Figure 4

Quantitative analysis of PlexinA and PlexinB branches and varicosities reveals opposing roles in local axonal growth and synapse formation. A, PlexinA RNAi within the pSc neuron increases the number of variable axonal branches, whereas PlexinB RNAi decreases the number of highly stereotyped (skeletal) branches. Significant decreases in the number of highly skeletal pSc branches were also observed in the PlexinA RNAi neurons due to an increase in branch routing errors. Asterisks denote significance of p < 0.05. B, The pSc axonal arbor sizes with PlexinA RNAi are significantly shifted towards larger values. The distribution of pSc arbor sizes with PlexinB RNAi on the other had is shifted towards smaller values. Frequency distributions of the total axonal branch lengths among the three genotypes, 455-Gal4, and 455-Gal4; UAS-PlexA-RNAi, and 455-Gal4; UAS-PlexB-RNAi were significantly different from each other (p < 0.05). The average arbor size for 455-Gal4 control flies was 1045 μm and approximately 30% of flies had total branch lengths of 1000 μm (blue bars). The average arbor size in PlexinB RNAi pSc neurons was 918 μm with 25% of the neurons having total branch lengths of 800 μm (green bars). Reducing PlexinA expression in pSc neurons had the opposite effect, and increased their average total branch lengths to 1219 μm with 35% of arbors at 1200 μm (yellow bars). Histogram bin width, 100 μm. C, Quantification of axonal varicosities is an indirect measure of synaptic contacts. Varicosities (arrowheads) along branches were counted to obtain a lower estimate of synapse number per pSc axon. Scale bar, 5 μm. D, PlexinA knockdown significantly increases varicosities along the pSc axon. Frequency distribution of the number of varicosities in 455-Gal4; UAS-PlexA-RNAi pSc axons (yellow bars) was shifted to the right compared to the 455-Gal4 distribution (blue bars) (p < 0.01), with no significant difference in 455-Gal4; UAS-PlexB-RNAi (green bars) compared to control.