Chronic Cytosolic Calcium Changes Induced by Alzheimer's Peptides Aβ Allows for Sorting of Aβ-Resistant Cell Subpopulations

Abstract

The Alzheimer's Aβ peptides interaction with the plasma membrane has been shown to result in a chronic increase in the cytosolic calcium concentrations of cells in culture. Such alteration may cause a variety of secondary effects which may lead to cell degeneration and reduced cell culture growth. We have proposed that the alteration in the cytosolic calcium concentrations induced by Aβ is initiated by early formation of Aβ ion channels in the plasma membrane. To study the Aβ-induced cytosolic calcium change dynamics we monitored the level of cytosolic calcium from hundreds of individual cells in culture exposed to Aβ for prolonged period of time. Because of the continuous cytotoxic effect of Aβ on cells, the number of viable cells in the cultures varied with time of exposure. Histograms based on the individual intracellular calcium levels showed that while most cells in the culture remain unaffected, a defined subpopulation of cells show increasingly higher than normal cytosolic calcium levels. After days of continuous exposure to Aβ cells more sensitive to Aβ died, and consequently the averaged cytosolic calcium for remaining cells in the whole culture approached control values. Aβ ion channel blockers prevented the induced cytosolic calcium changes and preserved cell viability, confirming the participation of Aβ ion channels. Confocal microscopic analysis using fluorescent annexin V on cells loaded with fura 2 showed that those cells that remained after prolonged exposure to Aβ did not display the proposed extracellular Aβ receptor phosphatidyl serine. Further addition of Aβ to this sorted cell subpopulation did not induce cytosolic calcium changes and cells continued to grow and divide. Application of this procedure allows for sorting out Aβ resistant subpopulation of cells.