The innervation of FGF-induced additional limbs in the chick embryo.

Motoneurones that supply the vertebrate limb innervate their muscle targets in a highly reproducible manner. As development proceeds, these limb-specific motoneurones send out axons, which grow towards the developing limb and then congregate at its base to form the plexus. In the plexus, in response to unknown positional cues, these axons rearrange, often changing their original spatial relationships, before sorting out to emerge in the defined nerve trunks that innervate the limb. Several proposals have been put forward to explain how this reproducible innervation pattern is achieved. These include (1) that early differences in the motoneurone identity dictate their future axonal trajectories, (2) that axons actively respond to attractive or repulsive positional cues provided by the limb bud itself, or (3) that motor axons are passively deployed, following pathways of least mechanical resistance. We have addressed the question of the relative roles of motoneurone identity and the signals that the axons encounter on their journey towards the limb bud. Using the developing chick embryo as our experimental model we tested the effect of providing an additional limb target for motor axons leaving the flank level of the spinal cord. To do this we placed FGF-soaked beads in the presumptive flank of 2-day-old chick embryos. This treatment induces an additional limb containing muscles. We investigated whether such additional limbs are innervated and by which neurones. We show that rather than the additional limbs being solely supplied by axons diverted from the two existing limb plexuses, motoneurones that normally supply the flank alter their trajectories to enter the induced limb. Once in the limb, axons respond to positional cues within the bud to generate the stereotypical innervation pattern. Our results show that the tendency of 'flank' motoneurones to innervate flank can be overcome by the presence of an additional limb.