Figure 1.
Arousal effect of caffeine was abolished in BG–A2AR KO mice. A–C, Typical sections from the Rosa26–Dlx5/6–Cre reporter mouse were stained with mouse polyclonal antibodies against β-gal to visualize Cre-expressing neurons indirectly. A, Robust expression of β-gal is seen in the striatum of the reporter mouse. B, At a single-cell level, double immunofluorescence for β-gal (green) and ChAT (magenta) on an adjacent section to A shows that cholinergic interneurons in the striatum also express β-gal. The arrows in the top and bottom of B indicate neurons with dual immunolabeling for β-gal and ChAT. C, In the classical arousal/sleep-related cell groups of the basal forebrain and anterior hypothalamus, i.e., the nucleus of the HDB, SI, or VLPO, only moderate immunoreactivity for β-gal is detected in the HDB. β-gal immunolabeling is absent in neurons of the SI and VLPO. Scale bars: A, 500 μm; B, 20 μm; C, 250 μm. D, E, The BG–A2AR KO mice and WT littermates were treated with vehicle or caffeine (2 or 30 mg/kg, i.p.). Time course (D) and total time (E) of wakefulness during the first 3 h after caffeine injection (arrows) were assessed with EEG/EMG recordings. Data are presented as the mean ± SEM (n = 4–5). *p < 0.05, **p < 0.01 compared with vehicle treatment within corresponding genotype. ##p < 0.01 compared with corresponding WT littermates. ††p < 0.01 compared between caffeine doses.