Fig. 3. The dynamics of dendritic pruning. Projections of z-stacks taken from movies of a wild-type pupae (C161-GAL4>CD8::GFP). (A) Overview of Movie 2 (see supplementary material). At 14 h 10 min APF as the arbors of ddaD and ddaE are pruning, fragments and blebs of GFP are found close to the arbor (arrowheads). Filopodia extend on ddaD's primary branch (arrows indicate sites of future severing). By 17 h 10 min ddaD has severed at two sites (arrows). The primary branch of ddaE is thinned (arrowhead). Asterisk indicates where arbor temporarily moved out of the z-stack. By 19 h 00 min the primary branches of both neurons have been separated from the cell body. Filopodia extend from the cell bodies (arrow). Between 19-24 h the severed branches decrease in length, thin (arrowhead) and generate swellings (arrow). Scale bar: 40 µm. (B) Proximal thinning of dendrites. Selected frames from Movie 2 focusing on a proximal region of ddaD and ddaE. Arrowheads indicate beads. Filopodia extend and retract on the thinning branch throughout the sequence (arrows, 17 h 00 min). Between 17 h 00 min and 17 h 50 min ddaE's primary branch becomes noticeably thinner. Just prior to 18 h ddaE is severed, isolating the distal arbor. (C) Distal branch severing. Overview of distal severing in ddaD; box indicates area enlarged in the panels on the right. At 14 h 10 min beads are present at the site labeled with the arrow. The beads are dynamic and change position during the movie. At 15 h 10 min there is no obvious thinning at the site where the arbor will sever (arrowhead) Branch labeled with asterisk retracts a short distance following severing. Scale bar: 20 µm. (D) Branch fragmentation. Branch severed from the primary branch (lower arrow in A) shows thinning at the ends by 17 h 50 min. At 18 h 50 min the branch begins to fragment into blebs, and by 20 h 00 min the only trace of the branch are a few blebs of GFP remaining in the field. Arrow indicates beads; arrowhead denotes thinning. Scale bar: 40 µm. (E) Branch retraction. Frames from a movie showing a retraction event on the primary branches of ddaD. The primary branch shows dynamic filopodia (arrowhead) along its length. Retraction is evident at 18 h 30 min and is back to it cell body by 21 h 00 min. The ventral branch to the left undergoes fragmentation (arrow). Scale bar: 35 µm.