Figure 2.

ADDLs are diffusible, extremely potent CNS neurotoxins. ADDLs in culture medium diffuse through culture support filters and cause extensive cell death in stratum granulosum (DG) and CA3 areas of organotypic hippocampal slice cultures. ADDL toxicity is extensive even at nanomolar doses, but nondiffusing fibrils at 20 μM are not toxic. (A) DG and CA3 area of a hippocampal slice treated for 24 hr with 5 μM ADDLs. Dead cells are highlighted in false yellow color (see Materials and Methods). Up to 40% of the cells in this region die following chronic exposure to ADDLs. (B) DG and CA3 area of another hippocampal slice treated with 20 μM fibrillar Aβ_1–42 for 24 hr. No cell death is seen. (C) Live cells in the same area of the slice treated with fibrillar Aβ_1–42. (D) ADDLs were added to duplicate mouse hippocampal slices for 24 hr at the indicated concentrations. The Live/Dead assay and image analysis (see Materials and Methods) were used to determine percent dead cells in fields containing the DG and CA3 areas. Data show that even after a 1,000-fold dilution to 5 nM Aβ_1–42, ADDLs still kill >20% of the cells, a value greater than half the maximum cell death. (Inset) Comparison of cell death in DG and CA3 observed with vehicle alone (Upper) or with ADDLs at 5 μM (Lower). ADDL concentration was determined by measuring the supernatant protein and subtracting the amount of soluble clusterin present. This formula assumes the amount of clusterin that pellets is negligible.