Figure 5.

Gliosis surrounding blood vessels with perivascular CM-DiI⁺ macrophages and primary findings of erythrocyte tracking experiments. (a) Representative images of cortical blood vessels (highlighted with the dotted line), stained with GFAP, a specific marker of astrocytes. Increased GFAP expression (green) is associated with inflammation and gliosis. No significant gliosis was observed surrounding CM-DiI^- vessels. In contrast, we observed a large increase in the perivascular GFAP expression surround blood vessels labelled with perivascular CM-DiI⁺ cells (red). Scale bar represents 30 µm. (b) Quantification of perivascular gliosis. CM-DiI⁺ vessels were compared to CM-DiI^- vessels in the same animal for both 2 and 5 days after SAH. ***P < 0.0001, *P < 0.05, paired t-test (n = 4 independent samples/group). (c) Schematic of the primary findings in the erythrocyte tracing studies. At 10 minutes, free intact erythrocytes were found in the basal cisterns and ventricles. Intact erythrocytes were also found in the perivascular spaces of apical, basal and subcortical arterioles. At 2 days after SAH, there was decreased blood and erythrocytes in the basal cistern. Blood in the ventricles was phagocytosed by discrete cells. Erythrocyte uptake was observed by perivascular and meningeal macrophages (in purple) but not microglia. Perivascular gliosis was observed primarily around arterioles with perivascular macrophages that engaged in erythrocyte clearance (swollen endfeet, in blue). At 5 days after SAH, little or no blood was found in basal cisterns. Blood in the ventricles were contained by discrete cells. Some turnover of the perivascular macrophages and more perivascular inflammation were observed. Overall, we found that perivascular and meningeal macrophages are involved in erythrocyte clearance, and contribute to perivascular inflammation and neuronal cell death after SAH. Note that erythrocytes, macrophages and astrocytic endfeet are not drawn to scale.