Pioglitazone improves reversal learning and exerts mixed cerebrovascular effects in a mouse model of Alzheimer's disease with combined amyloid-β and cerebrovascular pathology - PubMed (original) (raw)

Pioglitazone improves reversal learning and exerts mixed cerebrovascular effects in a mouse model of Alzheimer's disease with combined amyloid-β and cerebrovascular pathology

Panayiota Papadopoulos et al. PLoS One. 2013.

Abstract

Animal models of Alzheimer's disease (AD) are invaluable in dissecting the pathogenic mechanisms and assessing the efficacy of potential new therapies. Here, we used the peroxisome proliferator-activated receptor gamma agonist pioglitazone in an attempt to rescue the pathogenic phenotype in adult (12 months) and aged (>18 months) bitransgenic A/T mice that overexpress a mutated human amyloid precursor protein (APPSwe,Ind) and a constitutively active form of transforming growth factor-β1 (TGF-β1). A/T mice recapitulate the AD-related cognitive deficits, amyloid beta (Aβ) and cerebrovascular pathologies, as well as the altered metabolic and vascular coupling responses to increased neuronal activity. Pioglitazone normalized neurometabolic and neurovascular coupling responses to sensory stimulation, and reduced cortical astroglial and hippocampal microglial activation in both age groups. Spatial learning and memory deficits in the Morris water maze were not rescued by pioglitazone, but reversal learning was improved in the adult cohort notwithstanding a progressing Aβ pathology. While pioglitazone preserved the constitutive nitric oxide synthesis in the vessel wall, it unexpectedly failed to restore cerebrovascular reactivity in A/T mice and even exacerbated the dilatory deficits. These data demonstrate pioglitazone's efficacy on selective AD hallmarks in a complex AD mouse model of comorbid amyloidosis and cerebrovascular pathology. They further suggest a potential benefit of pioglitazone in managing neuroinflammation, cerebral perfusion and glucose metabolism in AD patients devoid of cerebrovascular pathology.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1. Pioglitazone (pio) restored stimulus-evoked CBF and CGU in A/T mice.

A, The impaired hyperemic response to whisker stimulation in adult and aged A/T mice was rescued by pioglitazone compared to age-matched WT controls, as measured by LDF (n=4 mice/group) (traces of adult mice shown, green: WT; turquoise: WTpio; violet: A/T and pink: A/Tpio). Values represent the percent increase in CBF induced by whisker stimulation relative to baseline. B, Pioglitazone also improved the decreased CGU response to whisker stimulation in the somatosensory cortex of aged A/T compared to WT mice (n=3-6 mice/group). The % activation ratio denotes the percentage of corrected standard uptake value (SUV) in the activated contralateral (left side (L)) vs. ipsilateral (right side (R)) somatosensory cortex. Error bars represent SEM. ★p<0.05, ★★p<0.01, ★★★p<0.001 using two-way ANOVA followed by Newman-Keuls post-hoc test.

Figure 2. Effects of pioglitazone (pio) on spatial learning and memory in adult A/T mice.

A, A/T mice (▼) displayed impaired learning during hidden-platform testing compared to aged-matched wild-type (WT) littermates (●) in the Morris watermaze. These deficits were not due to visual or motor disabilities as all groups had comparable abilities to find the visible platform (days 1-3). Pioglitazone did not improve the deficit in A/T mice (▽) and did not affect the performance of WT controls in the hidden platform training (days 4-8). (○).B, A/T mice, treated (▽) and non-treated (▼) with pioglitazone displayed significant deficit in memory retention as assessed during the probe trial in the target quadrant. C, When the probe trial was subdivided in 15s segments for better understanding of the pattern of behavior throughout the 60s-long probe trial in target quadrant, pioglitazone did not exert any benefit in A/T mice. D, A/T mice showed an inclination for the original quadrant during the probe trial, as shown by the increased time spent in this quadrant and smaller average distance to the platform location. E, Patterns of behaviour in original quadrant segmented in 4 slices of 15s each during the probe trial showed that pioglitazone had a positive effect in A/T mice (▽) in increasing percent time spent and average distance to platform during in the last quarter of the probe trial. F, Pie-representation of typical swimming patterns of % time spent and % distance traveled in target quadrant (red, #4) compared to the original quadrant (yellow, #2) during the probe trial. The drawings (on the left) are representations of single mouse swimming pattern best matching the average of of each group, as displayed by the pie patterns (on the right). Error bars represent SEM (n=6-9 mice/group). ★p<0.05 ★★, ** p<0.01, ★★★p<0.001 when compared to untreated WT controls (★) or A/T mice (*) using two-way ANOVA and repeated measures ANOVA followed by Newman-Keuls post-hoc test.

