Characterization of Tauopathy in a Rat Model of Post-Stroke Dementia Combining Acute Infarct and Chronic Cerebral Hypoperfusion

Abstract

Post-stroke dementia (PSD) is a major neurodegenerative consequence of stroke. Tauopathy has been reported in diverse neurodegenerative diseases. We investigated the cognitive impairment and pathomechanism associated with tauopathy in a rat model of PSD by modeling acute ischemic stroke and underlying chronic cerebral hypoperfusion (CCH). We performed middle cerebral artery occlusion (MCAO) surgery in rats to mimic acute ischemic stroke, followed by bilateral common carotid artery occlusion (BCCAo) surgery to mimic CCH. We performed behavioral tests and focused on the characterization of tauopathy through histology. Parenchymal infiltration of cerebrospinal fluid (CSF) tracers after intracisternal injection was examined to evaluate glymphatic function. In an animal model of PSD, cognitive impairment was aggravated when BCCAo was combined with MCAO. Tauopathy, manifested by tau hyperphosphorylation, was prominent in the peri-infarct area when CCH was combined. Synergistic accentuation of tauopathy was evident in the white matter. Microtubules in the neuronal axon and myelin sheath showed partial colocalization with the hyperphosphorylated tau, whereas oligodendrocytes showed near-complete colocalization. Parenchymal infiltration of CSF tracers was attenuated in the PSD model. Our experimental results suggest a hypothesis that CCH may aggravate cognitive impairment and tau hyperphosphorylation in a rat model of PSD by interfering with tau clearance through the glymphatic system. Therapeutic strategies to improve the clearance of brain metabolic wastes, including tau, may be a promising approach to prevent PSD after stroke.

Keywords: animal model; chronic cerebral hypoperfusion; glymphatic system; post-stroke dementia; tauopathy.

Figure 1

(a) The time line of the experiment. (b) Allocation of rats into groups. (c) Quantification of the infarct volume and representative images of cresyl violet staining in MS and MB rats. n = 6 in both groups; MCAO, middle cerebral artery occlusion; BCCAo, bilateral common carotid artery occlusion; SS, sham + sham; SB, sham + BCCAo; MS, MCAO + sham; MB, MCAO + BCCAo; ns, not significant.

Figure 2

Exploration time and the discrimination index in the novel object recognition test (a,b) and the novel object location test (c,d). n = 6–8 per group; NOR, novel object recognition; NOL, novel object location; SS, sham + sham; SB, sham + BCCAo; MS, MCAO + sham; MB, MCAO + BCCAo; * p < 0.05 and ** p < 0.01 compared to SS; ## p < 0.01 using the t-test.

Figure 3

Tauopathy in the cortex. (a) Triple-label immunohistochemistry of tau hyperphosphorylation at the serine 396 residue (Tau pS396) in conjunction with oligodendrocytes (Olig2) and neurons (NeuN). Quantitative analysis of the signal intensities of Tau pS396 (b), the signal intensities of Olig2 (c), the number of Olig2-positive cells (d), the signal intensities of NeuN (e), and the number of NeuN-positive cells (f). n = 6 per group; scale bar = 20 μm; SS, sham + sham; SB, sham + BCCAo; MS, MCAO + sham; MB, MCAO + BCCAo; MCAO, middle cerebral artery occlusion; BCCAo, bilateral common carotid artery occlusion; c, contralateral; i, ipsilateral; SI, signal intensity; CN, cell number; * p < 0.05 and *** p < 0.001 compared to SS; # p < 0.05 and ### p < 0.001 compared to the ipsilateral MS.

Figure 4

Tauopathy in the corpus callosum. (a) Triple-label immunohistochemistry of tau hyperphosphorylation at serine 396 residue (Tau pS396) in conjunction with oligodendrocytes (Olig2) and neurons (NeuN). Quantitative analysis of the signal intensities of Tau pS396 (b), the signal intensities of Olig2 (c), and the number of Olig2-positive cells (d). n = 6 per group; scale bar = 20 μm; SS, sham + sham; SB, sham + BCCAo; MS, MCAO + sham; MB, MCAO + BCCAo; MCAO, middle cerebral artery occlusion; BCCAo, bilateral common carotid artery occlusion; c, contralateral; i, ipsilateral; SI, signal intensity; CN, cell number; * p < 0.05, ** p < 0.01 and *** p < 0.001 compared to SS; ## p < 0.01 and ### p < 0.001 compared to the ipsilateral MS.

Figure 5

Histological correlation of tauopathy in the peri-infarct area of MB rats. Double-label immunohistochemistry of tau hyperphosphorylation at serine 396 residue (Tau pS396) and various markers, including NeuN, glial fibrillary acidic protein (GFAP), myelin basic protein (MBP), microtubule associated protein 2 (MAP2), Olig2, and Oligodendrocyte. Signals of NeuN, GFAP, MBP, and MAP2 were captured in the peri-infarct cortex and Olig2 and Oligodendrocyte in the peri-infarct corpus callosum. scale bar = 20 μm; MB, MCAO + BCCAo.

Figure 6

(a) Parenchymal infiltration of cerebrospinal fluid tracer (CSF) with Texas Red^®-conjugated dextran of 3kDa (TR-d3) and its quantification in dorsal cortex (DC, b), ventral cortex (VC, c), cingulate gyrus (CG, d), and basal forebrain (BF, e). (f) Parenchymal infiltration of CSF tracer with fluorescein isothiocyanate (FITC)-conjugated dextran of 40kDa (FITC-d40) and its quantification in DC (g), VC (h), CG (i), and BF (j). n = 6 per group; scale bar = 1 mm; SS, sham + sham; SB, sham + BCCAo; MS, MCAO + sham; MB, MCAO + BCCAo; MCAO, middle cerebral artery occlusion; BCCAo, bilateral common carotid artery occlusion; c, contralateral; i, ipsilateral; * p < 0.05, ** p < 0.01, and *** p < 0.001 compared to SS.