Bafilomycin A1 alkalinizes vesicular lumen, whereas Rose Bengal acidifies it, consistent with the actions of these agents on V-ATPase and VGLUTs, respectively. A–D, Astrocytes expressing synapto-pHluorin (raw data). The addition of bafilomycin A1 to the astrocytes at rest expressing synapto-pHluorin (A) causes an increase in pHluorin fluorescence (B, E). Quite the contrary, the addition of Rose Bengal to synapto-pHluorin-expressing astrocytes at rest (C) causes decrease of pHluorin fluorescence (D, E). The image in B is acquired after incubation with bafilomycin A1 (5 μm; 60 min), whereas the image in D is acquired after incubation with Rose Bengal (0.5 μm; 30 min). The pseudocolor scale is a linear representation of the fluorescence intensities ranging from 0 to 255. Changes in synapto-pHluorin fluorescence are expressed as dF/Fo (percentage) after background subtraction. Bars represent the means and SEMs. Concentrations (c) and exposure times (t) of pharmacological agents are given in micromolars and minutes, respectively.