Figure 3. Effects of pioglitazone (pio) on cerebrovascular reactivity and protein alterations in adult A/T mice.

A, The impaired cerebrovascular dilatations to acetylcholine (ACh) and calcitonin gene-related peptide (CGRP) in A/T mice (▼) were aggravated or reversed to weak constrictions by pioglitazone (▽). This was accompanied by a full recovery of baseline NO synthesis in adult A/T mice measured during NOS inhibition (L-NNA, 10-5M). Dilatations induced by the NO donor SNP were unimpaired in A/T mice, but were reversed to small constrictions or shifted to the right at high SNP concentrations in pioglitazone-treated A/T mice. Contractile responses to 5-HT and ET-1 were unaltered in A/T mice and pioglitazone had no detrimental effects. Error bars represent SEM (n=3-6 mice/group). B, Vasodilator synthesizing enzyme COX-2 and proteins associated with vascular fibrosis VEGF, CTGF and MMP-9 were not significantly altered by genotype or treatment, as measured by Western blot in pial vessels of adult A/T mice relative to their treated counterparts and WT mice. Actin was used as a reference for loading (n=4 mice/group). Similar results were obtained in the aged cohort, but data were not illustrated for clarity purposes. Error bars represent SEM. ★p<0.05, ★★, ** p<0.01, ★★★, *** p<0.001 when compared to untreated WT controls (★) or A/T mice (*) using two-way ANOVA (part A) and repeated measures ANOVA (part B) followed by Newman-Keuls post-hoc test.

Figure 4. Pioglitazone (pio) had no effect on amyloidosis in A/T mice.

A, Levels of soluble and insoluble Aβ1-40 and Aβ1-42 in adult A/T mice, as assayed in cortex and hippocampus by ELISA, were comparable between pioglitazone-treated- and untreated A/T mice. B, Western blot analysis with 6E10 antibody confirmed no effect of pioglitazone on soluble Aβ species in cortex of adult A/T mice. On the gel, (★) represents the APP band and (*), the 4kDa band of monomeric Aβ including Aβ1-40 and Aβ1-42, as quantified in the bar graph. C, Thioflavin S-stained dense-core Aβ plaque load in cortex and hippocampus were unaltered by pioglitazone in adult A/T mice. D, Similarly, 6E10-immunostaining of diffuse and dense-core Aβ plaques in 5µm-thick paraffin sections showed no effect of pioglitazone on cortical and hippocampal Aβ plaque load in adult A/T mice. Error bars represent SEM. ★p<0.05, ★★p<0.01, ★★★p<0.001 for comparison to A/T mice using two-way ANOVA followed by Newman-Keuls post-hoc test (n=4 mice/group).

Figure 5. Pioglitazone (pio) reduced cortical astroglial activation.

A, In the cortex (but not in the hippocampus) of aged A/T mice, pioglitazone significantly reduced the increase in GFAP-immunoreactive (IR) surface area relative to WT controls (n=4 mice/group). B, GFAP analysis of both the aged and adult groups rendered the same results of a selective cortical effect of pioglitazone on astroglial activation. Scale bar = 300 µm and 100 µm (insert). Error bars represent SEM. ★p<0.05, ★★p<0.01, ★★★p<0.001 using two-way ANOVA followed by Newman-Keuls post-hoc test.

Figure 6. Pioglitazone (pio) reduced hippocampal microglial activation.

A, Pioglitazone significantly reduced the increased surface area of Iba-1-immunolabelled microglial cells in the hippocampus of both aged and adult A/T mice compared to WT controls, as shown in the aged group. B, Both the area occupied by and staining intensity of Iba-1-positive microglial cells in the hippocampus were reduced by pioglitazone at both ages. No effect was seen in the cortex (n=4 mice/group). Scale bar = 100 µm and 25 µm (insert). Error bars represent SEM. ★p<0.05, ★★p<0.01 using two-way ANOVA followed by Newman-Keuls post-hoc test.

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Pioglitazone improves reversal learning and exerts mixed cerebrovascular effects in a mouse model of Alzheimer's disease with combined amyloid-β and cerebrovascular pathology - PubMed (original) (raw)