Fig. 1: Fer2-expressing 0273 neurons drive reward seeking despite shock.

a, Left, experimental protocol. Starved wild-type flies were trained to associate an odour (the CS+) with sucrose. t, test period. Right, learned CS+ approach can be competed with in a time-dependent manner by presenting the CS+ with 90 V shock (n = 16). Groups on the far left and far right show 60 s tests of sucrose-trained flies without electrified CS+ and 60 s shock avoidance of mock-trained flies, respectively. b, Top left, schematic of DANs labelled by 0273-GAL4 (other labelled neurons are not shown) that project from the PAM cluster to horizontal lobe mushroom body compartments. Bottom left, experimental protocol. Right, starved transgenic flies trained with CsChr activation of 0273 neurons do not show a time-dependent increase in CS+/90 V avoidance (n = 12). c, Left, experimental protocol. US, unconditioned stimulus. Right, starved flies trained with 0273-neuron activation approach reward-predicting CS+ despite 90 V shock. Mock-trained and sucrose-trained flies exhibit shock avoidance (n = 10). Different letters above bars in a–c indicate groups that are significantly different from each other (P < 0.05; one-way ANOVA then Tukey’s honestly significant difference (HSD)). Data are mean ± s.e.m.; dots are individual data points that correspond to independent behavioural experiments. Exact statistical values and comparisons are presented in Supplementary Information. d, UMAP projections of scRNA-seq data show that neuron-driven CsChr expression (left) overlaps with Fer2 expression (right). e, Top left, CsChr-positive cells express Fer2 in both biological replicates (Rep 1 and Rep 2) whereas Fer2 expression is almost absent in the rest of the brain. Right, CsChr-expressing cells co-express marker genes for cholinergic (63.1% of all cells), GABAergic (18.4%), dopaminergic (16.7%) or glutamatergic (1.8%) neurons.