Figure 2.

Ultrastructural analysis of vacuoles and myelin. A, Semithin sections of the sciatic nerve show no difference between the genotypes at 14 months of age (14m). B, Western blot analysis detects the ClC-2 protein in brain but not in sciatic nerve (sciatic n.). KO lysates showed the specificity of the ClC-2 antibody. Actin served as a loading control. C–E, Electron micrographs showing typical vacuoles in white matter tracts of ClC-2 KO cerebellum. These vacuoles (asterisks) contained aberrant myelin sheets (arrows) and were surrounded by thin myelin sheaths (arrowheads). a, Axon. D, Myelinated axon surrounded by a vacuole and a normal-appearing oligodendrocyte (n). F, Part of WT white matter cerebellum. G, The structure of myelin sheaths in the area of best compaction revealed no difference in myelin compaction. Distances between MDLs were unchanged in the KO [10.7 ± 0.1 nm (WT) vs 10.8 ± 0.1 nm (KO); n < 90, 3 animals each]. 7w, 7 weeks old. H, Semithin sections of the optic nerve of 7-week-old (7w) WT and Clcn2 ^−/− mice revealed no vacuolation in the KO (asterisks indicate capillary). Higher magnification is shown in supplemental Fig. S3 (available at www.jneurosci.org as supplemental material). Scale bars: A, 20 μm; C–F, 1 μm; G, 20 nm; H, 10 μm.