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. Author manuscript; available in PMC: 2016 May 18.
Published in final edited form as: Curr Biol. 2015 Apr 23;25(10):1282–1295. doi: 10.1016/j.cub.2015.03.028

Figure 7. unc-3 coordinates synaptogenesis and neurotransmitter choice in ventral nerve cord (VNC) cholinergic motor neurons.

Figure 7

A: For simplicity, the axons of four DA and four DB cholinergic VNC MNs are shown in green. In total, there are 9 DA and 7 DB MNs along the C.elegans VNC. The cell bodies of the rest of cholinergic VNC MNs (VA, VB, AS, VC) are shown as grey dots. The synapses of DA and DB MNs onto dorsal body wall muscle (BWM) are highlighted with black dots. Dotted box shows the region of the dorsal nerve cord where synapses were analyzed in Figure 7 B–D. Dorsal BWM is not shown.

B: The number of presynaptic specializations of the DA and DB MNs onto dorsal BWM is significantly reduced in unc-3(e151) animals. Presynapse was visualized by driving RAB-3 protein expression (fused to gfp) in DA and DB MNs using the unc-3 promoter (transgene otIs437). Presynaptic boutons were counted in a 200 µm region located at the most anterior part of the dorsal nerve cord (DNC). *** : p value < 0.001. N = 15. Mean values are represented with a vertical red line.

C: Double immunofluorescence staining for gfp and the AChR subunit UNC-38 allows to visualize the presynaptic specializations of DA and DB MNs (RAB-3::GFP, green signal), as well as the post-synaptic specializations of dorsal BWM (UNC-38, red signal) in WT and unc-3(e151) animals.

D–E: The average number of muscle arms (asterisks) per BWM of the dorsal right quadrant was evaluated in WT and unc-3(e151) animals. The transgenic strain trIs30 allows for visualization of the muscle cell in yellow (him-4prom::MB::YFP) and the dorsal nerve cord in red (hmr-1bprom::DsRed2 and unc-129prom::DsRed2). No significant differences in the number of muscle arms per BWM between WT and unc-3(e151) animals were observed. Five different muscle cells (#9, #11, #13, #15, #17) were analyzed. N = 10. N.S, not significant. Mean values are represented with a horizontal red line.

F–I: The expression of madd-4L (F) and madd-4B (H) transcriptional reporters is significantly affected in VNC cholinergic MNs of unc-3(e151) mutants. Scale bar 50 µm. Anterior is to the left. Results for three independent transgenic madd-4L reporter lines (otEx4946, otEx5410, sEx14990) are quantified in G. Red horizontal lines represent the mean value. N > 20 for otEx4946, otEx5410, and sEx14990 in WT and unc-3(e151) animals. Results for two independent transgenic madd-4B reporters lines (otEx5601, otEx5603) are quantified in I. Red horizontal lines represent the mean value. N = 25 – 30 for otEx5601 and otEx5603 in WT and unc-3(e151) animals. *** : p value < 0.001.