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T. L. LeVatte, M. L. Archibald, X. Wang, B. C. Chauhan; Increase in Endothelin B Receptor Expression in Optic Nerve Astrocytes in Endothelin-1 Induced Chronic Experimental Optic Neuropathy. Invest. Ophthalmol. Vis. Sci. 2007;48(13):3289.
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To quantify endothelin B (ETB) receptor expression in the optic nerve following retinal ganglion cell (RGC) loss in an experimental model of endothelin-1 (ET-1) induced chronic optic in rat.
Brown Norway rats (275-300 g), whose RGCs had been retrogradely labeled with 2% fluorogold from both superior colliculi, were surgically implanted with osmotic minipumps delivering 10-7 M (n=5), 10-9 M (n=3) or 10-11 M (n=3) ET-1 to the optic nerve approx. 1 mm behind one globe while the other eye served as a control. Animals were sacrificed after 21 days of ET-1 delivery. The retinas were whole-mounted for estimates of RGC survival and the optic nerves removed for Western immunoblot analysis using a polyclonal antibody. The blots were also stained for beta-actin to normalize the densitometric measurements to adjust for uneven protein loading. In another group of animals (n=2) optic nerves used for immunohistochemistry using confocal microscopy. Cryostated coronal optic nerve sections (20 µm) were double-labeled with antibodies against ETB and glial fibrillary acidic protein (GFAP). RGC survival estimates were made with fluorescent micrographs taken at 1, 2, and 3 mm from the optic nerve head in each quadrant.
ETB receptor expression was higher in the experimental eye compared to the fellow control eye in 8 (73%) of the 11 animals with a mean (+ SD) increase of 29.8 (+ 34.8)% in the densitometric analyses of western immunoblots. As expected, the experimental nerves showed stronger labeling for ETB in the experimental eyes compared to control eyes with ETB-positive cells almost completely co-localizing with GFAP-positive cells indicating that ETB receptor upregulation occurs in optic nerve astrocytes. There was a weak inverse relationship between RGC survival and increase in ETB receptor expression (r2=0.51).
There is an upregulation of ETB receptor expression in optic nerve astrocytes in ET-1 induced chronic optic neuropathy causing RGC loss.
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