Abstract
We wished to know whether the cell death and phagocytosis seen near the outgrowing nerve front in the hindlimb delineate axon pathways and, if so, whether the cells died only in the presence of growth cones. We unilaterally deleted the lumbosacral neural tube and reconstructed the patterns of neurite outgrowth and phagocytes during the stage when neurites first begin to colonize the thigh. In the control limbs, sensory and motor nerve pathways coincided with sites of phagocytosis, including those pathways that had yet to be colonized by growth cones. For instance, phagocytes were clustered at foci within the muscle masses where muscle nerves form a day later. However, they were not seen in adjacent, nonpathway regions such as posterior sclerotome or dorsal and ventral to the region of the plexus in which axons extend only posteriorly. Phagocytes were also seen in defined regions that are probably inaccessible to growth cones because they are too distant from pathways (i.e., subjacent to the apical ectodermal ridge) or express substances that are typical of precartilagenous tissues which may prohibit axon advance. In the experimental limbs, we conservatively estimated that neurite outgrowth was reduced to less than one-tenth (neurites were visible only with electron microscopy) or less than one-third of normal. Outgrowth extended less far distally and, in half the cases, motor innervation was completely abolished. Despite the extensive reduction in neurite outgrowth, the distribution of phagocytes was indistinguishable from that of the control side. Furthermore, the number of phagocytes did not differ significantly. We conclude that cell death delineates axon pathways remarkably well and does so without an interaction with growth cones; it is an independent characteristic of the axonal pathways and may be directly or indirectly important to axonal pathfinding. This is the first identification of a feature that characterizes prospective nerve pathways in the hindlimb.
