Fig. 4

Quantification of FD40 from dorsal dura and electron microscopical imaging of meningeal efflux pathways. A, FD40 fluorescence measurement from homogenized dorsal dura after BPL. Unpaired t-test, *P < 0.05, n = 5–6 mice per group, filled circles represent individual mice. B, Schematic representation of brain cortex and dorsal dura indicating the regions in dura which were selected for EM analysis. White boxes indicate magnified regions. The superior sagittal sinus (SSS), the confluence of sinuses (COS). C A dural sample without HRP (negative control) displays no staining. Endothelial cells (EC), smooth muscle cells (SMC), the vascular basement membrane (asterisks). D, The reaction product of HRP, polymerized 3′3-diaminobenzidine (DAB), is shown as a black precipitate around the SSS. HRP is localized in the basement membrane (asterisks) and in the junctions (arrows) of ECs lining the SSS and SMCs. E, Higher magnification of an area in D, showing distribution of HRP in a junction of adjacent ECs. Similar observations from 5 different mice. F, Silver-enhanced nanogold-conjugated IgG (arrowheads) is localized in the region around the COS in B, in the dural tissue consisting of collagen (Col) fibers, and inside ECs of a meningeal lymphatic vessel (LV). G, Higher magnification of an area in F showing clusters of nanogold-conjugated IgG in lymphatic ECs and on the surface of the vessel lumen (white arrowheads). H, A blood capillary (C) in the region around the COS. I, Higher magnification of an area in H showing nanogold-conjugated IgG in the basement membrane of ECs. J, Nanogold-conjugated IgG is distributed in the region close to the SSS. K, Higher magnification of an area in J shows nanogold-conjugated IgG in the basement membrane between ECs and SMCs of the SSS. Similar observations from 8 different mice