Axonal Growth-Associated Proteins

Science, 1986

Growth cones are specialized structures that form the distal tips of growing axons. During both normal development of the nervous system and regeneration of injured nerves, growth cones are essential for elongation and guidance of growing axons. Developmental and regenerative axon growth is frequently accompanied by elevated synthesis of a protein designated GAP43. GAP-43 has now been found to be a major component of growth-cone membranes in developing rat brains. Relative to total protein, GAP-43 is approximately 12 times as abundant in growth-cone membranes as in synaptic membranes from adult brains. Immunohistochemical localization of GAP-43 in frozen sections of developing brain indicates that the protein is specifically associated with neuropil areas containing growth cones and immature synaptic terminals. The results support the proposal that GAP-43 plays a role in axon growth. G AP-43 IS ONE OF A SMALL GROUP of axonally transported "growthassociated" proteins whose synthesis is increased 20to 100-fold during successful regeneration of axons in the central nervous system (CNS) of nonmammalian vertebrates (1-4) and in peripheral nerves of mammals (5). In adult mammalian CNS pathways that do not regenerate, GAP-43 synthesis and transport fails to increase beyond low background levels in response to injury (5, 6). During mammalian CNS de

Journal of Cell Biology, 1981

In an effort to determine whether the "growth state" and the "mature state" of a neuron are differentiated by different programs of gene expression, we have compared the rapidly transported (group I) proteins in growing and nongrowing axons in rabbits . We observed two polypeptides (GAP-23 and GAP-43) which were of particular interest because of their apparent association with axon growth . GAP-43 was rapidly transported in the central nervous system (CNS) (retinal ganglion cell) axons of neonatal animals, but its relative amount declined precipitously with subsequent development. It could not be reinduced by axotomy of the adult optic nerves, which do not regenerate ; however, it was induced after axotomy of an adult peripheral nervous system nerve (the hypoglossal nerve, which does regenerate) which transported only very low levels of GAP-43 before axotomy. The second polypeptide, GAP-23 followed the same pattern of growth-associated transport, except that it was transported at significant levels in uninjured adult hypoglossal nerves and not further induced by axotomy. These observations are consistent with the "GAP hypothesis" that the neuronal growth state can be defined as an altered program of gene expression exemplified in part by the expression of GAP genes whose products are involved in critical growth-specific functions. When interpreted in terms of the GAP hypothesis, they lead to the following conclusions: (a) the growth state can be subdivided into a "synaptogenic state" characterized by the transport of GAP-23 but not GAP-43, and an "axon elongation state" requiring both GAPS; (b) with respect to the expression of GAP genes, regeneration involves a recapitulation of a neonatal state of the neuron ; and (c) the failure of mammalian CNS neurons to express the GAP genes may underly the failure of CNS axons to regenerate after axon injury .

Cell, 1984

Neuroscience, 1994

In optic fibers, as in most axons of the central nervous system, the axonal growth-associated protein, GAP-43, is abundant during development but absent in adults. Since optic fibers can be induced to regenerate in culture, we examined whether this was associated with an increased expression of GAP-43 in adult mouse optic fibers that were regenerating from organotypic retinal explants on to laminin substrates. We found that simply placing adult mouse retina in culture under serum-free conditions was sufficient to induce GAP-43, which was detectable after about four to five days in vitro, coincident with the initiation of neurite outgrowth. In explants taken from animals in which the optic nerve was crushed in the orbit eight days prior to culturing, GAP-43 was observed within one day, as was neurite outgrowth. This priming effect was also seen in uivo as an increased level of GAP-43 reactivity in retinal ganglion cells and optic fibers in histological sections taken eight days after nerve crush. Reactivity in the adult fibers in culture was comparable to that observed in optic neurites growing from embryonic retinal explants and could be maintained for at least four weeks in culture. In the adult neurites, especially with longer times in culture, GAP-43 tended to be concentrated into varicosities that were often found in terminal-like arbors that formed in culture. Placing adult retina in culture under serum-free conditions is sufficient to induce re-expression of GAP-43 for an indefinite period of time. This suggests that GAP-43 expression and the propensity for growth in uivo may be repressed by a factor that is absent in vitro. In adult mammals, axons of the CNS do not, as a general rule, regenerate when severed.

Neuron, 1991

Although maturing neurons undergo a precipitous decline in the expression of genes associated with developmental axon growth, structural changes in axon arbors occur in the adult nervous system under both normal and pathological conditions. Furthermore, some neurons support extensive regrowth of long axons after nerve injury. Analysis of adult dorsal root ganglion (DRG) neurons in culture now shows that competence for distinct types of axon growth depends on different patterns of gene expression. In the absence of ongoing transcription, newly isolated neurons can extend compact, highly branched arbors during the first day in culture. Neurons subjected to peripheral axon injury 2-7 d before plating support a distinct mode of growth characterized by rapid extension of long, sparsely branched axons. A transition from "arborizing" to "elongating" growth occurs in naive adult neurons after ϳ24 hr in culture but requires a discrete period of new transcription after removal of the ganglia from the intact animal. Thus, peripheral axotomy-by nerve crush or during removal of DRGs-induces a transcription-dependent change that alters the type of axon growth that can be executed by these adult neurons. This transition appears to be triggered, in large part, by interruption of retrogradely transported signals, because blocking axonal transport in vivo can elicit competence for elongating growth in many DRG neurons. In contrast to peripheral axotomy, interruption of the centrally projecting axons of DRG neurons in vivo leads to subsequent growth in vitro that is intermediate between "arborizing" and "elongating" growth. This suggests that the transition between these two modes of growth is a multistep process and that individual steps may be regulated separately. These observations together suggest that structural remodeling in the adult nervous system need not involve the same molecular apparatus as long axon growth during development and regeneration.

