The Ddc-positive neurons are necessary for normal memory performance. (A) When the UASGAL4-RNAi-white transgene was expressed with Ddc-GAL4 driver, flies' memory performance was strongly reduced compared with all control genotypes [H (4, n = 643) = 28.3, P < 0.0001]. This finding was in contrast to TH-GAL4-driven UASGAL4-RNAi-white expression, where no deficits were found. Significant differences after multiple comparisons are presented, Ddc-GAL4/UASGAL4-RNAi-white with UASGAL4-RNAi-white/+ and Ddc/+ (**, P < 0.01). (B) Tests for the ability of flies with different genotypes to sense and avoid a 41°C temperature source did not find significant differences between genotypes [H (4, n = 382) = 3.37, P = 0.50]. (C) Blocking synaptic transmission by expressing the TeTxLC reveals a necessary role of the Ddc-positive neurons in memory formation. Flies with TeTxLC expression in the Ddc-positive and TH-positive neurons, and genetic controls, were trained in the heat box and tested for memory. Only flies expressing TeTxLC with the Ddc-GAL4 driver had a deficit in conditioned memory performance [H (4, n = 459) = 50.5, P < 0.0001, multiple comparisons indicate significant differences between Ddc-GAL4/UASGAL4-TeTxLC and both Ddc-GAL4/+ and UASGAL4-TeTxLC/+ performances (**, P < 0.01)]. (D) Tests for the ability to sense and avoid a 41°C high-temperature source in the thermosensitivity assay found a difference between these genotypes [H (4, n = 359) = 13.9, P < 0.01], which was caused by a difference between TH-GAL4/UASGAL4-TeTxLC and Ddc-GAL4/UASGAL4-TeTxLC flies (P < 0.05). The values represent means, and error bars are SEMs.