Fig. 5. In vitro culture in the absence of ecdysone reveals the step at which neurogenesis is blocked. (A) Disc from an early wandering A101 larva cultured in vitro for 24 hours in the presence of 20E shows the differentiation of the normal complement of neurons and outgrowth of their axons (22C10). (B) Disc from an early wandering larva cultured in vitro for 24 hours without ecdysone. Only the early born SOPs are visible with the A101 reporter line. Insets in A and B show higher magnification of the L3-2 region. (C-C'') Disc from an early wandering A101 larva cultured in vitro without ecdysone for 24 hours. The SOPs are recognizable with A101-ß-gal (C) but SENS fails to accumulate in them (C''). (C') merged image of A101 and SENS expression. (D-E) Changing pattern of SENS expression in the margin from two broad bands in the early wandering stage (D) to the accumulation in the SOPs by the mid-late wandering stage (E). (F) Block of SENS accumulation requires a functional ecdysone receptor: wing disc from an early wandering larva expressing UAS-IR-EcR under the control of dpp-GAL4 cultured in vitro in the absence of ecdysone for 24 hours. SENS only accumulates in the margin (arrow) where it is intersected by the dpp-domain (compare with control in C''). (C-F) Area shown is the same as in Fig. 2D. In all panels, anterior is to the top.