Fig. 6. Evidence that scaffold axons are required for glia migration and direct glia along specific pathways. (A,A') Wild-type scaffold axon fascicles (anti-ß-gal; grayscale) in a wg-lacZ late third instar optic lobe project in a stereotyped fashion toward glial destinations. MNG and Xi glia (anti-Repo; red) are seen following along fascicles towards their appropriate destinations. A higher magnification view of the yellow boxed area in A is shown in A'. (B,B') In ds¹ homozygous animals, scaffold axon fascicles project aberrantly, and the distribution of glia is correspondingly aberrant. A higher magnification image of the yellow-boxed region of B is shown in B'; glia (yellow arrowheads) on an abnormal trajectory are closely associated with the misprojecting scaffold axon fascicle (anti-ß-gal; grayscale). (C,C') Scaffold axon projections are likewise aberrant in ds^33k homozygous animals. In this specimen, dorsal scaffold axon fascicles bifurcate onto aberrant trajectories, and glia are likewise misdirected. The higher magnification image of the boxed region of C shown in C' reveals mispositioned glia (anti-Repo; red) in association with the misrouted scaffold axon fascicles (anti-ß-gal staining in grayscale). (D,D') Random somatic clones were generated expressing both a GFP marker and an activated Ras protein (Ras1^N17) that inhibits axon extension. wg-lacZ (anti-ß-gal staining; blue) was used to mark the migratory axon scaffold. In the late third instar optic lobe shown, a clone encompasses part of the dorsal Wg domain (white outline in D and yellow outline in D'). Glia (anti-Repo, red), which do not express the UAS-Ras1^N17 transgene, stalled at the edge of this Wg domain (yellow arrowheads in D,D'), much as they do in `eyeless' mutant strains (see Fig. 5). Scaffold axon fascicles cannot be detected emanating from the dorsal Wg domain, but are clearly visible extending from the ventral Wg domain (yellow arrows); ventral glia migrate normally. (E,E') A specimen like that shown in D,D' in which somatic clones (white outline in E, yellow outline in E') express UAS-Ras1^N17 (labeled by co-expression of GFP, green in E). Clones of particular interest encompass both the dorsal and ventral Wg domains. Scaffold axons are absent from both and glia stall at both the dorsal and ventral margins of the Wg domains (yellow arrowheads in E'). Scale bars: in A, 20 µm (for A-D,D',E,E'); in A', 8 µm (for A',B',C').