Journal: Fluids and Barriers of the CNS
Article Title: Aquaporin 1 and the Na+/K+/2Cl− cotransporter 1 are present in the leptomeningeal vasculature of the adult rodent central nervous system
Figure Lengend Snippet: uDISCO clearance of the intact mouse head depicts the expression of aquaporin 1. a Mouse brain (P60) cleared by uDISCO and immunolabeled for AQP1 (AQP1int, green) reveals the vasculature network in the leptomeninges, including the middle cerebral arteries (MCA, arrows). AQP1 + cells also line the subarachnoid cisterns and the olfactory bulb. b Optical section reveals AQP1 + choroidal epithelial cells and olfactory ensheathing glia cells. c , d Higher magnification images of the areas depicted in b (blue and purple squares) showing AQP1 in the glomerular layer (arrow) and in choroidal epithelial cells (asterisk). e Representative micrograph of a parasagittal section of an adult mouse brain (P90) immunolabeled for AQP1 (AQP1ext, grey). AQP1ext + epithelial cells of the choroid plexus are observed in the fourth ( f ) and in the lateral ventricles ( g ). In contrast, olfactory ensheathing glia cells in the olfactory bulb are not immunolabeled ( h ). i Representative micrograph of a coronal section from adult mouse brain (P90) immunolabeled with AQP1 (AQP1int, grey). j Higher magnification of the depicted area in i (square) shows in detail AQP1int + epithelial cells in the choroid plexus of the lateral ventricles. k Olfactory ensheathing glia cells are also immunoreactive. Dashed line in k depicts the mitral cell layer. l Immunoblotting reveals a band of 35 kDa, corresponding to the glycosylated form of AQP1, detected in the BS, Cb, Ctx, Hip, Hyp and OB, obtained from young adult mice (P30). The non-glycosylated form of AQP1, corresponding to a band of 28 kDa, is detected in choroid plexi and kidney homogenates obtained from young adult mice (P30). The housekeeping protein GAPDH (37 kDa) was used as loading control. Control antigen confirms antibody-epitope specific binding. m Graphic shows the relative AQP1 protein levels, in relation to GAPDH. BS, brain stem; Cb, cerebellum; ChP, choroid plexus; Ctx, cerebral cortex; CPu, caudate putamen; EPL, external plexiform layer; Fi, fimbria; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; GL, glomerular layer; Hip, hippocampus; Hyp, hypothalamus; IC, internal capsule; IPL, internal plexiform layer; Kdy, kidney; LV, lateral ventricle; OB, olfactory bulb; PirCtx, piriform cortex, SCh, suprachiasmatic nuclei; Thal, thalamus; WM, white matter; 3V, third ventricle; 4V, fourth ventricle. Scale bars: a , b , e 1 mm; c , i 500 μm; d , 200 μm; f – h , j , k 50 μm
Article Snippet: The band corresponding to the housekeeping protein glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was detected by Western blot using a mouse anti-GAPDH antibody (Sigma-Aldrich, St. Louis, Missouri, USA), with an apparent molecular weight of 37 kDa, as previously described [ , ].
Techniques: Expressing, Immunolabeling, Mouse Assay, Binding Assay