Fig. 3.
PAM-β′1 subset of dopaminergic neurons is involved in forgetting. (A) PAM-β′1 neurons in the R94F11∩R24E12 split-Gal4 driver (R94F11-Gal4DBD; R24E12-p65AD). Throughout the figures, gray dashed lines indicate MB lobes. (B) Colocalization of PAM-β′1 with tyrosine hydroxylase, a dopamine marker. Arrowheads indicate PAM-β′1 somas visualized by a membrane-tagged reporter (myr::GFP) driven by R94F11∩R24E12. Representative images are single confocal sections. (C and D) Projection of the brain region including the MB lobes. NP2397 was expressed in putative PAM-DANs (arrowheads) whose processes innervated β′1, β′2, and γ4 (white dashed lines). The expression was substantially removed by R58E02-Gal80, which suppressed Gal4 activity in PAM-DANs. (E and F) Magnified views of the MB horizontal lobes. Intersection of NP2397 with R24E12-LexA and R48B04-LexA revealed PAM-DANs that innervated distinct lobe compartments (Materials and Methods). (G) Suppressing the expression of NP2397 in PAM-β′1 restored memory decay at 3 h. UAS-Kir2.1; Gal80ts flies were crossed to flies carrying NP2397 in combination with the indicated Gal80 transgene. Rescue of the 3-h memory increment was observed with R58E02-Gal80 (P = 0.54, n = 10, t test) and R24E12-Gal80 (P = 0.68, n = 8, t test) but not with TH-Gal80 (P = 0.03, n = 6, t test) and R48B04-Gal80 (P = 0.009, n = 12, t test). (H and I) VT28152 was selective for PAM-β′1, whereas R48B04 was mainly expressed in PAM-DANs innervating γ4, γ5, and β′2. (J) Kir2.1 inactivation of VT28152 neurons (P = 0.01, n = 6, t test) but not R48B04 neurons (P = 0.22, n = 9, t test) led to increased 3-h memory retention. Data in G and J are means ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001. [Scale bars, 50 μm (A), 10 μm (B), and 20 μm (C–F, H, and I).]