Differences in Nitric Oxide Production and Function: A Comparison of Retinal Ganglion Cells and Retinal Glial Cells Cultured under Hypoxic Condition

Abstract

Abstract: : Purpose: To compare the effects of hypoxia on nitric oxide synthase (NOS) expression, and the production and role of NO, between isolated retinal ganglion cells (RGCs) and retinal glial cells. Methods: Reverse transcription-polymerase chain reaction (RT-PCR) was employed to examine the presence of neuronal NOS mRNA, inducible NOS mRNA, and endothelial NOS mRNAs in the two cell types.RGCs and retinal glial cells were separately cultured under hypoxic (10% O₂) or control (20% O₂) conditions. Changes in NOS-mRNA expression were quantified by real-time PCR, and nitrite in the medium was measured up to 96 h of culture. The effects of non-NOS- and iNOS-selective inhibitors on hypoxia-induced release of nitrite in the culture medium were evaluated. The effects of hypoxia and methylene blue on RGC survival and growth of neurites were evaluated. Results: RT-PCR revealed the presence of three types of NOSs in the two types of cultured cells. Hypoxic culture conditions significantly changed the expression of all NOS mRNAs in retinal glial cells but not in RGCs. NO production showed significant changes corresponding to those of NOS mRNAs in retinal glial cells but not in RGCs, and both NOS inhibitors significantly reduced hypoxia-induced nitrite release in retinal glial cells. Hypoxia significantly reduced RGC survival, but did not affect the growth of neurites, whereas methylene blue inhibited the growth of neurites in the surviving RGCs. Conclusions: Retinal glial cells may be the major source of NO under hypoxic conditions and be involved in RGC survival, whereas NO from RGCs may be involved not in RGC survival but in synaptogenesis.

Keywords: retinal culture • retinal glia • nitric oxide