Differentiation of Fetal Brain-derived Neural Precursors Co-cultured with Neonatal Retinal Cells

Abstract

Abstract: : Purpose: The specification of retinal cell fate is largely regulated by local cell-cell interactions. The neonatal (P0) retinal cells have the ability to be altered in their cell fate choice after the cells had undergone their terminal mitosis and to promote differentiation of early retinal precursors. In an attempt to reveal the potency of precursors, we analyzed the differentiation of fetal brain-derived neural precursors in vitro and in vivo. Methods: Neural precursors were prepared from the telencephalic neuroepithelium of EGFP (enhanced green fluorescence protein) transgenic mice on E14 (embryonic day 14). Cell suspension (2x105 cells) was injected into intravitreal or subretinal spaces of neonatal rat eyes. After transplantation, sections were prepared for immunohistochemistry. In addition, neural retinas from neonatal mice were isolated and dissociated into single cells. After 4 days, retinal spheres were dissociated and cultured with/without neural precursors from EGFP transgenic mice on E14 on Chamber Slides (Nunc) in N2/DMEM/bFGF medium. After that, the cells were fixed and processed for the detection of neuronal or retinal cell type markers. Results: Retinal transplantation of neural precursors revealed that the transplanted cells survive for at least 14 days. Immunohistochemical studies demonstrated that retinal cell type markers were not expressed in the transplanted cells. Also, in another series of in vitro experiments, P0 retinal precursor cells can differentiate into specific retinal cell types; (bIII-tubulin-positive, syntaxin-positive and Pax-6-positive) amacrine cells, (rhodopsin-positive) rod photoreceptors, (GFAP-positive) glial cells. In contrast, brain-derived neural precursors expressed no retinal specific markers. Conclusions: Our data suggest that fetal brain-derived neural precursors have acquired regional specification on E14.

Keywords: retinal culture • transplantation • retinal development