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Amyloid beta (Aβ)-mediated synapse dysfunction and spine loss are considered to be early events in Alzheimer’s disease (AD) pathogenesis. N-methyl-D-aspartate receptors (NMDARs) have previously been suggested to play a role for Amyloid beta (Aβ) toxicity. Pharmacological block of NMDAR subunits in cultured neurons and mice suggested that NMDARs containing the GluN2B subunit are necessary for Aβ-mediated changes in synapse number and function in hippocampal neurons. Interestingly, NMDARs undergo a developmental switch from GluN2B- to GluN2A-containing receptors. This indicates different functional roles of NMDARs in young mice compared to older animals. In addition, the lack of pharmacological tools to efficiently dissect the role of NMDARs containing the different subunits complicates the interpretation of their specific role. In order to address this problem and to investigate the specific role for Aβ toxicity of the distinct NMDAR subunits in dentate gyrus granule cells of adult mice, we used conditional knockout mouse lines for the subunits GluN1, GluN2A and GluN2B. Aβ-mediated changes in synaptic function and neuronal anatomy were investigated in several-months old mice with virus-mediated overproduction of Aβ and in 1-year old 5xFAD mice. We found that all three NMDAR subunits contribute to the Aβ-mediated decrease in the number of functional synapses. However, NMDARs are not required for the spine number reduction in dentate gyrus granule cells after chronic Aβ-overproduction in 5xFAD mice. Furthermore, the amplitude of synaptic and extrasynaptic NMDAR-mediated currents was reduced in dentate gyrus granule of 5xFAD mice without changes in current kinetics, suggesting that a redistribution or change in subunit composition of NMDARs does not play a role in mediating Amyloid beta (Aβ) toxicity. Our study indicates that NMDARs are involved in AD pathogenesis by compromising synapse function but not by affecting neuron morphology. Electronic supplementary material The online version of this article (10.1186/s40478-018-0611-4) contains supplementary material, which is available to authorized users.

Alzheimer’s disease (AD) is an irreversible, devastating neurodegenerative brain disorder characterized by the loss of neurons and subsequent cognitive decline. Despite considerable progress in the understanding of the pathophysiology of AD, the precise molecular mechanisms that cause the disease remain elusive. By now, there is ample evidence that activated microglia have a critical role in the initiation and progression of AD. The present study describes the identification of Glycoprotein nonmetastatic melanoma protein B (GPNMB) as a novel AD-related factor in both transgenic mice and sporadic AD patients by expression profiling, immunohistochemistry and ELISA measurements. We show that GPNMB levels increase in an age-dependent manner in transgenic AD models showing profound cerebral neuron loss and demonstrate that GPNMB co-localizes with a distinct population of IBA1-positive microglia cells that cluster around amyloid plaques. Our data further indicate that GPNMB is part of a microglia activation state that is only present under neurodegenerative conditions and that is characterized by the up-regulation of a subset of genes including TREM2, APOE and CST7. In agreement, we provide in vitro evidence that soluble Aβ has a direct effect on GPNMB expression in an immortalized microglia cell line. Importantly, we show for the first time that GPNMB is elevated in brain samples and cerebrospinal fluid (CSF) of sporadic AD patients when compared to non-demented controls. The current findings indicate that GPNMB represents a novel disease-associated marker that appears to play a role in the neuroinflammatory response of AD. Electronic supplementary material The online version of this article (10.1186/s40478-018-0612-3) contains supplementary material, which is available to authorized users.

