Cilostazol prevents amyloid β peptide(25-35)-induced memory impairment and oxidative stress in mice

Abstract

Background and purpose: Cilostazol may be effective in dementia associated with a cerebral ischaemia. In this study, we examined whether it exerts beneficial effects on learning and/or memory impairment induced by Aβ(25-35) in mice, and compared its effects with those of aspirin.

Experimental approach: Aβ(25-35) (9 nmol) was administered to mice i.c.v. Learning and memory behaviour were evaluated by measuring spontaneous alternation in a Y-maze and a step-down type passive avoidance test, on the 5th and 8th days after injection respectively. Levels of lipid peroxidation (malondialdehyde) and cytokines in the frontal cortex and hippocampus were measured 2, 3, 5 and 7 days after the Aβ(25-35) injection. The effects of repeated administration of cilostazol and aspirin (both at 30 and 100 mg·kg(-1), p.o.) on any changes induced by Aβ(25-35) were evaluated.

Key results: Repeated administration of cilostazol significantly attenuated the impairment of spontaneous alternation and the shortened step-down latency induced by Aβ(25-35) . Aspirin did not show any beneficial effect. A significant increase in the levels of malondialdehyde (MDA) and IL-1β (only measured in hippocampus) was observed 2, 3 and 5 days after the Aβ(25-35) injection in the frontal cortex and hippocampus. Repeated administration of cilostazol (100 mg·kg(-1)) completely prevented the increase in MDA levels but failed to antagonize the increase in the expression of IL-1β induced by Aβ(25-35).

Conclusions and implications: These results suggest that the protective effect of cilostazol on Aβ(25-35)-induced memory impairment may be related to oxidative stress in the frontal cortex and the hippocampus.

Figure 1

Experimental protocols used in this study. CMC, carboxymethyl cellulose sodium salt.

Figure 2

Effects of repeated administration of cilostazol (A) and aspirin (B) on Aβ_25-35-induced impairment of spontaneous alternation in the Y-maze test. Aβ_25-35 (9 nmol per mouse, i.c.v.) was injected 5 days before the Y-maze test. Mice were treated with cilostazol (30 and 100 mg·kg⁻¹, p.o.) or aspirin (30 and 100 mg·kg⁻¹, p.o.) once a day for 5 days. On the 5th day, these drugs were injected again 60 min before testing. Data are shown as median (vertical column) and first and third quartiles (vertical line). The number of mice used is shown in parentheses. Significant levels; *P < 0.05, **P < 0.01 versus sham control (Mann–Whitney's U-test). #P < 0.05, ##P < 0.01 versus Aβ_25-35 alone (Bonferroni's test). CMC, carboxymethyl cellulose sodium salt.

Figure 3

Effects of repeated administration of cilostazol (A) and aspirin (B) on Aβ_25-35-induced impairment of learning and memory in a passive avoidance test. Aβ_25-35 (9 nmol per mouse, i.c.v.) was injected 7 days before the training trial. Mice were treated with cilostazol (30 and 100 mg·kg⁻¹, p.o.) or aspirin (30 and 100 mg·kg⁻¹, p.o.) once a day for 7 days. On the 7th day, these drugs were injected again 60 min before retention testing. A retention trial was carried out 24 h after the training trial. Data are shown as median (horizontal bar) and first and third quartiles (vertical column). The number of mice used is shown in parentheses. Significant levels; *P < 0.05, **P < 0.01 versus sham control (Mann–Whitney's U-test), #P < 0.05, ##P < 0.01 versus Aβ_25-35 alone (Bonferroni's test). CMC, carboxymethyl cellulose sodium salt.

Figure 4

Effects of acute administration of cilostazol on Aβ_25-35-induced impairment of spontaneous alternation (A) and learning and memory in a passive avoidance test (B). Mice were injected with Aβ_25-35 (9 nmol per mouse, i.c.v.) 5 days before the training trial for the Y-maze test. Cilostazol (30 and 100 mg·kg⁻¹, p.o.) was injected 60 min before the Y-maze test and the training trial of the passive avoidance test. The retention trial was carried out 24 h after the training trial. Data are shown as the median (vertical column) and first and third quartiles (vertical line) for the Y-maze and median (horizontal bar), and first and third quartiles (vertical column) for the passive avoidance tests. The number of mice in each experiment is shown in parentheses. Significant levels; **P < 0.01 versus sham control (Mann–Whitney's U-test). CMC, carboxymethyl cellulose sodium salt.

