Quantitative assessments of cerebral vascular damage with a silicon rubber casting method in photochemically-induced thrombotic stroke rat models

Previous studies have described microvascular disturbances downstream of occluded large vessels arising during the acute phase (several hours) following cerebral ischemic insult. Prolonged microvascular disturbances may cause delayed neuronal cell death in ischemic penumbral regions, leading to expanded brain infarctions and poor neurological and functional outcomes. The lack of simple and quantitative methods for investigating this microcirculation failure suggests the need to develop a new method for clarifying the precise distribution and persistence of post-ischemic microvascular disturbances. The present study used a silicone rubber casting method in quantitative analyses of microvascular conditions in photochemically-induced thromboembolic (PIT) stroke rat models. After the casting procedure in rats with PIT stroke, a 6 microm-thick coronal section was obtained, and quantitative analyses of microvascular density and measurements of the infarct area in the serial section were performed. The major findings of the present study are as follows: (1) Silicone rubber casting techniques can be applied to precise quantitative analyses of microvessels in the same individual in whom brain infarct volume was measured; (2) the persistence and spatial distribution of microvascular disturbances assessed at the ischemic core, ischemic penumbra, and non-ischemic regions strongly suggest that microvascular disturbances affect brain infarct expansion; (3) the current method demonstrated the protective effects of MK-801 on microvessels, indicating that the technique may be useful in investigating factors that provide vascular protection. The experimental procedure introduced here would facilitate future evaluations of vascular protective agents.