The molecular events causing memory loss and neuronal cell death in Alzheimer’s disease (AD) over time are still unknown. Here we found that picomolar concentrations of soluble oligomers of synthetic beta amyloid (Aβ42) aggregates incubated with BV2 cells or rat astrocytes caused a sensitised response of Toll-like receptor 4 (TLR4) with time, leading to increased production of TNF-α. Aβ aggregates caused long term potentiation (LTP) deficit in hippocampal slices and predominantly neuronal cell death in co-cultures of astrocytes and neurons, which was blocked by TLR4 antagonists. Soluble Aβ aggregates cause LTP deficit and neuronal death via an autocrine/paracrine mechanism due to TLR4 signalling. These findings suggest that the TLR4-mediated inflammatory response may be a key pathophysiological process in AD., Hughes et al. investigate the TLR4-mediated inflammatory response in Alzheimer’s disease and show that picomolar concentrations of soluble amyloid beta aggregates lead to a sensitised response of TLR4 with time, resulting in increased TNF-α production. They suggest the use of near physiological concentration of soluble aggregates can cause long-term potentiation deficit and neuronal death through an autocrine/paracrine mechanism due to TLR4 signalling.

Amyloid-β peptide (Aβ) forms plaques in Alzheimer’s disease (AD) and is responsible for early cognitive deficits in AD patients. Advancing cognitive decline is accompanied by progressive impairment of cognition-relevant EEG patterns such as gamma oscillations. The endocannabinoid anandamide, a TrpV1-receptor agonist, reverses hippocampal damage and memory impairment in rodents and protects neurons from Aβ-induced cytotoxic effects. Here, we investigate a restorative role of TrpV1-receptor activation against Aβ-induced degradation of hippocampal neuron function and gamma oscillations. We found that the TrpV1-receptor agonist capsaicin rescues Aβ-induced degradation of hippocampal gamma oscillations by reversing both the desynchronization of AP firing in CA3 pyramidal cells and the shift in excitatory/inhibitory current balance. This rescue effect is TrpV1-receptor-dependent since it was absent in TrpV1 knockout mice or in the presence of the TrpV1-receptor antagonist capsazepine. Our findings provide novel insight into the network mechanisms underlying cognitive decline in AD and suggest TrpV1 activation as a novel therapeutic target.

Soluble β -amyloid peptides (A β) and small Aβ oligomers represent the most toxic peptide moieties recognized in brains affected by Alzheimer’s disease (AD). Here we provide the first evidence that specific St. John’s wort (SJW) extracts both attenuate A β-induced histopathology and alleviate memory impairments in APP-transgenic mice. Importantly, these effects are attained independently of hyperforin. Specifically, two extracts characterized by low hyperforin content (i) significantly decrease intracerebral Aβ 42 levels, (ii) decrease the number and size of amyloid plaques, (iii) rescue neocortical neurons, (iv) restore cognition to normal levels, and (iv) activate microglia in vitro and in vivo. Mechanistically, we reveal that the reduction of soluble Aβ 42 species is the consequence of a highly increased export activity in the bloodbrain barrier ABCC1transporter, which was found to play a fundamental role in Aβ excretion into the bloodstream. These data (i) support the significant beneficial potential of SJW extracts on AD proteopathy, and (ii) demonstrate for the first time that hyperforin concentration does not necessarily correlate with their therapeutic effects. Hence, by activating ABC transporters, specific extracts of SJW may be used to treat AD and other diseases involving peptide accumulation and cognition impairment. We propose that the anti-depressant and anti-dementia effects of these hyperforin-reduced phytoextracts could be combined for treatment of the elderly, with a concomitant reduction in deleterious hyperforin-related side effects.

A fascinating yet perhaps overlooked trait of the p75 neurotrophin receptor (p75(NTR)) is its ability to bind ligands with no obvious neurotrophic function. Using cultured basal forebrain (BF) neurons, this study demonstrates selective internalization of amyloid β (Aβ) 1-42 in conjunction with p75(NTR) (labelled with IgG192-Cy3) by cholinergic cells. Active under resting conditions, this process was enhanced by high K(+) stimulation and was insensitive to inhibitors of regulated synaptic activity-tetrodotoxin or botulinum neurotoxins (BoNT type/A and/B). Blockade of sarco-endoplasmic reticulum (SERCA) Ca(2+) ATPase with thapsigargin and CPA or chelation of Ca(2+) with EGTA-AM strongly suppressed the endocytosis of p75(NTR), implicating the role of ER released Ca(2+). The uptake of IgG192-Cy3 was also reduced by T-type Ca(2+) channel blocker mibefradil but not Cd(2+), an indiscriminate blocker of high voltage-activated Ca(2+) currents. A strong co-localization of IgG192-Cy3 with late endosome (Rab7) or lysosome (Lamp1) qualifier proteins suggest these compartments as the primary destination for internalized IgG192 and Aβ. Selective uptake and labeling of BF cholinergic cells with IgG192-Cy3 injected into the prefrontal cortex was verified also in vivo. The significance of these findings in relation to Aβ clearance in the cerebral cortex and pathophysiology of Alzheimer's disease is discussed.

