Fig. 6. GABA and DA suppress the excitability of dFSB neurons in a temperature-sensitive manner.

An ATP-gated cation channel P2X₂ was expressed in dFSB neurons along with a synaptic calcium sensor sytGCaMP (23E10 > P2X₂, sytGCaMP) to quantify ATP-induced changes in intracellular Ca²⁺ levels as an indirect readout of the neural excitability. Transgenic flies were pre-entrained in LD cycles at 21 °C (blue) or 29 °C (orange). Whole brains were dissected out, transferred to an imaging chamber, and equilibrated with HL3 buffer for 5 minutes. Where indicated, dissected brains were incubated with 50 mM GABA (a) or 10 mM DA (b) for 5 minutes prior to the batch application of 5 mM ATP (shaded by gray boxes). A time series of the fluorescence images was recorded using a photoactivated localization microscopy and their quantification was performed using ZEN software. Data represent average ± SEM (n = 9–14). n.s., not significant; *P < 0.05 as determined by Aligned ranks transformation ANOVA, Wilcoxon rank-sum test.