Increased intraretinal PO2 in short-term diabetic rats.

In diabetic retinopathy, neovascularization is hypothesized to develop due to hypoxia in the retina. However, evidence for retinal hypoxia is limited, and the progressive changes in oxygenation are unknown. The objective of this study was to determine if retinal hypoxia occurs early in the development of diabetes. Intraretinal oxygen (PO2) profiles were recorded with oxygen-sensitive microelectrodes in control and diabetic Long-Evans rats at 4 and 12 weeks after induction of diabetes. Diabetes did not affect oxygen consumption in the photoreceptors in either dark or light adaptation. Oxygenation of the inner retina was not affected after 4 weeks of diabetes, although vascular endothelial growth factor levels increased. At 12 weeks, average inner retinal PO2, normalized to choriocapillaris PO2, was higher in diabetic rats than in age-matched controls, which was opposite to what was expected. Thus retinal hypoxia is not a condition of early diabetes in rat retina. Increased inner retinal PO2 may occur because oxygen consumption decreases in the inner retina.

Diabetes changes expression of genes related to glutamate neurotransmission and transport in the Long-Evans rat retina.

PURPOSE: This study investigated changes in the transcript levels of genes related to glutamate neurotransmission and transport as diabetes progresses in the Long-Evans rat retina. Transcript levels of vascular endothelial growth factor (VEGF), erythropoietin, and insulin-like growth factor binding protein 3 (IGFBP3) were also measured due to their protective effects on the retinal vasculature and neurons. METHODS: Diabetes was induced in Long-Evans rats with a single intraperitoneal (IP) injection of streptozotocin (STZ; 65 mg/kg) in sodium citrate buffer. Rats with blood glucose >300 mg/dl were deemed diabetic. Age-matched controls received a single IP injection of sodium citrate buffer only. The retinas were dissected at 4 and 12 weeks after induction of diabetes, and mRNA and protein were extracted from the left and right retinas of each rat, respectively. Gene expression was analyzed using quantitative real-time reverse-transcription PCR. Enzyme-linked immunosorbent assay was used to quantify the concentration of VEGF protein in each retina. Statistical significance was determined using 2×2 analysis of variance followed by post-hoc analysis using Fisher's protected least squares difference. RESULTS: Transcript levels of two ionotropic glutamate receptor subunits and one glutamate transporter increased after 4 weeks of diabetes. In contrast, 12 weeks of diabetes decreased the transcript levels of several genes, including two glutamate transporters, four out of five N-methyl-D-aspartate (NMDA) receptor subunits, and all five kainate receptor subunits. Diabetes had a greater effect on gene expression of NMDA and kainate receptor subunits than on the α-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA) receptor subunits, for which only GRIA4 significantly decreased after 12 weeks. VEGF protein levels were significantly increased in 4-week diabetic rats compared to age-matched control rats whereas the increase was not significant after 12 weeks. Transcript levels of VEGF and VEGF receptors were unchanged with diabetes. Erythropoietin and IGFBP3 mRNA levels significantly increased at both time points, and IGFBP2 mRNA levels increased after 12 weeks. CONCLUSIONS: Diabetes caused significant changes in the transcriptional expression of genes related to ionotropic glutamate neurotransmission, especially after 12 weeks. Most genes with decreased transcript levels after 12 weeks were expressed by retinal ganglion cells, which include glutamate transporters and ionotropic glutamate receptors. Two genes expressed by retinal ganglion cells but unrelated to glutamate neurotransmission, γ-synuclein (SNCG) and adenosine A1 receptor (ADORA1), also had decreased mRNA expression after 12 weeks. These findings may indicate ganglion cells were lost as diabetes progressed in the retina. Decreased expression of the glutamate transporter SLC1A3 would lead to decreased removal of glutamate from the extracellular space, suggesting that diabetes impairs this function of Müller cells. These findings suggest that ganglion cells were lost due to glutamate excitotoxicity. The changes at 12 weeks occurred without significant changes in retinal VEGF protein or mRNA, although higher VEGF protein levels at 4 weeks may be an early protective response. Increased transcript levels of erythropoietin and IGFBP3 may also be a protective response.

Oxygen consumption and distribution in the Long-Evans rat retina.

The purpose of this study was to investigate the oxygen distribution and consumption in the pigmented Long-Evans rat retina in vivo during dark and light adaptation, and to compare these results to previous work on cat and albino rat. Double-barreled microelectrodes recorded both intraretinal PO(2) depth profiles and the electroretinogram (ERG), which was used to identify the boundaries of the retina. Light adaptation decreased photoreceptor oxygen consumption per unit volume (Q(av)) from 3.0 ± 0.4 ml·100 g(-1) min(-1) (mean ± SEM) in darkness to 1.8 ± 0.2 ml·100 g(-1) min(-1) and increased minimum outer retinal PO(2) at the inner segments (P(min)) from 17.4 ± 3.0 to 29.9 ± 5.3 mmHg. The effects of light on outer retinal PO(2) and Q(av) were similar to those previously observed in cat, monkey, and albino rats; however, dark-adapted P(min) was higher in rat than cat. The parameters derived from fitting the oxygen diffusion model to the rat data were compared to those from cat. Oxygen consumption of the inner segments (Q(2)) and choroidal PO(2) (P(C)) in rat and cat were similar. P(min) was higher in rat than in cat for two reasons: first, rat photoreceptors have a shorter oxygen consuming region; and second, the retinal circulation supplied a greater fraction of consumed oxygen to rat photoreceptors. The average PO(2) across the inner retina (P(IR)) was not different in dark adaptation (25.4 ± 4.8 mmHg) and light adaptation (28.8 ± 5.4 mmHg) when measured from PO(2) profiles. However, with the microelectrode stationary at 9-18% retinal depth, a small consistent decrease in PO(2) occurred during illumination. Flickering light at 6 Hz decreased inner retinal PO(2) significantly more than an equivalent steady illumination, suggesting that changes in blood flow did not completely compensate for increased metabolism. This study comprehensively characterized rat retinal oxygenation in both light and dark, and determined the similarities and differences between rat and cat retinas.