The deposition of the β-amyloid (Aβ) peptide in senile plaques and cerebral Aβ-amyloid angiopathy can be seeded in β-amyloid precursor protein (APP)-transgenic mice by the intracerebral infusion of brain extracts containing aggregated Aβ. Previous studies of seeded β-amyloid induction have used relatively short incubation periods to dissociate seeded β-amyloid induction from endogenous β-amyloid deposition of the host, thus precluding the analysis of the impact of age and extended incubation periods on the instigation and spread of Aβ lesions in brain. In the present study using R1.40 APP-transgenic mice (which do not develop endogenous Aβ deposition up to 15 months of age) we show that: (i) seeding at 9 months of age does not induce more Aβ deposition than seeding at 3 months of age, provided that the incubation period (6 months) is the same; and (ii) very long-term (12 months) incubation after a focal application of the seed results in the emergence of Aβ deposits throughout the forebrain. These findings indicate that the presence of Aβ seeds, and not the age of the host per se, is critical to the initiation of Aβ aggregation in the brain, and that Aβ deposition, actuated in one brain area, eventually spreads throughout the brain.

Aggregation of amyloid-β (Aβ) and Tau protein are hallmarks of Alzheimer's disease (AD), and according to the Aβ-cascade hypothesis, Aβ is considered toxic for neurons and Tau a downstream target of Aβ. We have investigated differentiated primary hippocampal neurons for early localized changes following exposure to Aβ oligomers. Initial events become evident by missorting of endogenous Tau into the somatodendritic compartment, in contrast to axonal sorting in normal neurons. In missorted dendritic regions there is a depletion of spines and local increase in Ca2+, and breakdown of microtubules. Tau in these regions shows elevated phosphorylation at certain sites diagnostic of AD-Tau (e.g., epitope of antibody 12E8, whose phosphorylation causes detachment of Tau from microtubules, and AT8 epitope), and local elevation of certain kinase activities (e.g., MARK/par-1, BRSK/SADK, p70S6K, cdk5, but not GSK3β, JNK, MAPK). These local effects occur without global changes in Tau, tubulin, or kinase levels. Somatodendritic missorting occurs not only with Tau, but also with other axonal proteins such as neurofilaments, and correlates with pronounced depletion of microtubules and mitochondria. The Aβ-induced effects on microtubule and mitochondria depletion, Tau missorting, and loss of spines are prevented by taxol, indicating that Aβ-induced microtubule destabilization and corresponding traffic defects are key factors in incipient degeneration. By contrast, the rise in Ca2+levels, kinase activities, and Tau phosphorylation cannot be prevented by taxol. Incipient and local changes similar to those of Aβ oligomers can be evoked by cell stressors (e.g., H2O2, glutamate, serum deprivation), suggesting some common mechanism of signaling.

Introduction The self-assembly of Aβ peptides into a range of conformationally heterogeneous amyloid states represents a fundamental event in Alzheimer’s disease. Within these structures oligomeric intermediates are considered to be particularly pathogenic. To test this hypothesis we have used a conformational targeting approach where particular conformational states, such as oligomers or fibrils, are recognized in vivo by state-specific antibody fragments. Results We show that oligomer targeting with the KW1 antibody fragment, but not fibril targeting with the B10 antibody fragment, affects toxicity in Aβ-expressing Drosophila melanogaster. The effect of KW1 is observed to occur selectively with flies expressing Aβ(1–40) and not with those expressing Aβ(1–42) or the arctic variant of Aβ(1–42) This finding is consistent with the binding preference of KW1 for Aβ(1–40) oligomers that has been established in vitro. Strikingly, and in contrast to the previously demonstrated in vitro ability of this antibody fragment to block oligomeric toxicity in long-term potentiation measurements, KW1 promotes toxicity in the flies rather than preventing it. This result shows the crucial importance of the environment in determining the influence of antibody binding on the nature and consequences of the protein misfolding and aggregation. Conclusions While our data support to the pathological relevance of oligomers, they highlight the issues to be addressed when developing inhibitory strategies that aim to neutralize these states by means of antagonistic binding agents.

