(A) Representative photomicrographs of the hindbrain showing cell bodies (arrowheads) and vessels (arrows) labeled with fluorescent leptin (1ug per embryo, white labeling) 5 and 45 min after it was intracerebroventricularly injected into E12 embryos. (B) Relative expression of Leprb mRNA in various brain areas and peripheral organs of E12 WT mouse embryos (n = 3 per group). (C) Relative expression of Leprb mRNA in the hindbrain of E10, E12 and E14 WT mouse embryos (n = 3–5 per age). Numbers in bar graphs indicate Ct values. (D) Confocal images of VAChT-immunoreactive cells (green fluorescence) and leptin-induced pSTAT3-immunopositive cells (red fluorescence) in the DMV of E12 WT embryos. (E) Confocal images and quantification of the overall density of VAChT-positive fibers in organotypic cultures of isolated E12 hindbrains. The cultures were incubated with leptin (100ng/ml) or vehicle for 48h (n = 5–6 explants per group). DMV, dorsomedial nucleus of the vagus nerve; HIND, hindbrain; V4, fourth ventricle. Scale bar, 100 um. ***P < 0.001 versus vehicle (E); and ****P < 0.0001 between indicated groups (D). Values are shown as mean the ± SEM.