Pax2+ Astrocytes In The Goldfish Optic Nerve Head During Continuous Growth And Regeneration

Abstract

Purpose: : During visual system development the optic nerve head (ONH) is a key region that allows the entrance of the retinal ganglion cell axons from the retina to the optic nerve. The ONH has a specific glial cell population which produces numerous molecules involved in the correct formation of the optic nerve, such as the transcription factor Pax2. The fish visual system is in continuous growth and can regenerate because the peripheral retina provides new retinal cells. Thus the adult fish ONH may retain some characteristics similar to those described during development.

Methods: : We used goldfish control and injured with a cryo-lesion of the peripheral retina (CL) and an optic nerve crush (OC). They were maintained from 2 to 210 days post-injury, in accordance with the European normative for animal care. We used double immunofluorescence analyses to label astrocytes (Pax2, GFAP, GS, S100, ZO1 and cytokeratin), proliferating cells (PCNA), growing axons (Zn8) and immunogold electron microscopy. With qPCR we analyzed pax2a gene expression levels in the ONH during regeneration.

Results: : In the ONH Pax2⁺ cells are astrocytes, different from those in the rest of the optic nerve and S100⁺ cells. They are GFAP⁺, cytokeratin⁺, ZO1⁺ and GS^- in control animals. During the regenerative process the ONH Pax2⁺ astrocytes and S100⁺ cells undergo modifications. During the first days after CL, the Pax2⁺ cell number is reduced and coincides with the absence of young axons. In contrast, there is an increase of S100⁺ processes. When the regenerating Zn8⁺ axons reach the ONH, there is an increase in the numbers of Pax2⁺ cell and the pax2a gene expression level in both the CL and OC. At the same time reactive S100⁺/GFAP⁺/GS⁺ astrocytes, located in the posterior ONH, organize the exit of the growing axons to the optic nerve. During the whole regenerative process we detected a ring of proliferating cells in the posterior ONH which generates mostly new Pax2⁺/PCNA⁺ astrocytes.

Conclusions: : Pax2⁺ astrocytes in the ONH form a unique glial cell population that participates actively in the ganglion cell axon regeneration together with S100⁺ astrocytes. Moreover, the transcription factor pax2a seems to play a fundamental role in this process.

Keywords: astrocytes: optic nerve head • regeneration • ganglion cells