Are Promoted by CPG15 In Vivo

We studied the morphological development of motor neuron axon terminal arbors in stage 47 albino Xenopus tadpoles, which are free swimming and have a functional motor circuitry, even though the tail musculature and innervation are still developing. (A) A schematic diagram of tadpole tail musculature shows that motor neuron axons leave the spinal cord through the ventral nerve root and travel caudally before they branch and form synapses. The tadpole tail is segmented as evident by the chevron-shaped myotome pattern. Each myotome contains a parallel series of muscle cells (light gray) that extend from one myotome junction to the next. Motor neuron axons that project to the tail musculature leave the ventrolateral spinal cord and traverse laterally in bundles until they reach the skeletal muscles of the tail. Once there, they continue extending caudally, where they branch and form complex synaptic terminals onto the ends of many muscle cells over one to several segments (Roberts and Clarke, 1982; Westerfield and Eisen, 1985). (B) Systemic injections of FITC-dextran and TR-αBTX into the heart allow the visualization of the outline of muscle cells (in green) and postsynaptic AChR (in red), respectively. The majority of synapses are at the ends of the muscle cells at the myotome junctions (Anderson and Cohen, 1974; Blackshaw and Warner, 1976; Chow and Cohen, 1983). (C) Higher magnification of an optical section through a myotome junction shows that each muscle cell has more than one postsynaptic site at each end. (D) To study the organization of motor neuron axon arbors as well as differentiated presynaptic structures on the branches together with AChR clusters on postsynaptic cells at the NMJ, we coelectroporated motor neurons in the spinal cord with plasmids expressing YFP and the synaptic vesicle protein synaptophysin fused to CFP and finally injected tadpoles with TR-αBTX immediately prior to the imaging session. (F and G) To identify synaptic sites on axon branches, we colocalized synaptophysin-CFP puncta with TR-αBTX-labeled AChR clusters in optical sections (see Experimental Procedures)