The influence of GAPT extraction on synapse loss of APPswe/PS1dE9 transgenic mice via adjusting Bcl-2/Bax balance

Abstract

Introduction: The degeneration of memory-focused synapses play important roles in Alzheimer's disease (AD) pathogenesis, while it is not well known how β amyloid interferes neuron apoptosis and how a herbal combination GAPT influence synapse loss and neuronal apoptosis pathways of APP/PS1 transgenic mice.

Methods: Three-month and six-month APPswe/PS1dE9 transgenic mice were used. Spatial and memory ability were measured by Morris Water Maze, Neuron and synapse number were assessed by electron microscope; Aβ, Bcl-2/Bax were determined by immunohistochemistry and western blot.

Results: APP/PS1 mice not only had increased Aβ accumulation, impaired memory performance, less synapse number, and much more necrosed neurons, but also had significant reduction in the Bcl-2/Bax ratio. However, GAPT and donepezil showed improved memory performance, less Aβ accumulation, increased neuron and synapse number, as well as restored balance of Bcl-2/Bax.

Discussion: GAPT may improve cognitive functions via both reducing Aβ deposition and restoring Bcl-2/Bax balance of neuron.

Keywords: APP/PS1 transgenic mice; Alzheimer's disease; Aβ deposition; Bcl-2/Bax balance; GAPT; Neuron apoptosis.

Fig. 1

Flowchart of experimental protocols. Six-month-old mice were treated for 3 months, named as short-treatment-period group, and 3-month-old mice were treated for 6 months, named as long-treatment-period group.

Fig. 2

Spatial learning and memory tests of each group in the Morris water maze. (A) Normal control; (B) APP/PS1; (C) APP/PS1 + donepezil; (D) APP/PS1 + Gl; (E) APP/PS1 + Gm; and (F) APP/PS1 + Gh. The average escape latencies of normal control mice were significantly decreased from day 1 to day 5. The average escape latencies were significantly longer in APPswe/PS1dE9 transgenic mice compared with their age-matched normal control mice. Although mice treated with GAPT and donepezil all showed decrease in escape latencies on the fifth training day, of them APP/PS1 + Gm had a shortest escape latency. Abbreviations: Gh, high-dose group; Gl, low-dose group; Gm, medium-dose group; PS1, presenilin 1.

Fig. 3

Representative searching strategies. If the animal spent more than 70% of the time traveling within a specified distance of the median line, then the search strategy was defined as a linear pattern. The tendency pattern was defined similarly to the linear pattern except that the designated area was expanded to a distance of up to 50% of the radius of the swimming pool. For the peripheral pattern, the central point of the animal's travel area was first determined. With this point as the center of the circle and 75% of the radius of the swimming pool determined the circle's boundary. If the animal spent more than 70% of the time traveling outside this boundary, then the search pattern was defined as a peripheral pattern.

Fig. 4

Ultrastructure of neurons in each group. Pyramidal cell layer of the dorsal CA1 subfield of the hippocampus at a magnification of ×20,000 were taken by a JEOL transmission electron microscope. The normal neuron structure of hippocampus CA1 area (A) and the degenerated, shrank, and necrosed neuron (B and C). Their nucleuses were out of normal shape and chromatin pyknosed and cell organelles were swelling without clear structures. The relative normal neuron structure of hippocampus CA1 area (D, E, F). Their nucleuses were relatively normal compare to A or B.

Fig. 5

Ultrastructure of neuron synapses in each group in short-term treatment. The synapses were also taken by the JEOL transmission electron microscope. The synapse number of the model group is significantly lower than the normal control group. There were less synapses in B than in A, while there were more synapses in C, D, E than in B. And the synapse number in F was similar in B. From the table and bar chart, it is clear that the synapse numbers of Gl, Gm, and Gh of both STP and LTP groups and donepezil LTP groups are all significantly increased without dose-dependency. Abbreviations: Gh, high-dose group; Gl, low-dose group; Gm, medium-dose group; LTP, long treatment period; STP, short treatment period. ^▲▲Compare to normal group, P < .01; ^★compare to APP/PS1 group, P < .05; ^★★compare to APP/PS1 group, P < .01.

Fig. 6

Ultrastructure of neuron synapses in each group in long-term treatment. The synapses were also taken by the JEOL transmission electron microscope. There were less synapses in B than in A, while there were more synapses in C, D, E than in B. And the synapse number in F was similar in B. And the structure of synapses in B and F were also immature. From the table and bar chart, it is clear that the synapse number of the model group is significantly lower than the normal control group. The synapse numbers of Gl, Gm, and Gh of both STP and LTP groups and donepezil LTP groups are all significantly increased without dose-dependency. Abbreviations: Gh, high-dose group; Gl, low-dose group; Gm, medium-dose group; LTP, long treatment period; STP, short treatment period. ^▲▲Compare to normal group, P < .01; ^★★compare to APP/PS1 group, P < .01.

Fig. 7

Amyloid beta (Aβ) plaque accumulation detected by immunohistochemistry in each group. Aβ plaque accumulation as indicated by brown and yellow staining in the extracellular space. There are obviously more positively stained plaques in (B) compared with (A), whereas less in (C, D, E, and F).The color version of this figure is available in the online edition.

Fig. 8

Amyloid beta (Aβ) accumulation detected by Western blot in each group. Western blot analysis showed that significantly more total Aβ was expressed in the APP/PS1 group than in the normal control mice. When compared with the APP/PS1 group, total Aβ expression was lower in all other groups. ^▵▵Compare to normal group, P < .01; ^▴compare to APP/PS1 group, P < .05; ^▴▴compare to APP/PS1 group, P < .01.

Fig. 9

Bcl-2/Bax ratio detected by Western blot in each group. There was a significant decrease in the Bcl-2/Bax ratio in APP/PS1 mice whereas the Bcl-2/Bax ratio was significantly increased in GAPT-treated groups. ^▵▵Compare to normal group, P < .01; ^▴compare to APP/PS1 group, P < .05; ^▴▴compare to APP/PS1 group, P < .01.