SUC and ATP evoke overlaying Ca2+ transients in NAc astrocytes. A1: Confocal image of SUC-responsive cells (green and red circled yellow spots) participated in ATP-stimulated concerted Ca2+ burst (green spots marked by white arrows). Red arrows show directions of SUC (vertical) and ATP (horizontal with x) applications. Scale bar is 50 μm. A2: Representative (dF/F0)max plots showing effects of 50 μM SUC alone and with 100 μM ATP puff in A1. Red and green traces correspond to the SUC-responsive cells circled in A1 while gray traces show the fluorescence changes of cells marked by white arrows in A2. B: Confocal images showing fluorescence of 5 μM Fluo-4 AM loaded cells in NAc slices during control (left image), 50 μM SUC application (middle images) and washout (right image). Time refers to the fluorescence-time plot (C). Scale bar is 50 μm. C: (dF/F0)max plots showing Ca2+ transients in response to 50 μM SUC in the NAc slice. Each coloured trace corresponds to a Ca2+ transient circled in (B). D: Comparison of cell number- and fluorescence intensity-based summary plots of the effects of SUC on Ca2+ fluorescence in rat NAc slices. *p < 0.05 and **p < 0.01 compared to control, +p < 0.05 and ++p < 0.01 compared to 50 μM (Mann-Whitney with Bonferroni post hoc tests; ). E: SUC-responsive repetitive Ca2+ transients co-localized with astroglial markers. The NAc slice showing SUC-responsive repetitive Ca2+ transients was subsequently double-immunostained for astroglial marker proteins GFAP (green, left) and Cx43 (red, middle) while the observation chamber kept continuously on the stage of the confocal microscope. Our post-Calcium protocol (cf. Methods section) showed yellow co-localisation of Cx43 and GFAP proteins (right) with a SUC-responsive cell shown in A. Scale bar is 5 μm.