Co-Localization of Amyloid Beta and Tau Pathology in Alzheimer's Disease Synaptosomes

Abstract

The amyloid cascade hypothesis proposes that amyloid beta (Abeta) pathology precedes and induces tau pathology, but the neuropathological connection between these two lesions has not been demonstrated. We examined the regional distribution and co-localization of Abeta and phosphorylated tau (p-tau) in synaptic terminals of Alzheimer's disease brains. To quantitatively examine large populations of individual synaptic terminals, flow cytometry was used to analyze synaptosomes prepared from cryopreserved Alzheimer's disease tissue. An average 68.4% of synaptic terminals in the Alzheimer's disease cohort (n = 11) were positive for Abeta, and 32.3% were positive for p-tau; Abeta and p-tau fluorescence was lowest in cerebellum. In contrast to synaptic p-tau, which was highest in the entorhinal cortex and hippocampus (P = 0.004), synaptic Abeta fluorescence was significantly lower in the entorhinal cortex and hippocampus relative to neocortical regions (P = 0.0003). Synaptic Abeta and p-tau fluorescence was significantly correlated (r = 0.683, P < 0.004), and dual-labeling experiments demonstrated that 24.1% of Abeta-positive terminals were also positive for p-tau, with the highest fraction of dual labeling (39.3%) in the earliest affected region, the entorhinal cortex. Western blotting experiments show a significant correlation between synaptic Abeta levels measured by flow cytometry and oligomeric Abeta species (P < 0.0001). These results showing overlapping Abeta and tau pathology are consistent with a model in which both synaptic loss and dysfunction are linked to a synaptic amyloid cascade within the synaptic compartment.

Synaptic Amyloid-β Oligomers Precede p-Tau and Differentiate High Pathology Control Cases

“…ajp.amjpathol.org -The American Journal of Pathology anti-Ab antibodies are sensitive to conformation and differentially label aggregated Ab preparations. 46 As previously reported, synaptic terminals from control cases show little to no Ab immunolabeling ( Figure 1C) 26,33,42,47 ; representative plots from AD cases illustrate the rise in synaptic Ab with increasing neuropathologic disease stage and the degree to which synaptic terminal Ab increases between plaque stage 0 (no plaque deposition) and stage C (end-stage plaque deposition) (Figure 1, D and E). 48 Figure 2A shows group differences in aggregate flow cytometric data [F(3,30) Z 6.13, P Z 0.002] in both early-stage (Braak II to IV) and late-stage (Braak V to VI) AD cases.…”

“…Because Western blot analysis of P-2 samples shows a strong signal for postsynaptic density protein 95, 26 it is possible that some of the Ab signal in P-2 samples arises from extracellular Ab bound to postsynaptic surface receptors, particularly heparan sulfate proteoglycan. 66 However, prior electron microscopic studies reported by our group and others document the spherical and primarily presynaptic structure of synaptosomes, 33,40 and incubation of AD synaptosomes with a combination of trypsin and heparinase does not reduce Ab labeling. Confirmation of presynaptic localization of oligomers by electron microscope would be optimal; however, detection of intraneuronal, particularly oligomeric, Ab is limited by the technical difficulty of detecting Ab antigens, which are hydrophobic and difficult to detect by conventional immunocytochemical methods, especially in postmortem tissue.…”

“…33 The anti-tau Ser396/404 phosphospecific PHF-1 antibody was a generous gift from Peter Davies (Albert Einstein College of Medicine, Bronx, NY). The conformation-dependent anti-Ab antibodies (prefibrillar oligomer-specific M55 and fibrilspecific M116) were a generous gift from Charles G. Glabe (University of California Irvine, Irvine, CA).…”

“…Detailed information about individual cases is presented in Table 1. For cryopreservation, samples were minced and slowly frozen in isotonic sucrose on the day of autopsy as previously described 33 ; the same protocols were used at each of the three institutions that provided samples.…”

“…22,33,59 With the use of cryopreserved cortex samples from the same transgenic rat age groups as above (3, 9, and 20 months), we investigated the association of synaptic p-tau with disease stage and amyloid pathology. P-2 samples were immunolabeled for p-tau; flow cytometric analyses are illustrated in Figure 6, AeC and show a representative increase in synaptic PHF-1 fluorescence in transgenic relative to wild-type animals at 20 months.…”