Deciphering cellular mechanisms regulating neuromuscular development

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Motoneurons innervate their appropriate muscle targets with remarkable precision during embryogenesis. To accomplish this task, the growth cone must make numerous pathfinding decisions as it grows to its correct target. Most decisions can be described as growing towards a desired guidance cue or away from a repulsive one. A directional decision shared by all developing motor axons is that to extend an axon out of the spinal cord. The last decision is more refined and involves determining whether to synapse with a fast or slow contracting muscle fibre. Ultimately, all guidance cues activate distinct intracellular signaling pathways that trigger asymmetrical changes in the growth cone causing it to turn towards or away from a cue. The neuromuscular junction (NMJ) is a tripartite synapse containing the motor axon, muscle fibre and terminal Schwann cell (tSC). While the NMJ has been extensively studied for over half a century, the role of the tSC has been largely ignored. This is changing as it is becoming increasingly evident that tSCs modulate neurotransmission and their abnormal function may underlie certain motor neuron diseases. One of the difficulties in studying tSCs is the lack of an in vitro model system that accurately reflects the physiology of the endogenous NMJ. My lab has developed a novel model system to study motor axon guidance and synapse formation using motoneurons derived from mouse embryonic stem (ES) cells. In the present proposal will build on our previous success using ES cell-derived motoneurons and ES/chick chimeras to determine: 1) how slow motoneurons selectively innervate slow muscle fibres during development, 2) develop an in vitro model system of a functional tripartite NMJ, and 3) determine what intracellular signaling pathway underlie the directed growth of motor axons out of the spinal cord and into the periphery.