The early development of retinal ganglion cell projections in the zebrafish, Brachydanio rerio.

Abstract

Retinal ganglion cell (RGC) axons and dendrites were labelled with the fluorescent carbocyanine dye, DiI, then photoconverted, and examined light- and electron microscopically. Injections into the optic chiasm of 32-72 hr embryos retrogradely labelled the RGCs and their dendrites. The first RGCs to extend an axon into the brain were located in the nasoventral retina, and spread progressively through dorsal to temporal retina. At 32-36 hrs the majority of RGCs labelled did not have dendrites, but by 38 hrs most did. Since optic axons do not reach their targets until 44 hrs, our results demonstrate that axonal contact with the target does not initiate dendritic development. Also, since RGCs did not have an apical process at 32-36 hrs, we conclude that the apical process of the RGC neuroepithelial precursor did not turn into a dendrite. The development of the retinofugal projection was studied by anterograde labelling of the RGC axons following the intraocular injection of DiI. In 6-7 day larvae the optic axons arborized in eight fields (AFs) evident in wholemounts, and two more evident only in sections. The first optic axons left the eye at 32 hrs and reached the tectum around 44 hrs. By 52 hrs the first non-tectal AFs were evident, and by 72 hrs all 10 AFs were innervated. Initially, optic axons arborized in only one AF, but as development proceeded some axons arborized in more than one. Light microscopic examination of wholemounts and sections revealed that the optic axons entered the brain rostral to the postoptic commissure, and paralleled the tract of the postoptic commissure (TPOC) part way to the tectum. Electron microscopic examination of seven leading optic growth cones revealed that they were separated from the TPOC by neuroepithelial endfeet, and did not contact it. Instead, the growth cones grew rostral to the TPOC in the morphologically identifiable intermediate zone (IZ), composed of neuroepithelial processes and endfeet. The majority of processes contacted by the growth cone and its filopodia within the IZ were neuroepithelial, suggesting that the guidance cues for the optic axons are located on the neuroepithelial processes and not on the TPOC.