Cell death of motoneurons in the chick embryo spinal cord. IV. Evidence that a functional neuromuscular interaction is involved in the regulation of naturally occurring cell death and the stabilization of synapses.

Embryos immobilized with neuromuscular blocking agents for differing periods between 4.5 and 9 days of incubation had an increased number of motoneurons in the brachial and lumbar lateral motor columns. Treatment with alpha-cobratoxin (alpha-CTX) on days 4--9, for instance, was able to prevent virtually all natural cell death during this period; control embryos had an average of 22,500 lumbar motoneurons on day 5.5, and 13,500 on day 10, whereas treated embryos had approximately 21,000 cells on day 10. Curare, alpha-CTX, alpha-bungarotoxin (alpha-BTX) and botulinum toxin were all about equally effective in preventing cell death. Similar treatment begun after day 12, however, had no effect on cell number. If even a partial immobilization was continued after day 10 (in embryos totally immobilized earlier) most of the excess neurons were maintained, in some cases right up to hatching, at which time the embryos died due to respiratory failure. In contrast, when administration of the immobilizing agents was stopped, allowing the embryos' motility to return to control levels, the excess neurons underwent a delayed cell death and total cell number fell to below control levels by days 16--18. Limb muscles from embryos with excess motoneurons exhibited relatively normal differentiation and had acetylcholinesterase (AChE) stained endplates which were innervated. Following curare treatment the two wing muscles, anterior and posterior latissimus dorsi, were found to have an increased number of AChE-stained endplates, whereas the only leg muscle examined quantitatively--the ischioflexorius (IFL)--did not; the IFL, did, however, have a markedly reduced variance in endplate distance, as well as other apparent differences suggesting an altered pattern of innervation. Our findings imply that the number of motoneurons undergoing natural cell death is closely related to muscle activity. Thus, functional interactions at the developing neuromuscular junction seem to be critical in controlling cell death. If a retrograde trophic factor is involved its action is somehow related to muscle activity.