Synapse loss correlates with a cognitive decline in Alzheimer's disease (AD), but whether this is caused by fibrillar deposits known as senile plaques or soluble oligomeric forms of amyloid ␤ (A␤) is controversial. By using array tomography, a technique that combines ultrathin sectioning of tissue with immunofluorescence, allowing precise quantification of small structures, such as synapses, we have tested the hypothesis that oligomeric A␤ surrounding plaques contributes to synapse loss in a mouse model of AD. We find that senile plaques are surrounded by a halo of oligomeric A␤. Analysis of >14,000 synapses (represented by PSD95-stained excitatory synapses) shows that there is a 60% loss of excitatory synapses in the halo of oligomeric A␤ surrounding plaques and that the density increases to reach almost control levels in volumes further than 50 m from a plaque in an approximately linear fashion (linear regression, r 2 ‫؍‬ 0.9; P < 0.0001). Further, in transgenic cortex, microdeposits of oligomeric A␤ associate with a subset of excitatory synapses, which are significantly smaller than those not in contact with oligomeric A␤. The proportion of excitatory synapses associated with A␤ correlates with decreasing density (correlation, ؊0.588; P < 0.0001). These data show that senile plaques are a potential reservoir of oligomeric A␤, which colocalizes with the postsynaptic density and is associated with spine collapse, reconciling the apparently competing schools of thought of ''plaque'' vs. ''oligomeric A␤'' as the synaptotoxic species in the brain of AD patients.Alzheimer ͉ array tomography ͉ neurodegeneration ͉ synaptotoxicity L oss of connectivity caused by neuronal death and synapse loss is thought to underlie cognitive decline in neurodegenerative conditions, such as Alzheimer's disease (AD). Synapse loss appears to be particularly important in the pathogenesis of AD. Indeed, it is known that synapses are lost during AD and that in AD tissue, synapse loss correlates strongly with cognitive decline (1-3). There is a growing consensus, based primarily on cell-based assays, that amyloid ␤ (A␤), the main component of senile plaques, is toxic to synapses (4-6). In both AD patients and animal models of the disease, synapse loss is greatest near senile plaques, indicating a link between amyloid pathology and synaptotoxicity in vivo. Work by several groups has shown a decrease in dendritic spine density and synaptophysin-positive synapses radiating out from the surface of plaques in mouse models of AD (7-10). Whether this is caused by fibrillar plaques or soluble oligomeric A␤ is controversial. We used multiphoton imaging of the living brain to show that this spine loss is caused by impaired spine stability over time near plaques and postulated that a plaque-related diffusible bioactive molecule was responsible (11). Here, we test the hypothesis that oligomeric A␤ is directly synaptotoxic.We hypothesize that soluble oligomeric A␤ associates with the postsynaptic density and causes the loss of synapses and spines observ...

Amyloid ␤ (A␤)-containing plaques are surrounded by dystrophic neurites in the Alzheimer's disease (AD) brain, but whether and how plaques induce these neuritic abnormalities remain unknown. We tested the hypothesis that soluble oligomeric assemblies of A␤, which surround plaques, induce calcium-mediated secondary cascades that lead to dystrophic changes in local neurites. We show that soluble A␤ oligomers lead to activation of the calcium-dependent phosphatase calcineurin (CaN) (PP2B), which in turn activates the transcriptional factor nuclear factor of activated T cells (NFAT). Activation of these signaling pathways, even in the absence of A␤, is sufficient to produce a virtual phenocopy of A␤-induced dystrophic neurites, dendritic simplification, and dendritic spine loss in both neurons in culture and in the adult mouse brain. Importantly, the morphological deficits in the vicinity of A␤ deposits in a mouse model of AD are ameliorated by CaN inhibition, supporting the hypothesis that CaN-NFAT are aberrantly activated by A␤ and that CaN-NFAT activation is responsible for disruption of neuronal structure near plaques. In accord with this, we also detect increased levels of an active form of CaN and NFATc4 in the nuclear fraction from the cortex of patients with AD. Thus, A␤ appears to mediate the neurodegeneration of AD, at least in part, by activation of CaN and subsequent NFAT-mediated downstream cascades.

In Alzheimer disease (AD), deposition of neurofibrillary tangles and loss of synapses in the neocortex and limbic system each correlate strongly with cognitive impairment. Tangles are composed of misfolded hyperphosphorylated tau proteins; however, the link between tau abnormalities and synaptic dysfunction remains unclear. We examined the location of tau in control and AD cortices using biochemical and morphologic methods. We found that, in addition to its well-described axonal localization, normal tau is present at both presynaptic and postsynaptic terminals in control human brains. In AD, tau becomes hyperphosphorylated and misfolded at both presynaptic and postsynaptic terminals, and this abnormally posttranslationally modified tau is enriched in synaptoneurosomal fractions. Synaptic tau seems to be hyperphosphorylated and ubiquitinated, and forms stable oligomers resistant to SDS denaturation. The accumulation of hyperphosphorylated tau oligomers at human AD synapses is associated with increased ubiquitinated substrates and increased proteasome components, consistent with dysfunction of the ubiquitin-proteasome system. Our findings suggest that synaptic hyperphosphorylated tau oligomers may be an important mediator of the proteotoxicity that disrupts synapses in AD.

