Abstract
Recent studies demonstrated that the excitotoxic amino acid homocysteine induces apoptotic death of retinal ganglion cells in vivo. In the present study, an in vitro rat retinal ganglion cell (RGC-5), culture system was used to analyze the toxicity of acute exposure to high levels of homocysteine, the mechanism of homocysteine-induced toxicity, and the usefulness of type 1 sigma receptor (sigmaR1) ligands as neuroprotectants. When cultured RGC-5 cells were subjected to treatment with 1 mM D,L-homocysteine, a significant increase in cell death was detected by terminal dUTP nick end labeling (TUNEL) analysis and analysis of activated caspase. When cells were treated with homocysteine- or glutamate in the presence of MK-801, an antagonist of the N-methyl-D-aspartate (NMDA) receptor, the cell death was inhibited significantly. In contrast, NBQX, an antagonist of the AMPA/Kainate receptor, and nifedipine, a calcium channel blocker, did not prevent the homocysteine- or glutamate-induced cell death. Semiquantitative RT-PCR and immunocytochemical analysis demonstrated that RGC-5 cells were exposed to homocysteine or glutamate express type 1 sigma receptor at levels similar to control cells. Treatment of RGC-5 cells with 3 or 10 microM concentrations of the sigmaR1-specific ligand (+)-pentazocine inhibited significantly the apoptotic cell death induced by homocysteine or glutamate. The results suggest that homocysteine is toxic to ganglion cells in vitro, that the toxicity is mediated via NMDA receptor activation, and that the sigmaR1-specific ligand (+)-pentazocine can block the RGC-5 cell death induced by homocysteine and glutamate.