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B. S. Winkler, G. Mane, R. Deda, M. Thomas, M. L. Duenow; Detoxicant Actions of Sulfhydryl Compounds in Retina. Invest. Ophthalmol. Vis. Sci. 2010;51(13):1425.
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© ARVO (1962-2015); The Authors (2016-present)
A recent publication from our laboratory (Winkler, IOVS, 49:3259, 2008) proposed that the exceptional vulnerability of photoreceptor cells to metabolic poisons and oxidants is linked to a specific deficiency in glutathione (GSH) and that the daily renewal of outer segments serves as a surrogate antioxidant/protectant. To test this hypothesis, we determined whether an elevation in the level of a GSH-mimetic in retinal cells protects against iodoacetate (IAA), a well-known inhibitor of glyceraldehyde-3-phosphate dehydrogenase (G3PDH) that selectively damages photoreceptor cells.
Isolated rat retinas were incubated in HEPES-buffered media of normal ionic concentration (pH=7.4, ambient air, 37C) for 1-3 hrs in the presence and absence of 10 mM N-acetylcysteine (NAC), a membrane permeable SH-containing compound that has antioxidant and detoxicant actions similar to GSH. During the incubations, samples of the media were withdrawn for estimation of lactate production and the level of NAC. At the end of the incubations, some control and NAC-treated retinas were transferred to media containing 0.01 mM iodoacetate (IAA) for 15 min. At the end of the 15 min exposure to IAA, retinas were rinsed in two successive 30 sec washes in pre-warmed control media and then homogenized in appropriate solutions for measurements of soluble SH-content and activity of G3PDH.
The content of soluble SH was similar in retinas incubated in control media over 3 hrs (20 nmoles/retina), but retinas incubated in media containing 10 mM NAC showed a time-dependent increase in the level of soluble SH. After 1 hr, SH-content was increased 3-fold and after 3 hrs SH-content was increased 5-fold. Aerobic lactate production was increased by 33% in NAC-treated retinas relative to the rate in control retinas, i.e., 1.35 micromoles lactate/hr/retina. Incubating control retinas in 0.01 mM IAA for 15 min led to a 60% inhibition of G3PDH, whereas in retinas pre-treated in NAC-containing media IAA did not inhibit the activity of G3PDH.
These results provide evidence of the beneficial effects of supplementation of retinas with a membrane-permeant thiol (NAC) against a specific chemical toxin that is known to selectively target photoreceptor cells. Protection was signalled by the lack of inhibition of G3PDH in retinas supplemented with NAC and is likely due to detoxification of IAA by NAC via a carboxymethylation reaction. In contrast, if NAC or GSH is not present (e.g., in photoreceptor cells), then this reaction will not occur, resulting in inhibition of G3PDH and death of the cells.
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