Induction and regulation of nitric oxide synthase in retinal Müller glial cells

Müller glial cells from the rat retina were examined for their capacity to produce nitric oxide (NO). Treatment of retinal Müller glial (RMG) cells with lipopolysaccharide (LPS), interferon-gamma, and tumor necrosis factor-alpha induced NO synthesis as determined by nitrite release in media. Simultaneous addition of LPS, interferon-gamma, and tumor necrosis factor-alpha caused the largest increase in NO synthesis. NO biosynthesis was detected after 12 h and was dependent on the dose of LPS, interferon-gamma, and tumor necrosis factor-alpha. Stereoselective inhibitors of NO synthase (NOS), cycloheximide and transforming growth factor-beta, blocked cytokine-induced NO production. Cytosol from LPS/cytokine-treated RMG cultures, but not from unstimulated cultures, produced a calcium/calmodulin-independent conversion of L-arginine to L-citrulline that was completely blocked by NOS inhibitor. The expression of NOS in RMG cells was confirmed by northern blot analysis, in which stimulation of these cells led to an increase in NOS mRNA levels. We conclude that RMG cells can express an inducible form of NOS similar to the macrophage isoform. High NO release from activated RMG cells might represent a protection from infection but may also contribute to the development of retinal pathologies.