Neurotoxic effects of thioflavin S-positive amyloid deposits in transgenic mice and Alzheimer's disease

Abstract

Despite extensive deposition of putatively neurotoxic amyloid-beta (Abeta) protein in the brain, it has not been possible to demonstrate an association of Abeta deposits with neuronal loss in Alzheimer's disease (AD), and neuronal loss is minimal in transgenic mouse models of AD. Using triple immunostaining confocal microscopy and analyzing the images with the cross-correlation density map method from statistical physics, we directly compared Abeta deposition, Abeta morphology, and neuronal architecture. We found dramatic, focal neuronal toxicity associated primarily with thioflavin S-positive fibrillar Abeta deposits in both AD and PSAPP mice. These results, along with computer simulations, suggest that Abeta develops neurotoxic properties in vivo when it adopts a fibrillar beta-pleated sheet conformation.

Fig 1.

(a) Color superposition of triple-labeled images of human brain tissue (AD case) from a dual photon microscope showing coregistration of neurons with Aβ-immunoreactive and ThioS-labeled deposits in layer III of the STS: Neu-N-stained neurons (red), Aβ-immunoreactive deposits (blue), and ThioS labeling (green), showing tangles in addition to ThioS-positive Aβ deposits. A schematic picture of concentric rings around one deposit illustrates the density map method used in the calculation. ThioS-positive Aβ channel (green) was used to classify the plaques into ThioS-positive and non-ThioS deposits. (b) Color superposition of confocal double-labeled immunostained images of transgenic mouse brain tissue showing coregistration of neurons (red) with Aβ-immunoreactive (blue) and ThioS-labeled (green) deposits. The arrows point to two ThioS deposits that coincide with two holes in the neuronal density.

Fig 2.

(a) Normalized neuronal density ρ_N(r) averaged over five AD cases with average neuron densities of 1,563, 763, 1,087, 1,597, and 862 neurons per mm² for (i) all Aβ deposits with n = 1,287 (black), (ii) non-ThioS Aβ deposits with n = 996 (cyan), (iii) ThioS-positive Aβ deposits with n = 223 (red), and (iv) randomly shuffled Aβ deposits with n = 1,287 (gray). (Inset) Normalized neuronal density ρ_N(r) for different size classes of Aβ deposits with respect to the radius R of the deposit, averaged over five different AD cases: (i) R < 7 μm (red), (ii) 7 < R < 10 μm (orange), (iii) 10 < R < 15 μm (green), and (iv) R > 15 μm (blue). (b) Neuronal density ρ(r) in transgenic mice with an average neuronal density of 2,468 per mm² with respect to the center of mass of the Aβ deposits. We plot ρ(r) for all Aβ deposits with n = 982 (black), non-ThioS (cyan) with n = 819, and ThioS-positive (red) Aβ deposits with n = 166. (Inset) Neuronal densities ρ(r) around ThioS-positive Aβ deposits of different effective radia R: (i) R < 12.3 μm (red) with n = 31, (ii) 12.3 < R < 24.8 μm (orange) with n = 78, and (iii) R > 24.8 μm (green) with n = 58.

Fig 3.

Schematic pictures and diagrams for possible biological scenarios of Aβ deposit effects on neuronal landscape. (A) Nontoxic diffuse deposit. (B) Nontoxic deposit displacing neurons. (C) Focally toxic deposit. (D) Focally toxic deposit with a toxic penumbra.

Fig 4.

(a) Neuronal density ρ(r) from the molecular dynamics simulations for the nontoxic and toxic model. In each case, ρ(r) is averaged over a set of sizes obtained from the ThioS size distribution with n = 254. Neuronal density ρ(r) for the nontoxic model shows a rather prominent peak of neuronal density that would correspond to a denser ring of neurons around the deposit. Neuronal density ρ(r) for the toxic model displays no such peak, indicating that neurons are not merely pushed away as the deposit grows but rather destroyed. The Insets correspond to typical computer realizations for each model, where the initial position for the nonoverlapping small circles (model neurons) is random and the total density and radius [N_neu = 1,850 in a 512 × 512 pixel² box, R = 4 pixels (3.5 μm)] is taken to be equivalent to the average density and radius of neurons in the PSAPP transgenic mice. The central deposit (only the hole shown) in each Inset has a radius of 18 pixels (15.7 μm). (b) Neuronal density ρ(r) in dependence on the distance from the surface of a ThioS core in transgenic mice. For each deposit ρ(r) is shifted by the radius of the ThioS core. Individual functions ρ(r) are then averaged over all 281 ThioS cores. (Inset) A confocal image of Neu-N-stained PsAPP mouse brain showing a hole in the otherwise rather homogeneous distribution of neurons.