Chronic cerebral hypoperfusion and plasticity of the posterior cerebral artery following permanent bilateral common carotid artery occlusion

Abstract

Vascular dementia (VaD) is a group of heterogeneous diseases with the common feature of cerebral hypoperfusion. To identify key factors contributing to VaD pathophysiology, we performed a detailed comparison of Wistar and Sprague-Dawley (SD) rats subjected to permanent bilateral common carotid artery occlusion (BCCAo). Eight-week old male Wistar and SD rats underwent BCCAo, followed by a reference memory test using a five-radial arm maze with tactile cues. Continuous monitoring of cerebral blood flow (CBF) was performed with a laser Doppler perfusion imaging (LDPI) system. A separate cohort of animals was sacrificed for evaluation of the brain vasculature and white matter damage after BCCAo. We found reference memory impairment in Wistar rats, but not in SD rats. Moreover, our LDPI system revealed that Wistar rats had significant hypoperfusion in the brain region supplied by the posterior cerebral artery (PCA). Furthermore, Wistar rats showed more profound CBF reduction in the forebrain region than did SD rats. Post-mortem analysis of brain vasculature demonstrated greater PCA plasticity at all time points after BCCAo in Wistar rats. Finally, we confirmed white matter rarefaction that was only observed in Wistar rats. Our studies show a comprehensive and dynamic CBF status after BCCAo in Wistar rats in addition to severe PCA dolichoectasia, which correlated well with white matter lesion and memory decline.

Keywords: Bilateral common carotid artery occlusion; Chronic cerebral hypoperfusion; Dolichoectasia; Sprague-Dawley strain; Vascular dementia; Wistar strain.

Fig. 1. Reference memory function after bilateral common carotid artery occlusion (BCCAo) assessed using a five-radial arm maze with tactile cues.

(A) Time line showing the experimental design. Before undergoing BCCAo, SD and Wistar rats were trained to remember the arms with a food pellet. From the 8^th day post-BCCAo, the ability of SD and Wistar rats to find the food-placed arms was tested over 10 days. (B) Representative image of different tactile cues applied for radial arm maze. (C) Representative image of a five-radial arm maze apparatus. The yellow box refers to the location of the food pellet, which is indicated as a red star. (D) Graph showing the number of errors during the memory test period. Wistar rats subjected to BCCAo (n=6) demonstrated a significantly higher number of incorrect choices compared with sham-treated Wistar (n=9), sham-operated SD (n=9), and BCCAo-treated SD (n=8) rats. Data are given as mean±SEM. ^*p<0.05 by repeated measures analysis of variance with Duncan's post hoc mean comparison. SD, Sprague–Dawley; SD sham, SD rats subjected to sham operation; SD BCCAo, SD rats subjected to BCCAO; Wistar sham, Wistar rats subjected to sham operation; Wistar BCCAo, Wistar rats subjected to BCCAo.

Fig. 2. Longitudinal whole-brain monitoring of cerebral blood flow (CBF) after bilateral common carotid artery occlusion (BCCAo) in SD and Wistar rats.

(A) Representative color-coded examples of serial laser Doppler perfusion images. Cortical microperfusion of Wistar and SD rats subjected to sham or BCCAo was repetitively monitored using a laser Doppler perfusion imaging system. (B) Graph showing CBF perturbation in MCA territory, indicated as two circles in the right image. After BCCAo, Wistar rats (n=7) and SD rats (n=9) showed a marked CBF reduction compared with sham-treated Wistar (n=7) and SD (n=8) rats. Notably, cerebral hypoperfusion in MCA territory was significantly greater in Wistar rats than SD rats. Data are given as mean±SEM. ^*p<0.05 compared to sham and ^†p<0.05 compared to Wistar rats subjected to BCCAo. Repeated measures analysis of variance with Duncan's post hoc mean comparison was performed. (C) Graph showing CBF perturbation in PCA territory, indicated as a circle in the right image. Wistar rats after BCCAo (n=7) showed marked CBF reduction compared to sham-treated Wistar (n=7), sham-treated SD (n=8), and BCCAo-operated SD (n=9) rats. Data are given as mean±SEM. ^*p<0.05 by repeated measures analysis of variance with Duncan's post hoc mean comparison. SD, Sprague–Dawley; SD sham, SD rats subjected to sham operation; SD BCCAo, SD rats subjected to BCCAO; Wistar sham, Wistar rats subjected to sham operation; Wistar BCCAo, Wistar rats subjected to BCCAo; MCA, middle cerebral artery territory; PCA, posterior cerebral artery.

Fig. 3. Structural plasticity of the brain vasculature at the circle of Willis in SD and Wistar rats after bilateral common carotid artery occlusion (BCCAo).

(A) Representative images of major blood vessels at 7 days (SD: n=7, Wistar: n=9), 14 days (SD: n=7, Wistar: n=6), and 21 days (SD: n=7, Wistar: n=8) after BCCAo or sham operation (SD: n=12, Wistar: n=7). (B) Table showing the ratio of the posterior communicating artery (PcomA) and posterior cerebral artery (PCA) diameter to that of the basilar artery in SD and Wistar rats. The relative PcomA and PCA were significantly thinner in Wistar rats than in SD rats. One-way analysis of variance also demonstrated that PCA plasticity was significantly increased at all time points after BCCAo compared with sham values for both SD and Wistar strains. Values are mean±SEM. ^†p<0.05: SD sham vs. Wistar sham; ^*p<0.05: sham vs. the rest of the time points assessed by one-way analysis of variance and Duncan's post hoc test. (C) Graph showing the fold change of relative PCA plasticity based on the sham-controls. Wistar rats showed significantly greater PCA plasticity than SD rats at all time points examined. ^*p<0.05 by two-tailed Student's t test. Values are mean±SEM. SD, Sprague–Dawley; MCA, middle cerebral artery; ACA, anterior cerebral artery; ICA, internal carotid artery; BA, basilar artery.

Fig. 4. White matter rarefaction in SD and Wistar rats.

(A) Representative microscopic images of the optic tract stained by Klüver-Barrera Luxol fast blue. Only Wistar rats subjected to bilateral common carotid artery occlusion (BCCAo) showed prominent vacuolation and serpentine white matter fibers, indicative of white matter damage. Scale bar in SD sham=50 µm (valid for all the other images). (B) Graph showing the severity of white matter lesion. Semi-quantitative analysis of white matter rarefaction showed a significantly higher grade of white matter damage after BCCAo in Wistar rats (n=7) compared with SD rats (n=8). Data are given as mean±SEM. ^*p<0.05 by Pearson Chi-square test. SD, Sprague–Dawley.