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Fig. 8 Inhibition of gamma-secretase and the timing of Notch signaling in pronephric development. All whole-mount in situ hybridization was performed at 34 hpf. (A-C) The wild-type pattern of shippo1 expression (A) is dramatically expanded when gamma-secretase is inhibited with 100 μM DAPT starting at 9.5 hpf (B) but not when DAPT is added later, at 24 hpf (C). (D-F) Conversely, the number of shippo1-positive cells seen in control embryos (D) is dramatically reduced in the pronephros of embryos induced to ectopically express the Notch1a intracellular domain under the control of HS-GAL4 induction (nicd hs) at either 8 hpf (E) or 9 hpf (F). (G-I) trpM7 expression is lost when DAPT is added at 9.5 hpf (H), but not when it is added later, at 24 hpf (I). (J-L) slc13a1 expression is lost when DAPT is added at 9.5 hpf (K), and also when DAPT is added later, at 24 hpf (L).