Exp Brain Res, 1991

Although mature mammalian CNS neurons do not normally regenerate axons after injury, it is well established that they will regrow axons over long distances into peripheral nerve implants. We have autografted segments of sciatic nerve into the brains of adult albino rats and have used light and electron microscopic immunocytochemistry to examine the distribution of the growth associated protein GAP-43 in and around the graft in the first two weeks following implantation. GAP-43 was present, 3-14 days after grafting, in small non-myelinated axonal sprouts in the brain parenchyma around the proximal tip of the graft. At 11-14 days after implantation similar sprouts within the graft itself were GAP-43 immunoreactive. The sprouts were either naked or associated with other cell processes (chiefly of Schwann cells; to a lesser extent of astrocytes). We also show that small numbers of neuronal perikarya around the tip of the graft become GAP-43 immunoreactive 11-14 days after implantation. Thus mature mammalian CNS neurons regenerating axons into a PNS graft display a marked increase in their content of GAP 43. In addition, we report that small plaques of GAP-43 reaction product are sometimes present on the plasma membranes of Schwann cells or astrocytes adjacent to immunoreactive axons, and that narrow sheet-like or filopodial processes of astrocytes, Schwann cells and possibly other non-neuronal cell types, may contain small amounts of GAP-43.

Trends in Neurosciences, 1997

Several lines of investigation have helped clarify the role of GAP-43 (F1, B-50 or neuromodulin) in regulating the growth state of axon terminals. In transgenic mice, overexpression of GAP-43 leads to the spontaneous formation of new synapses and enhanced sprouting after injury. Null mutation of the GAP-43 gene disrupts axonal pathfinding and is generally lethal shortly after birth. Manipulations of GAP-43 expression likewise have profound effects on neurite outgrowth for cells in culture. GAP-43 appears to be involved in transducing intra-and extracellular signals to regulate cytoskeletal organization in the nerve ending. Phosphorylation by protein kinase C is particularly significant in this regard, and is linked with both nerve-terminal sprouting and long-term potentiation. In the brains of humans and other primates, high levels of GAP-43 persist in neocortical association areas and in the limbic system throughout life, where the protein might play an important role in mediating experience-dependent plasticity.

The Journal of Neuroscience, 2001

The highly regulated expression of neurofilament (NF) proteins during axon outgrowth suggests that NFs are important for axon development, but their contribution to axon growth is unclear. Previous experiments in Xenopus laevis embryos demonstrated that antibody-induced disruption of NFs stunts axonal growth but left unresolved how the loss of NFs affects the dynamics of axon growth. In the current study, dissociated cultures were made from the spinal cords of embryos injected at the two-cell stage with an antibody to the middle molecular mass NF protein (NF-M), and time-lapse videomicroscopy was used to study early neurite outgrowth in descendants of both the injected and uninjected blastomeres. The injected antibody altered the growth dynamics primarily in long neurites (Ͼ85 m). These neurites were initiated just as early and terminated growth no sooner than did normal ones. Rather, they spent relatively smaller fractions of time actively extending than normal. When growth occurred, it did so at the same velocity. In very young neurites, which have NFs made exclusively of peripherin, NFs were unaffected, but in the shaft of older neurites, which have NFs that contain NF-M, NFs were disrupted. Thus growth was affected only after NFs were disrupted. In contrast, the distributions of ␣-tubulin and mitochondria were unaffected; thus organelles were still transported into neurites. However, mitochondrial staining was brighter in descendants of injected blastomeres, suggesting a greater demand for energy. Together, these results suggest a model in which intra-axonal NFs facilitate elongation of long axons by making it more efficient.

Developmental Brain Research, 1984

Dissociated chick embryo peripheral and central nervous system cultures enriched for neurons by differential adherence, defined medium and cytosine arabinoside release to the culture environment molecular species which enhance the performance of neurons in limiting conditions. Culture medium conditioned by the neurons can be depleted of substrate-attached material by serial passage on poly-o-lysine substrate, leaving in the medium factors which promote neurite outgrowth on poly-o-lysine. The substrate-attached material which enhances neuron survival and neurite extension is heat-and trypsin-labile but is not affected by prior treatment with antisera to mouse NGF, human plasma fibronectin or laminin. The autostimulation phenotype displayed by neurons may play a role in neuronal survival or axonal growth during neuronal development.