Cholinergic degeneration and memory loss delayed by vitamin E in a Down syndrome mouse model - PubMed (original) (raw)

Cholinergic degeneration and memory loss delayed by vitamin E in a Down syndrome mouse model

Jason Lockrow et al. Exp Neurol. 2009 Apr.

Abstract

Down syndrome (DS) individuals develop several neuropathological hallmarks seen in Alzheimer's disease, including cognitive decline and the early loss of cholinergic markers in the basal forebrain. These deficits are replicated in the Ts65Dn mouse, which contains a partial trisomy of murine chromosome 16, the orthologous genetic segment to human chromosome 21. Oxidative stress levels are elevated early in DS, and may contribute to the neurodegeneration seen in these individuals. We evaluated oxidative stress in Ts65Dn mice, and assessed the efficacy of long-term antioxidant supplementation on memory and basal forebrain pathology. We report that oxidative stress was elevated in the adult Ts65Dn brain, and that supplementation with the antioxidant vitamin E effectively reduced these markers. Also, Ts65Dn mice receiving vitamin E exhibited improved performance on a spatial working memory task and showed an attenuation of cholinergic neuron pathology in the basal forebrain. This study provides evidence that vitamin E delays onset of cognitive and morphological abnormalities in a mouse model of DS, and may represent a safe and effective treatment early in the progression of DS neuropathology.

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Figures

Fig. 1

Treatment strategy. (A) Ts65Dn mice begin to show deficits in spatial and working memory tests between 4 and 6 months, while degeneration of basal forebrain cholinergic neurons is first evident at 6 months with decreased cell size, followed by a reduced cell number by 8–10 months. This study initiated vitamin E dietary supplementation at 4 months of age, prior to cognitive and morphological alterations in Ts65Dn mice, and mice were maintained on control or vitamin E-enriched diets through 2 weeks of behavioral testing in the radial arm maze. Mice were sacrificed between 8 and 10 months of age, immediately following behavioral testing. (B) Latency to find the platform (seconds) during the first and final (fifth) trials on the visible platform task. There were no significant differences in performance on the simple visible platform test.

Fig. 2

Vitamin E-treated Ts65Dn mice improve performance on RA maze. WMC (A), RM (B), and WMI (C) errors±SEM collapsed across 2-day trial blocks. Testing blocks were separated into an acquisition, or learning, phase (days 2–9) and an asymptotic, or maintenance, phase (days 10–15), as shown by the vertical dotted line. A 3D graph of WMI errors plotted across both days and trials showing normosomic (D), Ts65Dn control (E) and Ts65Dn Vitamin E (F). Normosomic and Ts65Dn Vitamin E groups exhibited learning on WMC, RM, and WMI, while the Ts65Dn control group only showed a modest error reduction on WMI. Each block was analyzed separately using a simple ANOVA, and group differences were represented by * (p<0.05). The Ts65Dn control group performed more WMC errors than normosomics and more WMI errors than normosomic and Ts65Dn vitamin E mice. There were no statistical differences between the normosomic and Ts65Dn Vit E groups in the asymptotic phase of testing (normosomic control, _n_=9; Ts65Dn control, _n_=7; Ts65Dn vitamin E, _n_=10).

Fig. 3

Working memory load. Trial by group interactions for the asymptotic phase (days 10–15) of testing are shown for WMC (A) and WMI (B), with the y-axis depicting mean errors per trial±SEM, averaged over the last three blocks of testing. Trial progression escalates the amount of information required for task completion, and therefore serves to model increases in memory load. Ts65Dn control mice committed more errors on trial 4, when working memory load was highest. This impairment in Ts65Dn memory load appears to be improved with vitamin E supplementation (*p<0.05, **p<0.01).