The paradoxical appearance of aggregated α-synuclein (αsyn) in naive transplanted embryonic stem cells in Parkinson's disease (PD) brains has recently been reported, highlighting the possibility of neuron to neuron transmission of αsyn in PD. Here, we demonstrate in a cellular model the presence of αsyn oligomers in the extracellular space, their uptake by neurons, retrograde axonal transport to cell soma, and detrimental effects on neighboring cells. Moreover, we demonstrate that Hsp70 chaperones αsyn in the extracellular space and reduces extracellular αsyn oligomer formation and related toxicity. These novel findings provide evidence that extracellular αsyn oligomers may represent a crucial player in the propagation of pathology in PD, with their modulation by Hsp70 representing a potential new target for therapeutic interventions.

The apolipoprotein E ε4 gene is the most important genetic risk factor for sporadic Alzheimer's disease, but the link between this gene and neurodegeneration remains unclear. Using array tomography, we analysed >50000 synapses in brains of 11 patients with Alzheimer's disease and five non-demented control subjects and found that synapse loss around senile plaques in Alzheimer's disease correlates with the burden of oligomeric amyloid-β in the neuropil and that this synaptotoxic oligomerized peptide is present at a subset of synapses. Further analysis reveals apolipoprotein E ε4 patients with Alzheimer's disease have significantly higher oligomeric amyloid-β burden and exacerbated synapse loss around plaques compared with apolipoprotein E ε3 patients. Apolipoprotein E4 protein colocalizes with oligomeric amyloid-β and enhances synaptic localization of oligomeric amyloid-β by >5-fold. Biochemical characterization shows that the amyloid-β enriched at synapses by apolipoprotein E4 includes sodium dodecyl sulphate-stable dimers and trimers. In mouse primary neuronal culture, lipidated apolipoprotein E4 enhances oligomeric amyloid-β association with synapses via a mechanism involving apolipoprotein E receptors. Together, these data suggest that apolipoprotein E4 is a co-factor that enhances the toxicity of oligomeric amyloid-β both by increasing its levels and directing it to synapses, providing a link between apolipoprotein E ε4 genotype and synapse loss, a major correlate of cognitive decline in Alzheimer's disease.

We raised monoclonal antibodies against senile plaque (SP) amyloid and obtained a clone 9D2, which labeled amyloid ®brils in SPs and reacted with~50/100 kDa polypeptides in Alzheimer's disease (AD) brains. We puri®ed the 9D2 antigens and cloned a cDNA encoding its precursor, which was a novel type II transmembrane protein speci®cally expressed in neurons. This precursor harbored three collagen-like Gly±X±Y repeat motifs and was partially homologous to collagen type XIII. Thus, we named the 9D2 antigen as CLAC (collagen-like Alzheimer amyloid plaque component), and its precursor as CLAC-P/collagen type XXV. The extracellular domain of CLAC-P/collagen type XXV was secreted by furin convertase, and the N-terminus of CLAC deposited in AD brains was pyroglutamate modi®ed. Both secreted and membrane-tethered forms of CLAC-P/collagen type XXV speci®cally bound to ®brillized Ab, implicating these proteins in b-amyloidogenesis and neuronal degeneration in AD.

Alzheimer’s disease (AD) is the most common progressive neurodegenerative disorder causing dementia. Massive deposition of amyloid β peptide (Aβ) as senile plaques in the brain is the pathological hallmark of AD, but oligomeric, soluble forms of Aβ have been implicated as the synaptotoxic component. The apolipoprotein E epsilon 4 (apoE ε4) allele is known to be a genetic risk factor for developing AD. However it is still unknown how apoE impacts the process of Aβ oligomerization. Here, we found that the level of Aβ oligomers in APOEε4/ε4 AD patient brains is 2.7 times higher than those in APOEε3/ε3 AD patient brains, matched for total plaque burden, suggesting that apoE4 impacts the metabolism of Aβ oligomers. To test this hypothesis, we examined apoE’s effect on Aβ oligomer formation. Using both synthetic Aβ and a split-luciferase method for monitoring Aβ oligomers, we observed that apoE increased the level of Aβ oligomers in an isoform dependent manner (E2 < E3 < E4). This effect appears to be dependent on the ApoE carboxy-terminal domain. Moreover, these results were confirmed using endogenous apoE isolated from the TBS-soluble fraction of human brain, which increased the formation of Aβ oligomers. Taken together, these data show that lipidated apoE, especially apoE4, increases Aβ oligomers in the brain. Higher levels of Aβ oligomers in the brains of APOEε4/ε4 carriers compared to APOEε3/ε3 carriers may increase the loss of dendritic spines and accelerate memory impairments, leading to earlier cognitive decline in AD.

In vivo experimental evidence indicates that acute neuronal activation increases Aβ release from presynaptic terminals, whereas long-term effects of chronic synaptic activation on Aβ pathology remain unclear. To address this issue, we adopted optogenetics and transduced stabilized step-function opsin, a channelrhodopsin engineered to elicit a long-lasting neuronal hyperexcitability, into the hippocampal perforant pathway of APP transgenic mice. In vivo microdialysis revealed a ∼24% increase in the hippocampal interstitial fluid Aβ42 levels immediately after acute light activation. Five months of chronic optogenetic stimulation increased Aβ burden specifically in the projection area of the perforant pathway (i.e., outer molecular layer of the dentate gyrus) of the stimulated side by ∼2.5-fold compared with that in the contralateral side. Epileptic seizures were observed during the course of chronic stimulation, which might have partly contributed to the Aβ pathology. These findings implicate functional abnormalities of specific neuronal circuitry in Aβ pathology and Alzheimer disease.