Abstract

In the absence of external cues, neurons in vitro polarize by using intrinsic mechanisms. For example, cultured hippocampal neurons extend arbitrarily oriented neurites and then one of these, usually the one nearest the centrosome, begins to grow more quickly than the others. This neurite becomes the axon as it accumulates molecular components of the apical junctional complex. All the other neurites become dendrites. It is unclear, however, whether neurons in vivo, which differentiate within a polarized epithelium, break symmetry by using similar intrinsic mechanisms. To investigate this, we use four-dimensional microscopy of developing retinal ganglion cells (RGCs) in live zebrafish embryos. We find that the situation is indeed very different in vivo, where axons emerge directly from uniformly polarized cells in the absence of other neurites. In vivo, moreover, components of the apical complex do not localize to the emerging axon, nor does the centrosome predict the site of axon emergence. Mosaic analysis in four dimensions, using mutants in which neuroepithelial polarity is disrupted, indicates that extrinsic factors such as access to the basal lamina are critical for normal axon emergence from RGCs in vivo.

Highlights

  • A key step in neuronal morphogenesis is the emergence of correctly oriented axons and dendrites

  • Some of these centrosomes are clearly in the apical tips of ath5:gap-rfp-positive cells. When these cells enter into the differentiation process that will lead to an retinal ganglion cells (RGCs), the apical process is retracted as we have described, and the centrosome remains associated with the tip of the retracting process (20 out of 20 cells in five different embryos), even when the cell is extending its axon on the opposite side (Figure 6g and Additional file 6)

  • Factors extrinsic to RGCs are required for efficient RGC polarization in vivo: the role of the retinal pigment epithelium (RPE) and the basal lamina The results described in the previous section suggest a role of the retinal neuroepithelium environment in the orientation of RGC polarity, but raise the question of which elements of the environment are most important for influencing the site of axon growth in RGCs

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Summary

Introduction

A key step in neuronal morphogenesis is the emergence of correctly oriented axons and dendrites. If hippocampal cells are cultured soon after their final mitotic divisions, multiple neurites emerge simultaneously at seemingly random orientations. From these young multipolar neurons, one neurite begins to elongate preferentially, marking the beginning of polarization. It becomes the axon [2], and as it grows it inhibits the other neurites from becoming axons. Aided by adenomatous polyposis coli and KIF3A (a kinesin superfamily protein), proteins that travel along microtubules, these apical components accumulate at the tips of growing axons. The centrosome, acting as a microtubule organizing (page number not for citation purposes)

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