22 research outputs found
Recommended from our members
Research data for paper: Elevated amyloid beta disrupts the nanoscale organization and function of synaptic vesicle pools in hippocampal neurons
Data for paper published in Cerebral Cortex on 03/04/22
Files:
Biasettietal-VPools-TEM (Transmission electron microscope)-maps: TEM-based vesicle pool maps by synapse and condition, used to generate distance plots.
Biasettietal-SypHy-fractions: Fluorescence SypHy values used to calculate pool fractions by synapse and condition.
Biasettietal-iGluSNFR: iGluSnFR response values by synapse, experiment and condition.
Biasettietal-EMdistVtoAZ: Coordinates of each vesicle from TEM images measured as distances to nearest point on the active zone; this is used to generate mean distances for each pool class (PC+: photoconverted, PC-: non-photoconverted) and also to construct cumulative distance plots; measurements were collected for both WT (wild-type) and APPSwe/Ind mice.
Biasettietal-iGluSnFRrationale: Background dataset used to establish iGluSnFR protocol used in the paper
Biasettietal-abeta-time-conc-rationale: Supporting figure outlining the rationale for time and concentration used in this work.
Abstract
Alzheimer’s disease is linked to increased levels of amyloid beta (Aβ) in the brain, but the mechanisms underlying neuronal dysfunction and neurodegeneration remain enigmatic. Here, we investigate whether organizational characteristics of functional presynaptic vesicle pools, key determinants of information transmission in the central nervous system, are targets for elevated Aβ. Using an optical readout method in cultured hippocampal neurons, we show that acute Aβ42 treatment significantly enlarges the fraction of functional vesicles at individual terminals. We observe the same effect in a chronically elevated Aβ transgenic model (APPSw,Ind) using an ultrastructure-function approach that provides detailed information on nanoscale vesicle pool positioning. Strikingly, elevated Aβ is correlated with excessive accumulation of recycled vesicles near putative endocytic sites, which is consistent with deficits in vesicle retrieval pathways. Using the glutamate reporter, iGluSnFR, we show that there are parallel functional consequences, where ongoing information signaling capacity is constrained. Treatment with levetiracetam, an antiepileptic that dampens synaptic hyperactivity, partially rescues these transmission defects. Our findings implicate organizational and dynamic features of functional vesicle pools as targets in Aβ-driven synaptic impairment, suggesting that interventions to relieve the overloading of vesicle retrieval pathways might have promising therapeutic value.</p
Recommended from our members
The involvement of Aß42 and tau in nucleolar and protein synthesis machinery dysfunction
Alzheimer’s disease (AD) is the most common form of dementia and is distinguished from other dementias by observation of extracellular Amyloid-b (Ab) plaques and intracellular neurofibrillary tangles, comprised of fibrils of Ab and tau protein, respectively. At early stages, AD is characterized by minimal neurodegeneration, oxidative stress, nucleolar stress, and altered protein synthesis machinery. It is generally believed that Ab oligomers are the neurotoxic species and their levels in the AD brain correlate with the severity of dementia suggesting that they play a critical role in the pathogenesis of the disease. Here, we show that the incubation of differentiated human neuroblastoma cells (SHSY5Y) with freshly prepared Ab42 oligomers initially resulted in oxidative stress and subtle nucleolar stress in the absence of DNA damage or cell death. The presence of exogenous Ab oligomers resulted in altered nuclear tau levels as well as phosphorylation state, leading to altered distribution of nucleolar tau associated with nucleolar stress. These markers of cellular dysfunction worsen over time alongside a reduction in ribosomal RNA synthesis and processing, a decrease in global level of newly synthesized RNA and reduced protein synthesis. The interplay between Ab and tau in AD remains intriguing and Ab toxicity has been linked to tau phosphorylation and changes in localization. These findings provide evidence for the involvement of Ab42 effects on nucleolar tau and protein synthesis machinery dysfunction in cultured cells. Protein synthesis dysfunction is observed in mild cognitive impairment and early AD in the absence of significant neuronal death
Recommended from our members
Misfolded amyloid-ß-42 impairs the endosomal–lysosomal pathway
Misfolding and aggregation of proteins is strongly linked to several neurodegenerative diseases, but how such species bring about their cytotoxic actions remains poorly understood. Here we used specifically-designed optical reporter probes and live fluorescence imaging in primary hippocampal neurons to characterise the mechanism by which prefibrillar, oligomeric forms of the Alzheimer’s-associated peptide, Aß42, exert their detrimental effects. We used a pH-sensitive reporter, Aß42-CypHer, to track Aß internalisation in real-time, demonstrating that oligomers are rapidly taken up into cells in a dynamin-dependent manner, and trafficked via the endo-lysosomal pathway resulting in accumulation in lysosomes. In contrast, a non-assembling variant of Aß42 (vAß42) assayed in the same way is not internalised. Tracking ovalbumin uptake into cells using CypHer or Alexa Fluor tags shows that preincubation with Aß42 disrupts protein uptake. Our results identify a potential mechanism by which amyloidogenic aggregates impair cellular function through disruption of the endosomal–lysosomal pathway
Recommended from our members
Structure dependent effects of Amyloid-ß on long-term memory in Lymnaea stagnalis
