Acute leakage patterns of fluorescent plasma flow markers after transient focal cerebral ischemia suggest large openings in blood-brain barrier

Abstract

Objective: This study tested the hypothesis that blood-brain barrier (BBB) opening during acute reperfusion permits the passage of smaller macromolecules but not larger ones and that this molecular size restriction disappears over time. Methods: Following 3 hours (h) of unilateral middle cerebral artery occlusion and either 3 or 21 h of reperfusion, Wistar rats (n = 42) were injected with Evans blue (EB, a fluorescent dye that binds instantly to plasma albumin yielding EB-tagged albumin, EB-Alb) and with one of three fluorescent dextrans ranging in size from 77- to 2000-kDa. During occlusion and reperfusion, ischemic status of the affected tissue was confirmed by magnetic resonance imaging (MRI). Blood-to-brain transfer of the dextrans relative to that of EB-Alb was examined by fluorescence microscopy within three regions with ischemic damage. Results: Increase in EB-Alb leakage from 3 to 21 h of reperfusion was significant (from 40-60% to 80-90% of fields examined; p < 0.05). Co-leakage of the largest dextran used 2000-kDa, with EB-Alb was observed in only 40% of the fields at 3+3 h, but nearly in all at 3 + 21 h (p < 0.01). Parenchymal distribution of the tracers differed among the fields and included considerable cellular uptake of EB-Alb and some of dextrans. Conclusions: Supporting the hypothesis, opening of the BBB was insufficient to allow passage of the largest dextran at 3 + 3 h in about 40% of the microvascular networks viewed. The number of total leaky microvascular beds increased by nearly 50% between 3 + 3 h and 3 + 21 h.