Alzheimer’s disease (AD) is characterized by neurotoxic amyloid-ß plaque formation in brain parenchyma and cerebral blood vessels known as cerebral amyloid angiopathy (CAA). Besides CAA, AD is strongly related to vascular diseases such as stroke and atherosclerosis. Cerebrovascular dysfunction occurs in AD patients leading to alterations in blood flow that might play an important role in AD pathology with neuronal loss and memory deficits. Platelets are the major players in hemostasis and thrombosis, but are also involved in neuroinflammatory diseases like AD. For many years, platelets were accepted as peripheral model to study the pathophysiology of AD because platelets display the enzymatic activities to generate amyloid-ß (Aß) peptides. In addition, platelets are considered to be a biomarker for early diagnosis of AD. Effects of Aß peptides on platelets and the impact of platelets in the progression of AD remained, however, ill-defined. The present study explored the cellular mechanisms triggered by Aß in platelets. Treatment of platelets with Aß led to platelet activation and enhanced generation of reactive oxygen species (ROS) and membrane scrambling, suggesting enhanced platelet apoptosis. More important, platelets modulate soluble Aß into fibrillar structures that were absorbed by apoptotic but not vital platelets. This together with enhanced platelet adhesion under flow ex vivo and in vivo and platelet accumulation at amyloid deposits of cerebral vessels of AD transgenic mice suggested that platelets are major contributors of CAA inducing platelet thrombus formation at vascular amyloid plaques leading to vessel occlusion critical for cerebrovascular events like stroke.

Cerebral β-amyloidosis can be exogenously induced by the intracerebral injection of brain extracts containing aggregated β-amyloid (Aβ) into young, pre-depositing Aβ precursor protein- (APP) transgenic mice. Previous work has shown that the induction involves a prion-like seeding mechanism in which the seeding agent is aggregated Aβ itself. Here we report that the β-amyloid-inducing activity of Alzheimer's disease (AD) brain tissue or aged APP-transgenic mouse brain tissue is preserved, albeit with reduced efficacy, after formaldehyde fixation. Moreover, spectral analysis with amyloid conformation-sensitive luminescent conjugated oligothiophene dyes reveals that the strain-like properties of aggregated Aβ are maintained in fixed tissues. The resistance of Aβ seeds to inactivation and structural modification by formaldehyde underscores their remarkable durability, which in turn may contribute to their persistence and spread within the body. The present findings can be exploited to establish the relationship between the molecular structure of Aβ aggregates and the variable clinical features and disease progression of AD even in archived, formalin-fixed autopsy material.

Pathological, genetic, and biochemical hallmarks of Alzheimer's disease (AD) are linked to amyloid-β (Aβ) peptide aggregation. Especially misfolded Aβ42peptide is sufficient to promote amyloid plaque formation. However, the cellular compartment facilitating the conversion of monomeric Aβ to aggregated toxic Aβ species remains unknown.In vitromodels suggest lipid membranes to be the driving force of Aβ conversion. To this end, we generated two novel mouse models, expressing either membrane-anchored or nonanchored versions of the human Aβ42peptide. Strikingly, membrane-anchored Aβ42robustly accelerated Aβ deposition and exacerbated amyloid-associated toxicity upon crossing with Aβ precursor protein transgenic mice. Thesein vivofindings support the hypothesis that Aβ–membrane interactions play a pivotal role in early-onset AD as well as neuronal damage and provide evidence to study Aβ–membrane interactions as therapeutic targets.

