A NOVEL TYPE OF PROGRAMMED NEURONAL DEATH IN THE CERVICAL SPINAL-CORDOF THE CHICK-EMBRYO

We examined the massive early cell death that occurs in the ventral ho rn of the cervical spinal cord of the chick embryo between embryonic d ays 4 and 5 (E4 and E5). Studies with immunohistochemistry, in situ hy bridization, and retrograde-tracing methods revealed that many dying c ells express Islet proteins and Lim-3 mRNA (motoneuron markers) and se nd their axons to the somatic region of the embryo before cell death. Together, these data strongly suggest that the dying cells are somatic motoneurons. Cervical motoneurons die by apoptosis and can be rescued by treatment with cycloheximide and actinomycin D. Counts of motoneur on numbers between E3.5 and E10 revealed that, in addition to cell dea th between E4 and E5, motoneuron death also occurs between E6 and E10 in the cervical cord. Studies with [H-3]thymidine autoradiography and morphological techniques revealed that in the early cell-death phase ( E4-E5), genesis of motoneurons, axonal elongation, and innervation of muscles is still ongoing. However, studies with [H-3]thymidine autorad iography also revealed that the cells dying between E4 and E5 become p ostmitotic before E3.5. Increased size of peripheral targets, treatmen t with neuromuscular blockade, and treatment with partially purified m uscle or brain extracts and defined neurotrophic agents, such as NGF, BDNF, neurotrophin-3, CNTF, bFGF, PDGF, S100-beta, activin, cholinergi c differentiation factor/leukemia inhibitory factor, bone morphogeneti c protein-2, IGF-I, interleukin-6, and TGF-beta 1, were all ineffectiv e in rescuing motoneurons dying between E4 and E5. By contrast, motone urons that undergo programmed cell death at later stages (E6-E10) in t he cervical cord are target-dependent and respond to activity blockade and trophic factors. Experimental approaches revealed that early cell death also occurs in a notochord-induced ectopic super-numerary moton euron column in the cervical cord. Transplantation of the cervical neu ral tube to other segmental regions failed to alter the early death of motoneurons, whereas transplantation of other segments to the cervica l region failed to induce early motoneuron death. These results sugges t that the mechanisms that regulate motoneuron death in the cervical s pinal cord between E4 and E5 are independent of interactions with targ ets. Rather, this novel type of cell death seems to be determined by s ignals that either are cell-autonomous or are derived from other cells within the cervical neural tube.