Ca2+ dysfunction to the other players involved in AD pathogenesis, hoping to provide a crossover body of evidence, useful to amplify the knowledge that will lead towards the discovery of an effective therapy

What Electrophysiology Tells Us About Alzheimer’s Disease::A Window into the Synchronization and Connectivity of Brain Neurons

Electrophysiology provides a real-time readout of neural functions and network capability in different brain states, on temporal (fractions of milliseconds) and spatial (micro, meso, and macro) scales unmet by other methodologies. However, current international guidelines do not endorse the use of electroencephalographic (EEG)/magnetoencephalographic (MEG) biomarkers in clinical trials performed in patients with Alzheimer’s disease (AD), despite a surge in recent validated evidence. This Position Paper of the ISTAART Electrophysiology Professional Interest Area endorses consolidated and translational electrophysiological techniques applied to both experimental animal models of AD and patients, to probe the effects of AD neuropathology (i.e., brain amyloidosis, tauopathy, and neurodegeneration) on neurophysiological mechanisms underpinning neural excitation/inhibition and neurotransmission as well as brain network dynamics, synchronization, and functional connectivity reflecting thalamocortical and cortico-cortical residual capacity. Converging evidence shows relationships between abnormalities in EEG/MEG markers and cognitive deficits in groups of AD patients at different disease stages. The supporting evidence for the application of electrophysiology in AD clinical research as well as drug discovery pathways warrants an international initiative to include the use of EEG/MEG biomarkers in the main multicentric projects planned in AD patients, to produce conclusive findings challenging the present regulatory requirements and guidelines for AD studies

Mechanisms of amyloid-beta cytotoxicity in hippocampal network function : rescue strategies in Alzheimer's disease

The origin and nature of cognitive processes are strongly associated with synchronous rhythmic activity in the brain. Gamma oscillations that span the frequency range of 30–80 Hz are particularly important for sensory perception, attention, learning, and memory. These oscillations occur intrinsically in brain regions, such as the hippocampus, that are directly linked to memory and disease. It has been reported that gamma and other rhythms are impaired in brain disorders such as Alzheimer’s disease, Parkinson’s disease, and schizophrenia; however, little is known about how these oscillations are affected. In the studies contained in this thesis, we investigated a possible involvement of toxic Amyloid-beta (Aβ) peptide associated with Alzheimer’s disease in degradation of gamma oscillations and the underlying cellular mechanismsin rodent hippocampi. We also aimed to prevent possible Aβ- induced effects by using specially designed molecular tools known to reduce toxicity associated with Aβ by interfering with its folding and aggregation steps. Using electrophysiological techniques to study thelocal field potentials and cellular properties in the CA3 region of the hippocampus, we found that Aβ in physiological concentrations acutely degrades pharmacologically- induced hippocampal gamma oscillations in vitro in a concentration- and time- dependent manner. The severity of degradation also increased with the amount of fibrillar Aβ present. We report that the underlying cause of degradation of gamma oscillations is Aβ-induced desynchronization of action potentials in pyramidal neurons and a shift in the equilibrium of excitatory-inhibitory synaptic transmission. Using specially designed molecular tools such as Aβ-binding ligands and molecular chaperones, we provide evidence that Aβ-induced effects on gamma oscillations, cellular firing, and synaptic dynamics can be prevented. We also show unpublished data on Aβ effects on parvalbumin-positive baskets cells or fast-spiking interneurons, in which Aβ causes an increase in firing rate during gamma oscillations. This is similar to what is observed in neighboring pyramidal neurons, suggesting a general mechanism behind the effect of Aβ. The studies in this thesis provide a correlative link between Aβ-induced effects on excitatory and inhibitory neurons in the hippocampus and extracellular gamma oscillations, and identify the Aβ aggregation state responsible for its toxicity. We demonstrate that strategies aimed at preventing peptide aggregation are able to prevent the toxic effects of Aβ on neurons and gamma oscillations. The studies have the potential to contribute to the design of future therapeutic interventions that are aimed at preserving neuronal oscillations in the brain to achieve cognitive benefits for patients

The Structural and Functional Connectome and Prediction of Risk for Cognitive Impairment in Older Adults

The human connectome refers to a comprehensive description of the brain's structural and functional connections in terms of brain networks. As the field of brain connectomics has developed, data acquisition, subsequent processing and modeling, and ultimately the representation of the connectome have become better defined and integrated with network science approaches. In this way, the human connectome has provided a way to elucidate key features of not only the healthy brain but also diseased brains. The field has quickly evolved, offering insights into network disruptions that are characteristic for specific neurodegenerative disorders. In this paper, we provide a brief review of the field of brain connectomics, as well as a more in-depth survey of recent studies that have provided new insights into brain network pathologies, including those found in Alzheimer's disease (AD), patients with mild cognitive impairment (MCI), and finally in people classified as being "at risk". Until the emergence of brain connectomics, most previous studies had assessed neurodegenerative diseases mainly by focusing on specific and dispersed locales in the brain. Connectomics-based approaches allow us to model the brain as a network, which allows for inferences about how dynamic changes in brain function would be affected in relation to structural changes. In fact, looking at diseases using network theory gives rise to new hypotheses on mechanisms of pathophysiology and clinical symptoms. Finally, we discuss the future of this field and how understanding both the functional and structural connectome can aid in gaining sharper insight into changes in biological brain networks associated with cognitive impairment and dementia

Dynamics of large-scale brain activity in health and disease

Tese de doutoramento em Engenharia Biomédica e Biofísica, apresentada à Universidade de Lisboa através da Faculdade de Ciências, 2008Cognition relies on the integration of information processed in widely distributed brain regions. Neuronal oscillations are thought to play an important role in the supporting local and global coordination of neuronal activity. This study aimed at investigating the dynamics of the ongoing healthy brain activity and early changes observed in patients with Alzheimer's disease (AD). Electro- and magnetoencephalography (EEG/MEG) were used due to high temporal resolution of these techniques. In order to evaluate the functional connectivity in AD, a novel algorithm based on the concept of generalized synchronization was improved by defining the embedding parameters as a function of the frequency content of interest. The time-frequency synchronization likelihood (TF SL) revealed a loss of fronto-temporal/parietal interactions in the lower alpha (8 10 Hz) oscillations measured by MEG that was not found with classical coherence. Further, long-range temporal (auto-) correlations (LRTC) in ongoing oscillations were assessed with detrended fluctuation analysis (DFA) on times scales from 1 25 seconds. Significant auto-correlations indicate a dependence of the underlying dynamical processes at certain time scales of separation, which may be viewed as a form of "physiological memory". We tested whether the DFA index could be related to the decline in cognitive memory in AD. Indeed, a significant decrease in the DFA exponents was observed in the alpha band (6 13 Hz) over temporo-parietal regions in the patients compared with the age-matched healthy control subjects. Finally, the mean level of SL of EEG signals was found to be significantly decreased in the AD patients in the beta (13 30 Hz) and in the upper alpha (10 13 Hz) and the DFA exponents computed as a measure of the temporal structure of SL time series were larger for the patients than for subjects with subjective memory complaint. The results obtained indicate that the study of spatio-temporal dynamics of resting-state EEG/MEG brain activity provides valuable information about the AD pathophysiology, which potentially could be developed into clinically useful indices for assessing progression of AD or response to medication