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DURING normal development of the vertebrate nervous system, large numbers of neurons in the central and peripheral nervous system undergo naturally occurring cell death1. For example, about half of all spinal motor neurons die over a period of a few days in developing avian, rat and mouse embryos1. Previous studies have shown that extracts from muscle and brain, secreted factors from glia, as well as several growth factors and neurotrophic agents, including muscle-derived factors, can promote the survival of developing motor neurons in vitro and in vivo2–15. But because neurotrophins and other known trophic agents administered alone or in combination are insufficient to rescue all developing motor neurons from cell death8, other neurotrophic molecules are probably essential for the survival and differentiation of motor neurons. Here we report that glial-cell-line-derived neurotrophic factor (GDNF), a potent neurotrophic factor that enhances survival of mammalian midbrain dopaminergic neurons16,17, rescues developing avian motor neurons from natural programmed cell death in vivo and promotes the survival of enriched populations of cultured motor neurons. Furthermore, treatment with this agent in vivo also prevents the induced death and atrophy of both avian and mouse spinal motor neurons following peripheral axotomy.

During normal vertebrate development, about half of spinal motoneurons are lost by a process of naturally occurring or programmed cell death. Additional developing motoneurons degenerate after the removal of targets or afferents. Naturally occurring motoneuron death as well as motoneuron death after loss of targets or after axotomy can be prevented by in vivo treatment with putative target (muscle) derived or other neurotrophic agents. Motoneurons can also be prevented from dying in vitro and in vivo (Y.Q.-W., R.W., D.P., J. Johnson and L. Van Eldik, unpublished data and refs 7, 13, 14) by treatment with central nervous system extracts (brain or spinal cord) and purified central nervous system and glia-derived proteins. Here we report that in vivo treatment of chick embryos with brain-derived neurotrophic factor rescues motoneurons from naturally occurring cell death. Furthermore, in vivo treatment with brain-derived neurotrophic factor (and nerve growth factor) also prevents the induced death of motoneurons that occurs following the removal of descending afferent input (deafferentation). These data indicate that members of the neurotrophin family can promote the survival of developing avian motoneurons.

The daily treatment of chick embryos with neuromuscular blocking agents during the period of massive naturally occurring death of spinal motoneurones (days 5–10) reduces the normal loss of these cells1,2. Because both pre- and postsynaptic blocking agents have this effect3, the cause seems to be related to a reduction in synaptic and/or muscular activity. If this interpretation is correct, an experimentally induced increase in nerve or muscle activity (and/or an increased activation of acetylcholine receptors, AChR), during the normal cell death period should accelerate or enhance the loss of motoneurones. Consistent with this prediction, we have found that daily treatment with the nicotinic receptor agonist, carbachol, or with the anti-cholinesterase (anti-ChE) agent, eserine, leads to a significant enhancement of cell death of spinal motoneurones3. However, because in the same study other related agents (for example nicotine) failed to alter normal cell death, we believed that a different approach might provide a better test of our prediction. We now report that short-term continuous electrical stimulation of the hindlimb (the musculature and nerve trunks) initiated at any time between stages 31 (day 7) and stage 34 (day 8) results in a significant reduction in the number of healthy motoneurones and an increase in the number of pyknotic (degenerating) motoneurones in the lumbar lateral motor column (LMC). As the number of pyknotic cells in the dorsal root ganglia (DRG) is unaltered, this effect seems to be specific to motoneurones.