Fig. 1.

HSL is present in neurons and distributed throughout the mouse brain. A: Western blots of presynaptic (Pre), postsynaptic (Post), and extrasynaptic (Extra) fractions prepared from the mouse cortex show enrichment in SNAP25, PSD95, and synaptophysin, respectively. B: Relative to total protein extracts (TE), synaptosomes, presynaptic and postsynaptic fractions show immunoreactivity against HSL, a band of 84 kDa that is absent from protein extracts of the HSL^−/− mouse cortex (picture on the right). Protein loaded in gels was 30 μg for comparison of synaptic fractions and TE, and 40 μg for comparing TE from HSL^−/− and wild-type (WT) littermates. C: Quantitative analysis of Western blot immunoreactivity suggests presynaptic and mostly postsynaptic HSL enrichment. D and E: HSL-specific (blue circles) and nonspecific (orange crosses) DAG lipase activity in brain areas and adipose tissue (D) and respective immunoreactivity in Western blotting (E). Protein loaded for immunoblotting was 40 μg for brain samples and 15 μg for adipose tissue. F: Representative fluorescence micrographs of the mouse brain cortex immunolabeled for HSL (magenta), NeuN (green), and MAP2 (yellow). HSL immunoreactivity appears within NeuN-positive (filled arrowhead) and NeuN-negative (open arrowhead) cells. G: The absence of immunoreactivity in cortical slices from HSL^−/− mice. The scale bars in micrographs are 20 μm. Data in bar graphs are represented as individual data points and mean ± SD of n = 6 in C and n = 3 in D. Symbols over data points indicate significant differences between control and HSL^−/− mice (∗∗P < 0.01 and ∗∗∗P < 0.001) based on Fisher’s least significant difference post hoc comparison after ANOVA. FC, frontal cortex; MAP2, microtubule-associated protein 2; OC, occipital cortex; PSD95, postsynaptic density protein 95; PTC, parietotemporal cortex.