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. 2018 Dec 3;10:44. doi: 10.3389/fnsyn.2018.00044

Figure 4.

Figure 4

pHoenix acidifies glutmatergic and GABAergic SVs in intact synapses. (A,B) Images of bafilomycin-treated (>2 h, 1 μm) cultures expressing pHoenix in glutamatergic (A) or GABAergic (B) synapses at baseline, after activation of pHoenix with 550 nm LED for 120 s, and a time point 100 s after end of light stimulation. Image intensity scales for all panels were set to 0–1300 a.u. (A) or 0–700 a.u. (B). Arrows indicate particular ROIs. Scale bar = 10 μm. (C) Average fluorescence response normalized to baseline fluorescence in bafilomycin-treated cultures expressing pHoenix in glutamatergic (black; n = 13 images; three independent cultures) or GABAergic (blue; n = 16 images; three independent cultures) synapses or expressing only SypHy in glutamatergic (gray; n = 16 images; two independent cultures) or GABAergic (light blue; n = 15 images; two independent cultures) stimulated with 550 nm LED for 120 s. The SypHy-only images are included for control purposes and were performed in a separate set of experiments. (D) Comparison of pHoenix fluorescence normalized to baseline in bafilomycin-treated cultures at time points immediately after 120 s, 550 nm light stimulation and 100 s after completion of light stimulation in glutamatergic (black) and GABAergic (blue) synapses. Mean ± SEM. Significance determined by paired t-test. Glu ***p < 0.0001; GABA, ***p < 0.0001.