Figure 5

Effects of repeated pre-administration of cilostazol on Aβ_25-35-induced impairment of spontaneous alternation (A) and learning and memory in a passive avoidance test (B). Mice were treated with cilostazol (100 mg·kg⁻¹, p.o.) repeatedly for 8 days beginning 8 days before Aβ_25-35 (9 nmol per mouse, i.c.v.) injection. The Y-maze test was carried out 5 days after (A) Aβ_25-35 injection, and the passive avoidance test was carried out 7–8 days after Aβ_25-35 injection (B). The retention trial was carried out 24 h after the training trial. Data are shown as the median (vertical column) and first and third quartiles (vertical line) for the Y-maze, and median (horizontal bar) and first and third quartiles (vertical column) for the passive avoidance tests. The number of mice used is shown in parentheses. Significant levels; **P < 0.01 versus sham control (Mann–Whitney's U-test). CMC, carboxymethyl cellulose sodium salt.

Figure 6

Effects of repeated administration of cilostazol on Aβ_25-35-induced impairment of spontaneous alternation (A) and learning and memory in a passive avoidance test (B). Mice were injected with Aβ_25-35 (9 nmol per mouse, i.c.v.) 7 days before the Y-maze test or 9 days before the passive avoidance test. Mice were treated with cilostazol (100 mg·kg⁻¹, p.o.) repeatedly for 5 days beginning 2 days after Aβ_25-35 (9 nmol per mouse, i.c.v.) injection. Cilostazol was administered 60 min before testing on the Y-maze test day. On the next day, cilostazol was injected. On the day of the training trial for the passive avoidance test, cilostazol was administered 60 min before testing. A retention trial was carried out 24 h after the training trial. Data are shown as the median (vertical column) and first and third quartiles (vertical line) for the Y-maze, and median (horizontal bar) and first and third quartiles (vertical column) for the passive avoidance tests. The number of mice in each experiment is shown in parentheses. Significant levels; *P < 0.05, **P < 0.01 versus sham control, #P < 0.05 versus Aβ_25-35 alone (Mann–Whitney's U-test). CMC, carboxymethyl cellulose sodium salt.

Figure 7

Effects of cilostazol on Aβ_25-35-induced increase in lipid peroxidation in the mouse frontal cortex (A) and hippocampus (B). Aβ_25-35 (9 nmol per mouse) was injected (i.c.v.) and the mice were killed 2, 3, 5 and/or 7 days later. Mice were treated with cilostazol (100 mg·kg⁻¹, p.o.) 60 min before being killed (acute) or repeatedly (3, 4, 6 or 8 times). On the 1st day, cilostazol was administered 60 min before Aβ_25-35. Lipid peroxidation levels were assessed by the thiobarbituric acid method. Each lipid peroxidation level was normalized per unit protein. Values represent means ± SEM (n = 6–9). Significant levels; *P < 0.05, **P < 0.01 versus sham control (Student's t-test), ##P < 0.01 versus Aβ_25-35 alone (Dunnett's test). MDA, malondialdehyde.

Figure 8

Effects of cilostazol on the levels of pro-inflammatory cytokines in mouse hippocampus after Aβ_25-35 injection. Aβ_25-35 (9 nmol per mouse) was injected (i.c.v.) and the mice were killed 2, 3 and/or 5 days later. Mice were treated with cilostazol (100 mg·kg⁻¹, p.o.) 60 min prior to death (acute) or repeatedly (3, 4 or 6 times). On the 1st day, cilostazol was administered 60 min before Aβ_25-35. Pro-inflammatory cytokine levels were assessed by the suspension array method. Each cytokine level was normalized per unit protein. Values represent means ± SEM (n = 4–6). Significant levels; *P < 0.05 versus corresponding sham control (Student's t-test). GM-CSF, granulocyte macrophage colony-stimulating factor.