(A, A'). CaMKII::YFP single-molecule fluorescence in situ hybridization (smFISH) in γ5β'2a MBON dendrites and soma (R66C08-GAL4>UAS-myr::SNAP). Images are maximum intensity projections of ten 0.2 μm z-sections. (B, B'). CaMKII::YFP smFISH in γ1pedc>α/β MBON dendrites and soma (MB112C-GAL4>UAS-myr::SNAP). Nuclear transcription foci are indicated (red arrowheads). Images are maximum intensity projections of ten 0.2 µm z-sections. (C) CaMKII::YFP smFISH signal/background in transcriptionally active γ5β'2a somata. Transcription foci are readily distinguished as the brightest puncta in the soma/nucleus (red data points). Note that only one transcription focus can be visualized per cell since the CaMKII::YFP allele is heterozygous. (D) Schematic of aversive training and control protocols followed by smFISH. The yellow and red circles represent the two odors. (E) CaMKII::YFP mRNA numbers in γ5β'2a MBON dendrites increase 10 min after odor–shock pairing, relative to control groups (one-way ANOVA: untrained-10 min p=0.001; odor only-10 min p=0.016; shock only-10 min p=0.002), and decrease to baseline by 2 hr (one-way ANOVA: 10 min-2 h p<0.001; 1–2 h p=0.004). CaMKII::YFP mRNA numbers in γ5β'2a MBON somata increase 1 hr after odor–shock pairing, relative to untrained (one-way ANOVA: p=0.001), odor only (one-way ANOVA: p=0.002), and 10 min post training (one-way ANOVA: p=0.025). The proportion of transcriptionally active γ5β'2a MBON somata is unchanged (X2=2.064, df = 5, p=0.840). (F) CaMKII::YFP mRNA numbers are not changed by aversive odor–shock pairing in γ1pedc>α/β MBON dendrites (one-way ANOVA: f = 1.473, p=0.212), their somata (one-way ANOVA: f = 2.183, p=0.067), and there is no detected change in CaMKII::YFP transcription (X2=3.723, df = 5, p=0.59). (G) Signal/background ratio of CaMKII::YFP transcription foci in γ5β'2a MBON somata. (H) Signal/background ratio of CaMKII::YFP mRNA localized in γ5β'2a MBON dendrites. Asterisks denote significant difference (p<0.05). Data are means ± standard error of mean. Individual data points are displayed.