The development of most regions of the vertebrate nervous system includes a distinct phase of neuronal degeneration during which a substantial proportion of the neurons initially generated die. This degeneration primarily adjusts the magnitude of each neuronal population to the size or functional needs of its projection field, but in the process it seems also to eliminate many neurons whose axons have grown to either the wrong target or an inappropriate region within the target area. In addition, many connections that are initially formed are later eliminated without the death of the parent cell. In most cases such process elimination results in the removal of terminal axonal branches and hence serves as a mechanism to "fine-tune" neuronal wiring. However, there are now also several examples of the large-scale elimination of early-formed pathways as a result of the selective degeneration of long axon collaterals. Thus, far from being relatively minor aspects of neural development, these regressive phenomena are now recognized as playing a major role in determining the form of the mature nervous system.

Corticospinal axons innervate their midbrain, hindbrain, and spinal targets by extending collateral branches interstitially along their length. To establish that the axon shaft rather than the axonal growth cone is responsible for target recognition in this system, and to characterize the dynamics of interstitial branch formation, we have studied this process in an in vivo-like setting using slice cultures from neonatal mice containing the entire pathway of corticospinal axons. Corticospinal axons labeled with the dye 1,1'-dioctodecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (or Dil) were imaged using time-lapse video microscopy of their pathway overlying the basilar pons, their major hindbrain target. The axon shaft millimeters behind the growth cone exhibits several dynamic behaviors, including the de novo formation of varicosities and filopodia-like extensions, and a behavior that we term "pulsation," which is characterized by a variable thickening and thining of short segments of the axon. An individual axon can have multiple sites of branching activity, with many of the branches being transient. These dynamic behaviors occur along the portion of the axon shaft overlying the basilar pons, but not just caudal to it. Once the collaterals extend into the pontine neuropil, they branch further in the neuropil, while the parent axon becomes quiescent. Thus, the branching activity is spatially restricted to specific portions of the axon, as well as temporally restricted to a relatively brief time window. These findings provide definitive evidence that collateral branches form de novo along corticospinal axons and establish that the process of target recognition in this system is a property of the axon shaft rather than the leading growth cone.

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In newborn rats each retina projects principally to the contralateral superior colliculus, but there is also a sparse projection to the whole of the ipsilateral superior colliculus. During the first 2 weeks postnatally the ipsilateral projection normally becomes restricted to the rostromedial part of the superior colliculus. The restriction of this projection is due to the preferential death of ipsilaterally projecting retinal ganglion cells and is apparently the result of competition between optic fibers from the two eyes, since it can be prevented by enucleation of the opposite eye at birth. To determine if electrical activity plays a role in the normal restriction of the ipsilateral retinocollicular projection, the sodium channel-blocking agent tetrodotoxin was administered to one or both eyes during the first 2 weeks postnatally. Tetrodotoxin blockade of activity in one eye resulted in the persistence of a sparse projection from the opposite eye throughout the ipsilateral superior colliculus and the survival of a substantial number of the ipsilaterally projecting retinal ganglion cells in that eye that would normally have died. When both eyes were treated with tetrodotoxin no restriction of the ipsilateral projection was seen on either side. These findings suggest that the competition between retinal ganglion cell axons (either for terminal space or an essential trophic factor), which normally leads to retinal ganglion cell death and the restriction of the ipsilateral retinocollicular projection, is mediated in some way by electrical activity.During the development of most regions of the vertebrate central nervous system, there is a period during which many of the constituent neurons die (1-3). This phenomenon, although widespread and well-documented, is poorly understood in terms of its cause and control. We report here the results of an experiment that casts some light on how cell death in one of the better-studied sets of neural connections, the rat retinocollicular projection, may be controlled.In the rat, the vast majority of retinal ganglion cells (RGCs) project to the contralateral side of the brain and especially to the contralateral superior colliculus (SC) (4,5), whereas a much smaller number of RGCs project to the ipsilateral SC. From just after birth until about postnatal day 10 (P10) about 50% of the RGCs in both retinas die (6-8), and over the same period the ipsilateral retinocollicular projection becomes progressively restricted in its distribution. At birth this ipsilateral projection covers the entire colliculus, but by P10 it is normally restricted to the rostromedial edge of the SC (9). This restriction appears to be due to the preferential death of ipsilaterally projecting RGCs (10,19). Thus, whereas about 50% of the cells that project to the contralateral SC die, at least 85% of the ipsilaterally projecting RGCs that lie outside the so-called "temporal crescent" die (unpublished data). It seems that this preferential elimination of ipsilaterally projecting RGCs is due to their e...

We used the indirect immunofluorescence method to determine the crossreactivity of a library of 57 monoclonal antibodies (mAbs) against each of the subunits of the nicotinic acetylcholine receptor (nAcChoR) isolated from Torpedo and Electrophorus electric organs or from fetal calf and human muscle, with specific neural elements in the midbrain of the chick. Out of 17 mAbs that recognized motor end plates on chick muscle, 14 produced a similar pattern of labeling in the midbrain: the neuronal perikarya and dendrites in the lateral spiriform nucleus (SpL) were intensely labeled, and there was moderate labeling of fibers in certain of the deeper layers of the optic tectum, which disappeared after the SpL was destroyed electrolytically. Two lines of evidence suggest that the mAbs may be crossreacting with nAcChoRs in the midbrain. First, all of the mAbs that stained the SpL also stained neuromuscular junctions in skeletal muscle, whereas none of the 40 mAbs that failed to stain end plates crossreacted with the SpL; second, in vitro immunological studies and blocking experiments on tissue sections (in which unlabeled mAbs were used to block the staining of a directly fluorescein-treated mAb) indicated the presence of mAbs specific for unique antigenic determinants on all four of the subunits (a, .3, y, and 8) from Torpedo nAcChoR in chick midbrain and muscle. On the other hand, the distribution of mAb staining in the optic tectum does not closely parallel that of either acetylcholinesterase staining or of "251-labeled a-bungarotoxin binding; no toxin binding has been observed autoradiographically in the SpL, but the nucleus does contain moderately dense acetylcholinesterase staining. Taken together, our observations suggest that there may be a cholinergic input to the SpL and that the projection fibers from the SpL to the optic tectum (which are also stained with an antiserum to[Leu]enkephalin) may contain presynaptic nAcChoRs. It is clear, however, that the distribution of the putative nAcChoRs, a-bungarotoxin binding sites, and acetylcholinesterase staining in the avian midbrain are quite different, although they do overlap to some degree in the deeper layers of the optic tectum.Until recently, the only available method for localizing nicotinic acetylcholine receptors (nAcChoR) in the nervous system has involved the use of radiolabeled a-bungarotoxin (a-BTX) and autoradiography. Many studies in which this approach was used have been published in the last few years; in some cases, it has been possible to relate the distribution of the labeled toxin to that of known cholinergic pathways (1). The finding of substantial toxin binding in the optic tectum of a number of different vertebrates has been especially intriguing, and this has been taken as evidence that in these animals the central projections of the retina may be cholinergic (2). There is, however, a growing body of evidence that a-BTX binding does not necessarily indicate the presence of functional nAcChoRs in several neuronal populations (3-6).The p...