L-Arginine Trigger Photoreceptor Death via Nitric Oxide Induced cGMP Accumulation

Abstract

Abstract: : Purpose: L-arginine is known non-toxic amino acid, so it is generally used as drug vehicle. However, it was reported that intravitreal injection of commercial drugs (tPA and antibiotics) with L-arginine vehicle results in dose-dependent retinal toxicity. Because these drugs alone was not toxic in retina, we thought that L-arginine may be related to its toxicity. We investigate the biochemical mechanism of L-arginine toxicity in primary cultured mouse retina cells. Methods: We investigated the dose-dependent primary cultured mouse retina cells toxicity of commercial tPA (with L-arginine vehicle) and L-arginine alone. In order to examine the involvement of nitric oxide in L-arginine toxicity, we measured the alterations of nitric oxide concentration using Griess reagent and cGMP concentration using ELISA kit. The ability of NMMA (NOS inhibitor) and amiloride (CNG channel blocker) to protect against L-arginine toxicity was assessed. Cell death level and death type were determined by LDH release and histological finding. Results: L-Arginine solution was treated in doses of 0, 0.5, 2.5, 5 mM. At dosage of 5 mM, cell death was increased extensively and occurred in 24 hour after treatment. But cell death was not increased over 40% 3days after treatment with L-arginine. The results of photomicrographs showed that dead cell type was dominant photoreceptor cell. Nuclear fragmentation analyzed by fluorescence micrographs indicated that retina cell death induced by L-arginine only is apoptosis rather than necrosis. Nitrite and cGMP formation increased by two fold in cultured retinal cells treated with L-arginine compared to control (P<0.05). The cell viability was increased by 16% and 18% in the presence of 100 uM NMMA and 10 uM amiloride, respectively (P<0.05). Conclusions: This studies demonstrate that L-arginine induced toxicity resulting from large amount of NO donor in retina, followed by accumulation cGMP and open CNG ion channel.

Keywords: apoptosis/cell death • drug toxicity/drug effects • photoreceptors