Fig. 4

Vitamin E reverses elevations in oxidative stress in Ts65Dn mice. (A) H2DCFDA assay of cortical tissue homogenates demonstrated a significant increase in cortical ROS levels in Ts65Dn controls over both normosomic and Ts65Dn vitamin E mice. Values are expressed as mean±SEM in µM DCF fluorescence per milligram protein. (B) Scattergram showing the significant correlation between cortical ROS levels and WMC errors in Trial 4 of the Asymptotic phase of testing for Ts65Dn control animals. Ts65Dn mice that made more WMC errors on the final trial of testing tended to have higher levels of ROS in the cortex (Ts65Dn control, _n_=7; ***p<0.001).

Fig. 5

Vitamin E prevented cholinergic degeneration in the medial septal nucleus (MSN) of Ts65Dn mice. Cholinergic neurons were identified by the high affinity NGF receptor TrkA immunostaining in 8–10-month mice. Normosomic animals demonstrated a strong TrkA immunoreactivity in the MSN (A), while Ts65Dn controls exhibited weaker staining as well as regional asymmetry of TrkA-positive neuron distribution in the MSN (B). These abnormalities were not seen in Ts65Dn vitamin E mice (C). Stereological cell counting showed that Ts65Dn mice exhibited neuronal loss (*p<0.05, +_p_=0.053) in the medial septal nucleus (D), while separate analysis of cohorts demonstrated the greatest cell loss in the 10-month-old cohort (F). Vitamin E-treated mice did not exhibit of BFCN compared with normosomic littermates. Cell area measurements also showed a significant reduction in TrkA-positive cell area in Ts65Dn controls (*p<0.05, ***p<0.001) compared to normosomic and Ts65Dn vitamin E mice (E), while cohort analysis demonstrated that vitamin E had the greatest effect on the 8-month-old cohort (G). Values are mean cell size±SEM (normosomic control, _n_=9; Ts65Dn control, _n_=7; Ts65Dn vitamin E, _n_=10). Scale bar=100 µm.

Fig. 6

APP metabolism is altered following vitamin E supplementation in Ts65Dn mice. Western blot depicting the levels of full length APP and its CTFs using O443 and secreted APP using 22C11 (A). Densitometric analysis of bands corresponding to APP-FL (B) and APP-CTFα (C) reveals that APP expression is elevated in both treated and untreated Ts65Dn mice (APP-FL: Normosomic=4.657±0.203; Ts65D_n_=5.787±0.159; Ts65Dn Vit E=5.981±0.135), while CTFα levels in Ts65Dn Vitamin E mice are statistically increased over their untreated counterparts (APP-CTF: Normosomic=0.899±0.070; Ts65D_n_=1.315±0.102; Ts65Dn Vit E=1.656±0.110). Secreted APP levels are increased in Ts65Dn and Ts65Dn Vitamin E mice, though the treated mice show a non-significant increase over Ts65Dn controls (D, sAPP: Normosomic=2.124±0.131; Ts65Dn=3.056±0.095; Ts65Dn Vit E=3.245±0.027). Ratio of APP-CTFα to APP-FL as a measurement for the relative metabolism of APP in prefrontal cortex tissues (E). Guinea pig brain (GPB) lysate (A) is a positive control for APP-FL. Data are mean±SEM, normalized by the total protein stain Ponceau S (_n_=4 per group).

Fig. 7

Vitamin E attenuated the loss of calbindin D-28k expression in the hippocampus of Ts65Dn mice. Immunohistochemical analysis of hippocampal neurons demonstrate robust Calbindin immunoreactivity in CA1 pyramidal neurons of normosomic animals (A). Ts65Dn control animals exhibit reduced staining at the cell body and apical dendrite (B), while vitamin E treatment partially recovers this loss (C). Densitometry revealed that Ts65Dn control mice had reduced CB-immunoreactivity in the CA1/CA2 region compared to normosomic controls (D, ***p<0.001). Ts65Dn vitamin E mice show a marginal increase in CB over Ts65Dn control (D, +_p_=0.07) mice, while a cohort analysis showed that this increase was most prominent at 8 months of age (E). Values are mean±SEM staining intensity normalized to background. Scale bar=100 µm.

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Cholinergic degeneration and memory loss delayed by vitamin E in a Down syndrome mouse model - PubMed (original) (raw)