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Diabetic Retinopathy (DR) is one of the most common complications of diabetes and a major cause of blindness worldwide. We studied the transcriptome of the diabetic retina using Series Analysis of Gene Expression (SAGE) technology and observed a 45.6% reduction in transcript levels of glutamine synthetase (GS) in streptozotocin-induced diabetic rats compared with normal rats. RT-PCR and colorimetric enzyme activity assays revealed significant differences in GS mRNA expression and enzyme activity as early as the first month of diabetes development, with a progressive decrease in GS mRNA level and enzyme activity over a 12-month period. Northern blot analysis indicated a linear correlation between the reduction in GS expression and the time course of diabetic retinopathy (r = 0.802, p < 0.0001), which was validated by real-time RT-PCR (r = 0.731, p < 0.001). Our results implicate GS as a possible biomarker for evaluating the severity of developed diabetic retinopathy over the time course of diabetes progression.