CNS staining for ionized-calcium binding protein 1 (Iba1) and lysosomal-associated membrane protein1 (LAMP-1).

IHC staining of the cerebellum for Iba1 (A-C) and LAMP-1 (D-F). (A) Molecular layer of cerebellum in 12-month-old WT mice showing nonreactive resident microglia characterized by fine cytoplasmic extensions. (B) In 12-month-old Gnptab^−/− mice, there are numerous enlarged Iba1/MAC2 positive macrophages in the molecular layers associated with Purkinje cell loss. (C) In 12-month-old Gnptg^−/− mice, there was only mild activation of microglia in the molecular layer. (D) LAMP-1 in 12-month-old WT cerebellum is limited primarily to the Purkinje cell layer. (E) In contrast, LAMP-1 labeling in 12-month-old Gnptab^−/− mice is prominent in the enlarged macrophages/microglia in the molecular layer but reduced in the areas of Purkinje cell loss. (F) There was a slightly increased amount of LAMP-1 staining in the Purkinje cell and molecular layers of 12-month-old Gnptg^−/− mice, suggesting a subclinical increase in lysosomal storage in these areas. In (G-I) similar genotype-related differences in LAMP-1 staining are shown in spinal cord white tracts in 12-month-old WT, Gnptab^−/− and Gnptg^−/− mice, respectively. In all genotypes LAMP-1 was detected within oligodendrocytes, but the extent of labeling was markedly increased in Gnptab^−/− mice (H) due to increased volume of cytoplasm and extensions of hypertrophic oligodendrocytes, as well as reactive microglia and macrophages. (I) In 12-month-old Gnptg^−/− mice, LAMP-1-positive oligodendrocytes are also larger and more prominent than in WT mice, but microglia/macrophages are uncommon. (J-L) Endothelial cells in meningeal veins in WT mice were consistently LAMP-1 negative (J), but microvesiculated endothelium of meningeal veins (not arteries) in both Gnptab^−/− (K) and Gnptg^−/− (L) mice was LAMP-1 positive, although markedly more so in the Gnptab^−/− mice. (Figs. A - I at 40X,Bar  =  100 mm; Figs. J - L at 60X, Bar  =  50 mm)