Fig. 1
The macrophage GABAergic system and GABA levels during infection. a Representative GABA-induced whole-cell currents in BMDMs (GABA, 300 µM). Bottom, the effects by GABAAR antagonist bicuculline (BIC, 50 µM). Right, quantification of GABA-induced currents at each GABA concentration. b, c BMDMs were infected with Mtb-ERFP (MOI of 5) for the indicated times and stained with GABA (green) and DAPI (nuclei; blue). b Cells were visualized by confocal microscopy. Scale bars, 15 μm. c Average mean fluorescence intensities (MFIs) of GABA. d–f Mice were infected i.v. with Mtb (1 × 106 CFU), and monitored at 14 dpi. Lung tissues were stained with GABA (red) and DAPI (nuclei; blue). d Representative in vivo imaging data from multiple lesions of Mtb-infected or -uninfected lungs (n = 8 per group). Scale bars, 50 μm (top) and 20 μm (bottom). e Average MFIs of GABA staining (n = 4 per group). f GABA in serum was measured by ELISA (n = 8 per group). g, h BMDMs were infected with Mtb (MOI of 5) or BCG (MOI of 5) for the indicated times. g GAD65 expression analyzed by flow cytometry. The units of the horizontal axis are the fluorescence (FITC) intensity. Representative gating strategy is shown in Supplementary Fig. 5. h Average MFIs of GAD65. i In BMDMs, 11 GABAAR subunit mRNAs were determined using the ∆∆Ct method, and normalized to the reference gene Gapdh. j BMDMs were infected with Mtb (MOI of 5) for the indicated times and GABAAR subunit expression was determined using qRT-PCR. k GABAAR subunit expression in lung tissues was determined using qRT-PCR (n = 6 mice for no infection, n = 7 mice for Mtb infection) as in d–f. *p < 0.05, **p < 0.01, and ***p < 0.001. ns: not significant, UI: uninfected. Statistical significance was determined by one-way ANOVA (c, h, j), Mann–Whitney U-test (f, k), or two-tailed unpaired t-test (e). Data shown (means ± SEM) represent the combined results of triplicates from three independent experiments (c, h, i, j), or of 15 (a), 2 (f), or 3 (b–e, g) independent experiments. Each symbol represents one animal (k)
