Direct Observation of Single Oligomers of the Alzheimer’s Amyloid-β Peptide on Live Cell Membranes

Abstract

Oligomeric species of the amyloid-β peptide are strongly implicated in the synaptic dysfunction and neuronal loss seen in Alzheimer's disease. While proposed mechanisms for oligomeric toxicity are abundant and diverse, many involve amyloid-β (Aβ) interaction with cell membranes, either via direct insertion into the cell membrane or by binding to specific cell-surface receptors. One such hypothesis holds that Aβ forms calcium-permeable channels within the membranes of neurons, disrupting homeostasis and triggering an apoptotic signaling cascade. However, both the exact identity of the toxic aggregate or aggregates and the mechanism by which the toxicity is mediated remain undetermined. Study of this system under physiological conditions presents a challenge, as oligomeric forms of the peptide in vivo are heterogeneous and metastable, and Aβ itself is normally present in the brain at only nanomolar concentrations. We recently used single molecule spectroscopy to detect stable, conductive low-order Aβ(1-40) oligomers in synthetic membranes exposed to nanomolar levels of the peptide. Similar oligomers may form on the membranes of live cells and, by disrupting cell membrane integrity, contribute to the functional abnormalities and neuronal death observed in Alzheimer's. With single molecule confocal laser scanning microscopy, we have identified small Aβ(1-40) oligomers bound to the membranes of SH-SY5Y neuroblastoma cells after ten minutes’ exposure to low (50nM) peptide concentrations. Additionally, we have used single-particle fluorescence intensity measurements to characterize the oligomeric states of these cell-bound aggregates. We find that small oligomers ranging from dimers to octamers form in solution under physiological conditions and that the size distribution shifts towards larger oligomers for Aβ(1-40) on the cell membrane, which may indicate that oligomer growth can occur following membrane binding.