SummaryA central question about human brain aging is whether cognitive enrichment slows the development of Alzheimer changes. Here, we show that prolonged exposure to an enriched environment (EE) facilitated signaling in the hippocampus of wild-type mice that promoted long-term potentiation. A key feature of the EE effect was activation of β2-adrenergic receptors and downstream cAMP/PKA signaling. This EE pathway prevented LTP inhibition by soluble oligomers of amyloid β-protein (Aβ) isolated from AD cortex. Protection by EE occurred in both young and middle-aged wild-type mice. Exposure to novelty afforded greater protection than did aerobic exercise. Mice chronically fed a β-adrenergic agonist without EE were protected from hippocampal impairment by Aβ oligomers. Thus, EE enhances hippocampal synaptic plasticity by activating β-adrenoceptor signaling and mitigating synaptotoxicity of human Aβ oligomers. These mechanistic insights support using prolonged exposure to cognitive novelty and/or oral β-adrenergic agonists to lessen the effects of Aβ accumulation during aging.

γ-Secretase is essential for the generation of the neurotoxic 42-amino acid amyloid β-peptide (Aβ(42)). The aggregation-prone hydrophobic peptide, which is deposited in Alzheimer disease (AD) patient brain, is generated from a C-terminal fragment of the β-amyloid precursor protein by an intramembrane cleavage of γ-secretase. Because Aβ(42) is widely believed to trigger AD pathogenesis, γ-secretase is a key AD drug target. Unlike inhibitors of the enzyme, γ-secretase modulators (GSMs) selectively lower Aβ(42) without interfering with the physiological function of γ-secretase. The molecular target(s) of GSMs and hence the mechanism of GSM action are not established. Here we demonstrate by using a biotinylated photocross-linkable derivative of highly potent novel second generation GSMs that γ-secretase is a direct target of GSMs. The GSM photoprobe specifically bound to the N-terminal fragment of presenilin, the catalytic subunit of γ-secretase, but not to other γ-secretase subunits. Binding was differentially competed by GSMs of diverse structural classes, indicating the existence of overlapping/multiple GSM binding sites or allosteric alteration of the photoprobe binding site. The β-amyloid precursor protein C-terminal fragment previously implicated as the GSM binding site was not targeted by the compound. The identification of presenilin as the molecular target of GSMs directly establishes allosteric modulation of enzyme activity as a mechanism of GSM action and may contribute to the development of therapeutically active GSMs for the treatment of AD.

Background: In Alzheimer’s disease (AD), amyloid-β (Aβ) is the major component of extracellular plaques, whereas the microtubule-associated protein tau forms the main component of intracellular tangles. In contrast to frontotemporal dementias and other neurodegenerative diseases, both proteins form pathological aggregates and are considered key players for the development of AD. However, the connection between Aβ and tau and the functional loss of neurons and synapses, which ultimately lead to cognitive impairments, is still not well understood. Objectives: Making use of primary neurons exposed to Aβ oligomers, we sought to determine how tau mediates the Aβ-induced neuronal dysfunction. Additionally, we asked how the microtubule cytoskeleton is involved in the combined Aβ and tau toxicity. Methods: We exposed mature primary rat neurons with developed synapses to Aβ oligomers and used immunofluorescence and electron microscopy to investigate tau, actin, neurofilament and microtuble cytoskeleton changes. Results: Aβ oligomers preferentially associate with synapses, notably dendritic spines, throughout the neuronal cell culture. As a consequence, endogenous tau gets missorted from the axonal into the somatodendritic compartment in a subset of cells. These missorted cells also display missorting of neurofilaments, and a dramatic loss of microtubules, which can be prevented by the microtubule stabilizer taxol. Conclusions: Aβ causes tau missorting, loss of neuronal cell polarity and loss of dendritic microtubules. This in turn leads to impaired organelle/mitochondria transport, whereby synapses cannot be maintained properly and eventually decay. The data support the view that the microtubule cytoskeleton is a valid therapeutic target in AD.