Inhibition of calpains improves memory and synaptic transmission in a mouse model of Alzheimer disease
Fabrizio Trinchese, Mauro Fa’, Shumin Liu, Hong Zhang, Ariel Hidalgo, Stephen D. Schmidt, Hisako Yamaguchi, Narihiko Yoshii, Paul M. Mathews, Ralph A. Nixon, Ottavio Arancio
Fabrizio Trinchese, Mauro Fa’, Shumin Liu, Hong Zhang, Ariel Hidalgo, Stephen D. Schmidt, Hisako Yamaguchi, Narihiko Yoshii, Paul M. Mathews, Ralph A. Nixon, Ottavio Arancio
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Research Article Neuroscience

Inhibition of calpains improves memory and synaptic transmission in a mouse model of Alzheimer disease

Abstract

Calpains are calcium-dependent enzymes that determine the fate of proteins through regulated proteolytic activity. Calpains have been linked to the modulation of memory and are key to the pathogenesis of Alzheimer disease (AD). When abnormally activated, calpains can also initiate degradation of proteins essential for neuronal survival. Here we show that calpain inhibition through E64, a cysteine protease inhibitor, and the highly specific calpain inhibitor BDA-410 restored normal synaptic function both in hippocampal cultures and in hippocampal slices from the APP/PS1 mouse, an animal model of AD. Calpain inhibition also improved spatial-working memory and associative fear memory in APP/PS1 mice. These beneficial effects of the calpain inhibitors were associated with restoration of normal phosphorylation levels of the transcription factor CREB and involved redistribution of the synaptic protein synapsin I. Thus, calpain inhibition may prove useful in the alleviation of memory loss in AD.

Authors

Fabrizio Trinchese, Mauro Fa’, Shumin Liu, Hong Zhang, Ariel Hidalgo, Stephen D. Schmidt, Hisako Yamaguchi, Narihiko Yoshii, Paul M. Mathews, Ralph A. Nixon, Ottavio Arancio

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Figure 5

Calpain inhibition reestablished the increase in CREB phosphorylation during synaptic plasticity in APP/PS1 mice and produced a normal distribution of the synaptic protein synapsin I.

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Working hypothesis on how calpain inhibitors (Calp-inh) may be able to i...
(A) Western blot for pCREB at Ser-133 from cultured hippocampal neurons after 5-minute glutamate treatment. Five-day-old cultures were either treated with vehicle, E64, or BDA-410 for 3 days prior to applying glutamate. (B) Quantitative western blot analysis of data shown in A. pCREB levels in vehicle-treated WT cultures were increased after glutamate (n = 11; P < 0.01). APP/PS1 cultures failed to present the pCREB increase (n = 7; P > 0.01 with 1-way ANOVA). However, both E64 and BDA-410 reestablished normal pCREB values (n = 5 and 8, respectively). E64 and BDA-410 did not affect phosphorylation in WT cultures (n = 5 and 7, respectively). All samples were normalized against α-tubulin. (C) Examples of hippocampal slices stained with a pCREB antibody and fixed 60 minutes after tetanus in WT and APP/PS1 animals treated for 5 months from 8 weeks of age with E64, BDA-410, or vehicle. Lower-power (original magnification, ×4) view of the entire slice (left and middle panels). Higher-power (original magnification, ×16) view of CA1 cell pyramidal area (right panels). (D) Plot showing blockade of CA1-pCREB increase after tetanus in APP/PS1 slices (n = 6 both for WT and APP/PS1 slices; P < 0.01), whereas treatment with both E64 and BDA-410 for 5 months from 8 weeks of age reestablished the tetanus-induced pCREB increase (n = 6 for both; P < 0.01 compared with tetanized slices from vehicle-treated APP/PS1 mice). Both E64 and BDA-410 had no effect on WT mice after tetanus (n = 6 for both). In the absence of theta-burst, neither E64 nor BDA-410 induced changes in WT or APP/PS1 mice (n = 4 for each group; data not shown). IF, immunofluorescence. (E) The number of synapsin I–immunoreactive puncta was increased in vehicle-treated APP/PS1 cultures (n = 6) compared with vehicle-treated WT cultures (n = 6; P < 0.01). However, the basal synapsin I–immunoreactive puncta number was normal after exposure to E64 (n = 7) or BDA-410 (n = 5). The inhibitors did not affect synapsin I immunoreactivity in WT cultures (E64, n = 6; BDA-410, n = 7; P < 0.01 for both compared with vehicle-treated APP/PS1 cultures). (F) Glutamate failed to increase synapsin I–immunoreactive puncta numbers in vehicle-treated APP/PS1 cultures (n = 8) compared with vehicle-treated WT cultures (n = 7, P < 0.01), whereas both E64 and BDA-410 reestablished the glutamate-induced immunoreactivity increase in transgenic cultures (E64, n = 9; BDA-410, n = 9; P < 0.01 for both compared with vehicle-treated APP/PS1 cultures). Both E64 and BDA-410 had no effect on glutamate-induced immunoreactivity increase in WT cultures (E64, n = 9; BDA-410, n = 8).