Amyloid-ß (Aß) peptides are implicated in the causation of memory loss, neuronal impairment, and neurodegeneration in Alzheimer's disease. Our recent work revealed that Aß 1–42 and Aß 25–35 inhibit long-term memory (LTM) recall in Lymnaea stagnalis (pond snail) in the absence of cell death. Here, we report the characterization of the active species prepared under different conditions, describe which Aß species is present in brain tissue during the behavioral recall time point and relate the sequence and structure of the oligomeric species to the resulting neuronal properties and effect on LTM. Our results suggest that oligomers are the key toxic Aß1–42 structures, which likely affect LTM through synaptic plasticity pathways, and that Aß 1–42 and Aß 25–35 cannot be used as interchangeable peptides
Shapeshifting tau: from intrinsically disordered to paired-helical filaments
Tau is an intrinsically disordered protein that has the ability to self-assemble to form paired helical and straight filaments in Alzheimer’s disease, as well as the ability to form additional distinct tau filaments in other tauopathies. In the presence of microtubules, tau forms an elongated form associated with tubulin dimers via a series of imperfect repeats known as the microtubule binding repeats. Tau has recently been identified to have the ability to phase separate in vitro and in cells. The ability of tau to adopt a wide variety of conformations appears fundamental both to its biological function and also its association with neurodegenerative diseases. The recently highlighted involvement of low-complexity domains in liquid–liquid phase separation provides a critical link between the soluble function and the insoluble dysfunctional properties of tau.</p
Recommended from our members
The structure of cross-ß tapes and tubes formed by an octapeptide, aSß1
Elaborate morphology: The aSß1 peptide, a fragment of a-synuclein, assembles into flat tapes consisting of a peptide bilayer, which can be modeled based on the cross-ß structure found in amyloid proteins. The tapes are stabilized by hydrogen bonding, whilst the amphiphilic nature of the peptide results in the thin bilayer structure. To further stabilize the structure, these tapes may twist to form helical tapes, which subsequently close into nanotubes
Recommended from our members
Effects of Aß exposure on longterm associative memory and its neuronal mechanisms in a defined neuronal network
Amyloid beta (Aß ) induced neuronal death has been linked to memory loss, perhaps the most devastating symptom of Alzheimer’s disease (AD). Although Aß -induced impairment of synaptic or intrinsic plasticity is known to occur before any cell death, the links between these neurophysiological changes and the loss of specific types of behavioral memory are not fully understood. Here we used a behaviorally and physiologically tractable animal model to investigate Aß -induced memory loss and electrophysiological changes in the absence of neuronal death in a defined network underlying associative memory. We found similar behavioral but different neurophysiological effects for Aß 25-35 and Aß 1-42 in the feeding circuitry of the snail Lymnaea stagnalis. Importantly, we also established that both the behavioral and neuronal effects were dependent upon the animals having been classically conditioned prior to treatment, since Aß application before training caused neither memory impairment nor underlying neuronal changes over a comparable period of time following treatment
Recommended from our members
Metal- and UV- catalyzed oxidation results in trapped amyloid-ß intermediates revealing that self-assembly is required for Aß-induced cytotoxicity
Dityrosine (DiY), via the cross-linking of tyrosine residues, is a marker of protein oxidation, which increases with aging. Amyloid-ß (Aß) forms DiY in vitro and DiY-cross-linked Aß is found in the brains of patients with Alzheimer disease. Metal- or UV- catalyzed oxidation of Aß42 results in an increase in DiY cross-links. Using DiY as a marker of oxidation, we compare the self-assembly propensity and DiY cross-link formation for a non-assembly competent variant of Aß42 (vAß) with wild-type Aß42. Oxidation results in the formation of trapped wild-type Aß assemblies with increased DiY cross-links that are unable to elongate further. Assembly-incompetent vAß and trapped Aß assemblies are non-toxic to neuroblastoma cells at all stages of self-assembly, in contrast to oligomeric, non-cross-linked Aß. These findings point to a mechanism of toxicity that necessitates dynamic self-assembly whereby trapped Aß assemblies and assembly-incompetent variant Aß are unable to result in cell death
Recommended from our members
Sequence Determinants for Amyloid Fibrillogenesis of Human a-Synuclein
Parkinson's disease (PD) and dementia with Lewy bodies (DLB) are characterized by the presence of filamentous inclusions in nerve cells. These filaments are amyloid fibrils that are made of the protein a-synuclein, which is genetically linked to rare cases of PD and DLB. ß-Synuclein, which shares 60% identity with a-synuclein, is not found in the inclusions. Furthermore, while recombinant a-synuclein readily assembles into amyloid fibrils, ß-synuclein fails to do so. It has been suggested that this may be due to the lack in ß-synuclein of a hydrophobic region that spans residues 73¿83 of a-synuclein. Here, fibril assembly of recombinant human a-synuclein, a-synuclein deletion mutants, ß-synuclein and ß/a-synuclein chimeras was assayed quantitatively by thioflavin T fluorescence and semi-quantitatively by transmission electron microscopy. Deletion of residues 73¿83 from a-synuclein did not abolish filament formation. Furthermore, a chimera of ß-synuclein with a-synuclein(73¿83) inserted was significantly less fibrillogenic than wild-type a-synuclein. These findings, together with results obtained using a number of recombinant synucleins, showed a correlation between fibrillogenesis and mean ß-strand propensity, hydrophilicity and charge of the amino acid sequences. The combination of these simple physicochemical properties with a previously described calculation of ß-strand contiguity allowed us to design mutations that changed the fibrillogenic propensity of a-synuclein in predictable way