(A) Quantification of inhibition of Ca2=-responses for ACh + GABA (left) or ACh + glutamate (right) compared to ACh. LNvs from larvae with Pdf-Gal4 and UAS-G-CaMP and either no other transgenes (Control, black bars, 89% inhibition), UAS-GABAB-R2-RNAi on Chr. III (Pdf > GABAB-R2-RNAi, light gray, 91% inhibition, p>.5 vs Control) or UAS-GABAB-R3-RNAi (Pdf > GABAB-R3-RNAi, dark gray, 19.5% inhibition, p<.001 vs Control).
(B) Adult head RNA levels of GABAB-R2 and Pdf were measured as in Figure 2 from either UAS-GABAB-R2-RNAi heterozygotes (GABAB-R2-RNAi / +) or flies with elav-Gal4 expressing UAS-GABAB-R2-RNAi (elav > GABAB-R2-RNAi). GABAB-R2 RNA levels were lower in elav > GABAB-R2-RNAi flies (p<0.01).
(C) GABAB-R3 and Pdf RNA levels measured by qPCR on amplified RNA from adult LNvs isolated from Pdf-RFP; Pdf-Gal4 / + controls or flies which also had UAS-GABAB-R3-RNAi (PdfRFP; Pdf > GABAB-R3-RNAi). GABAB-R3 RNA levels were reduced in Pdf-RFP; Pdf > GABABR3-RNAi flies (p<0.02). Pdf RNA levels were ~2.5-fold higher with GABAB-R3-RNAi flies and so the extent of knockdown with this RNAi transgene (>400-fold) may be over-estimated.
(D) Representative actograms of adult flies expressing either GABAB-R2-RNAi (Pdf > GABABR2-RNAi, left) or GABAB-R3-RNAi (Pdf > GABAB-R3-RNAi, right) in LNvs. Pdf > GABAB-R3-RNAi flies have longer period rhythms than Pdf > GABAB-R2-RNAi flies (24.9hr vs 24.3hr, p <.001).