Figure 5. Ovary-projecting OA neurons control the GSC increase.

(A, C–D, F–G) Frequencies of germaria containing 1, 2, and 3 GSCs (left vertical axis) and the average number of GSCs per germarium (right vertical axis) in virgin (V) and mated (M) female flies. The number of germaria analyzed is indicated inside the bars. (A) RNAi of Tdc2 and TβH by Tdc2-GAL4. OA was added into the standard food. (B) A schematic drawing of Drosophila central nervous system and the ovary-projecting OA neurons with the dsx⁺ OA neurons projecting to the ovary. (C) Tdc2 RNAi in dsx⁺ Tdc2⁺ neurons with the genotype indicated. (D–E) TrpA1-mediated activation of dsx⁺ Tdc2⁺ neurons. 17°C and 29°C were used as the permissive and restrictive temperatures, respectively, of TrpA1 channel. (D) GSC number. (E) The ratio of pH3⁺ GSCs and total GSCs. (F) The activation of Tdc2⁺ neurons with Oamb^Δ genetic background. (G) The inactivation of dsx⁺ Tdc2⁺ neurons. (H) Illustration showing the location of three clusters of Tdc2⁺ neurons in the caudal part of the abdominal ganglion (I–K, I’–K’). Negative images of TRIC labeling (anti-GFP) in the abdominal ganglions of virgin (I–K) and mated females (I’–K’) of TRIC (Tdc2 >UAS-mCD8::RFP, UAS-p65AD::CaM LexAop2-mCD8::GFP; nSyb-MKII::nlsLexADBDo;UAS-p65AD::CaM) flies, indicating intracellular Ca²⁺ transients. Scale bars, 20 μm. (L) The GFP intensities from the Tdc2⁺ median cluster, Tdc2⁺ dorsal cluster, and dsx⁺ Tdc2⁺ cluster of TRIC females show Ca²⁺ activity in virgin (gray) and mated females (red). Wilcoxon rank sum test was used for A, C, D, F, G, and L. Fisher’s exact test with Holm’s correction was used for E. ***p≤0.001, **p≤0.01, and *p≤0.05; NS, non-significant (p>0.05). All source data are available in Source data 1 and 3.

Figure 5—figure supplement 1. dsx⁺ Tdc2⁺ neurons control GSC increase.

(A–B) Frequencies of germaria containing 1, 2, and 3 GSCs (left vertical axis) and the average number of GSCs per germarium (right vertical axis) in virgin (V) and mated (M) female flies using nSyb-GAL4 (A) and Tdc2-GAL4 (B). (C–D) Images of dsx⁺ Tdc2⁺ neurons expressing the UAS > stop >mCD8::GFP reporter under Tdc2-GAL4 with dsx-FLP. GFP expression was detected only in the abdominal ganglion neurons projecting to the ovary. Images of the abdominal ganglion and the reproductive system are shown in C and D, respectively. (E–G) Immunofluorescence of dsx⁺ Tdc2⁺ neurons expressing UAS >mCD8::GFP reporter under Tdc2-GAL4 with tub >FRT >GAL80>FRT. GFP expression was only observed in the anti-Tdc2 positive neurons in the abdominal ganglion that projected to the ovary (E). Scale bars, 100 µm in C, D, E, and G; 10 µm in F. Wilcoxon rank sum test with Holm’s correction was used. ***p≤0.001; NS, non-significant (p>0.05). All source data are available in Source data 1.