Memantine improves cognition and reduces Alzheimer's-like neuropathology in transgenic mice

Abstract

Memantine is an N-methyl-d-aspartate receptor antagonist that is approved for the treatment of moderate to severe Alzheimer's disease (AD). In this study, three groups of triple-transgenic (3xTg-AD) mice with differing levels of AD-like pathology (6, 9, and 15 months of age) were treated for 3 months with doses of memantine equivalent to those used in humans. After the treatment, memantine-treated mice had restored cognition and significantly reduced the levels of insoluble amyloid-beta (Abeta), Abeta dodecamers (Abeta*56), prefibrillar soluble oligomers, and fibrillar oligomers. The effects on pathology were stronger in older, more impaired animals. Memantine treatment also was associated with a decline in the levels of total tau and hyperphosphorylated tau. Finally, memantine pre-incubation prevented Abeta-induced inhibition of long-term potentiation in hippocampal slices of cognitively normal mice. These results suggest that the effects of memantine treatment on AD brain include disease modification and prevention of synaptic dysfunction.

Figure 1

Memantine improves spatial cognitive deficits in 3xTg-AD mice. Memantine-treated 3xTg-AD mice were evaluated using a spatial reference paradigm and compared with sucrose-treated 3xTg-AD and NonTg mice. Mice were trained on the spatial reference version of the Morris water maze task before and after treatment. All three groups (6, 9, and 12 months old) of memantine-treated 3xTg-AD mice showed impairment compared with NTg mice during the pretreatment training phase (A, C, and E), which largely disappeared after 3 months of memantine treatment (A, C, and E). Similarly, all 3xTg-AD mice showed both short (1.5 hours) and long-term (24 hours) memory deficits during the pretreatment retention trials (B, D, and F), but the mice with mild and moderate pathology demonstrated a significant improvement after treatment (B and D). **P < 0.05, memantine-treated 3xTg-AD mice versus sucrose-treated NTg mice. *P < 0.05, memantine-treated 3xTg-AD mice versus sucrose-treated 3xTg-AD mice.

Figure 2

Memantine reduces accumulation and phosphorylation of tau in 3xTg-AD mice may be mediated by GSK3β. After 3 months of memantine treatment, total levels of the soluble fraction of tau protein in 3xTg-AD mice with severe pathology (A and B) were significantly lower, compared with the sucrose-treated animals, using HT7 antibody. Memantine treatment was also associated with significantly lower levels of tau phosphorylated at residues 212/214 (AT100), 199/202 (AT8), 212/214 (AT100), and 231 (AT180) (A and B). Levels of total tau (HT7) and phosphorylated tau at 199/202 residues had no significantly changes (C and D). Immunostaining using AT100 antibody revealed a significantly reduction in somatodendritic tau phosphorylated at residues 212/214 (E). Western blot analysis shows that the levels of inactive GSK3β (phospho-GSK3β-S9) were increased by memantine treatment in 3xTg-AD, compared with sucrose-treated mice (F and G). Levels of p25 and PP2A do not show differences with memantine treatment in mice with severe pathology (F and G). *P < 0.05, memantine-treated 3xTg-AD mice versus sucrose-treated 3xTg-AD mice. S, sucrose-treated; M, memantine-treated.

Figure 3

Memantine treatment alters levels of soluble and insoluble Aβ. Following 3 months of treatment, levels of soluble and insoluble Aβ_1-40 and Aβ_1-42 on homogenates of the whole brain hemisphere of 3xTg-AD mice were determined using ELISA and reported in pg of Aβ/mg of wet tissue weight. In the youngest group of animals, memantine treatment was associated with significantly lower levels of soluble Aβ_1-40 and Aβ_1-42 (A) and insoluble Aβ_1-40 (B). In the moderate pathology group, memantine-treated animals had significantly lower levels of soluble Aβ_1-40, but significantly higher levels of soluble Aβ_1-42, compared with their sucrose-treated counterparts (E). In this age group, memantine had no apparent effect on the levels of insoluble Aβ_1-40 and Aβ_1-42 (F). In the most severe pathology group, memantine treatment was associated with significantly higher levels of soluble Aβ_1-42 (I), but with significantly lower levels of insoluble Aβ_1-40 and Aβ_1-42 (J). Immunohistological staining of the subiculum of the mild and moderate pathology groups showed a modest reduction in accumulation of Aβ in memantine-treated 3xTg-AD mice, compared with their sucrose-treated counterparts (C, D, G, and H), using 6E10 antibody. However, in the severe pathology group the immunostaining using both antibodies 6E10 and D3 (antiAβ_1-42) showed a robust effect of the memantine reducing the accumulation of the Aβ in subiculum (K–N). *P < 0.05, memantine-treated 3xTg-AD mice versus sucrose-treated 3xTg-AD mice.

Figure 4

Memantine reduces oligomer formation in the 3xTg-AD mice. Western blot analysis of homogenates of whole brain hemisphere revealed no difference in the levels of APP, C99 fragment, C83 fragment, ADAM10, and BACE between the memantine- and sucrose-treated 3xTg-AD mice, using 6E10, CTF20, anti-ADAM10, anti-β-site APP cleaving enzyme (BACE) antibodies respectively, (A, B, E, F). However, the levels of the Aβ dodecamer (Aβ*56) (using 6E10 antibody) were significantly lower in memantine-treated animals (A and B). In addition, dot blot analysis indicated a significant reduction in the levels of all prefibrillar oligomers (A11 antibody) in memantine-treated mice (male and female), and lower levels of fibrillar oligomers (OC antibody) in female animals only (C and D). *P < 0.05, memantine-treated 3xTg-AD mice versus sucrose-treated 3xTg-AD mice.

Figure 5

Memantine prevents Aβ oligomer induced inhibition of LTP. Application of Aβ (42 nmol/L) for 90 minutes blocked LTP in slices of hippocampal CA1. Induction of LTP (arrow) and maintenance for 1 hour. Representative traces of single experiments are shown before (control) and 1 hour after tetanization (100 Hz/1 second). Memantine restored LTP in the presence of Aβ. There was no difference in the fEPSP potentiation after 1 hour of LTP induction when slice were incubated for at least 6 hours with memantine (1 μmol/L) under control conditions (not shown) and after wash-in of Aβ. Representative traces of single experiments are shown before (control) and 1 hour after HFS. Data represent the summary of recordings from slices of 10 animals.