Growth cone morphology varies with position in the developing mouse visual pathway from retina to first targets

Abstract

We have labeled the growth cones of retinal ganglion cell axons with HRP in intact mouse embryos. This has allowed us to visualize growth cone morphology during outgrowth along an entire CNS pathway from origin to target; to ask whether growth cone forms, and thus behaviors, differ at various points along the pathway; and to study the relationships of growth cones with the cellular environment. During the major period of axon outgrowth between embryonic day (E) 12 and 15, growth cones in the optic nerve are highly elongated (up to 40 microns) and have lamellopodial expansions, but the majority lack the microspikes or filopodia characteristic of many growth cones. Within the optic chiasm (E13–15), most growth cones shorten and spread, and project several short filopodia. In the optic tract, growth cones become more slender and again lack filopodia, resembling sleeker versions of optic nerve growth cones. Near the first target region (lateral geniculate nucleus), growth cones with filopodia arise from individual axon lengths and turn medially toward the target. Within target regions, the branches of immature axon arbors are tipped by minute swellings rather than by the enlarged growth cones prevalent during outgrowth toward targets. Electron-microscopic analysis of identified labeled growth cones in the optic nerve reveal intimate interactions between growth cones and glia or other growth cones in the form of invaginating contacts. In the optic nerve, growth cones contact immature glial (neuroepithelial) cells somewhere along their length, and also envelop bundles of neurites. In the chiasm, single growth cones simultaneously relate to many different profiles. These results demonstrate that in this single pathway from origin to targets, growth cone morphology varies systematically with position along the visual pathway. During outgrowth, simple growth cones are prominent when axons follow well-defined common pathways, and more elaborate filopodial forms appear when growth cones diverge, as they turn or come to decision regions. Together with observations in vitro and in nonmammalian nervous systems in situ, these data serve as reference points for testing to what extent growth cone form reflects intrinsic factors and interactions with the environment.

Retinal axon divergence in the optic chiasm: dynamics of growth cone behavior at the midline [published erratum appears in J Neurosci 1995 Mar;15(3):following table of contents]

“…At this site, ipsilateral-projecting fibers diverge from crossing fibers, by developing large, often bifurcated or multiply branched terminal endings at the border of a zone 150-200 pm proximal to the midline. These growth cones display even more complex morphologies than those described in our original study on growth cone form and position in the mouse visual system and chiasm in particular (Bovolenta and Mason, 1987). By inference from the images of curving growth cones and the sharp angle of the proximal neurite, we concluded that those branched fibers tipped with bifurcated or complex growth cones turn away from the midline back toward the ipsilateral optic tract.…”

“…A large number of studies have demonstrated in fixed dyelabeled preparations that simple streamlined growth cones are seen in straight paths and complex spread forms bearing filopodia are seen in decision regions (e.g., Tosney and Landmesser, 1985;Caudy and Bentley, 1986;Bovolenta and Mason, 1987;Nordlander, 1987;Holt, 1989;Norris and Kalil, 1990;Bovolenta and Dodd, 1991;Kim et al, 1991;Yaginuma et al, 1991;Wang et al, 1993). In the present study, we correlated form with behavior, within the single locale of the chiasm, and observed that the simple forms were common during advance and the complex forms during a pause in growth, in agreement with two other recent studies using video time-lapse imaging (Kaethner and Stuermer, 1992;Halloran and Kalil, 1994).…”

“…These studies have shown that in following precise trajectories, growth cones display dramatic alterations in their forms, thought to reflect the interactions that growth cones have with cellular and molecular cues in these regions. These analyses have also led to the conclusion that the variations in growth cone form are position specific, and that complex forms are more common in decision regions than in tracts, where streamlined forms predominate (e.g., Roberts and Taylor, 1983;Tosney and Landmesser, 1985;Caudy and Bentley, 1986;Bovolenta and Mason, 1987;Nordlander, 1987;Holt, 1989;Bovolenta and Dodd, 1990;Norris and Kalil, 1990;Kim et al, 199 1;Yaginuma et al, 1991;Wang et al, 1993).…”

Cellular Strategies of Axonal Pathfinding

“…These studies provided an index for analyzing growth cone morphology and behaviors in vivo. In the mid 1980s, numerous studies based on dye-labeling, static and dynamic imaging, and EM of growth cones in invertebrates and vertebrates convincingly showed that growth cone morphology mirrors three general behaviors that are related to substrata and cellular environments through which they grow (e.g., Tosney and Landmesser 1985;Caudy and Bentley 1986;Bovolenta and Mason 1987;Norris and Kalil 1990;Kim et al 1991). Torpedo-shaped growth cones, often with convex and concave lamellar "wings" extending from a central shaft, are observed during extension on axon bundles in tracts in vivo.…”

Recovery of Neurofilament Expression Selectively in Regenerating Reticulospinal Neurons

“…They have few microfilaments, are densely packed with phosphorylated neurofilaments (NFs) (Hall et al, 1991;Lurie et al, 1994;Hall and Lee, 1995;McHale et al, 1995;Pijak et al, 1996), elongate slowly (Ͻ120 m /d) Selzer, 1983, 1984;Lurie and Selzer, 1991a;Davis and McClelland, 1994a), and contain very little actin (G. F. Hall, J. Yao, K. S. Kosik, and M. E. Selzer, unpublished observations). Thus the mechanism of growth cone motility in these regenerating central axons must be different from that of embryonic neuronal growth cones in vitro (Letourneau, 1981;Gordon-Weeks, 1989) or in situ (Ho and Goodman, 1982;Keshishian and Bentley, 1983;Tosney and Landmesser, 1985;Bovalenta and Mason, 1987;Nordlander and Singer, 1987;Bovalenta and Dodd, 1990;Yaginuma et al, 1991), which contain no NFs and grow 1-3 mm /d in a process involving complex interactions between actin microfilaments, myosin, and microtubules (Lin and Forscher, 1995;.…”