Martha E Stokely1, Scott T Brady, Thomas Yorio. 1. Department of Biomedical Sciences, University of North Texas Health Science Center, Fort Worth, Texas 76107-2699, USA.
Abstract
PURPOSE: Increased levels of endothelins (ETs) are associated with glaucoma and have been said to contribute to the development of glaucomatous optic neuropathy. In glaucoma, movement of selected components of anterograde axonal transport essential in ganglion cell survival is impaired-specifically, the transport of mitochondria. This study evaluates the effect(s) of a single administration of intravitreous ET-1 on anterograde axonal transport in the rat optic nerve. METHODS: Proteins for anterograde axonal transport were pulse labeled by intravitreous injection of (35)S-methionine plus or minus ET-1 (2 nmol) in HEPES buffer (pH 7.4). At appropriate time intervals, optic nerves were dissected, sectioned while frozen, and homogenized in denaturing buffer, and transported protein was quantitated by liquid scintillation counting. Counts corrected for efficiency, quench, background, and decay were statistically evaluated (ANOVA, n = 7). RESULTS: Effects of treatment with intravitreous ET-1 on anterograde axonal transport were significant, biphasic, and prolonged (4 hours to 21 days). The initial phase was a significant enhancement of transport at times normally associated with small, fast-moving tubulovesicles (4 and 24 hours), followed by significant impairments at times normally associated with transport of mitochondria (28-36 hours), cytoplasmic matrix (4 days), and cytoskeletal proteins (21 days). The most pronounced effect of ET-1 was decreased axonal transport at times associated with normal anterograde transport of mitochondrial proteins (28, 32, and 36 hours, P = < 0.001, P < 0.015, and P < 0.001, respectively). This was mimicked by ET-3 at 28 hours. CONCLUSIONS: Effects of intravitreous ET-1 are consistent with a receptor-mediated role for elevated ETs in pathologic misregulation(s) of anterograde axonal transport.
PURPOSE: Increased levels of endothelins (ETs) are associated with glaucoma and have been said to contribute to the development of glaucomatous optic neuropathy. In glaucoma, movement of selected components of anterograde axonal transport essential in ganglion cell survival is impaired-specifically, the transport of mitochondria. This study evaluates the effect(s) of a single administration of intravitreous ET-1 on anterograde axonal transport in the rat optic nerve. METHODS: Proteins for anterograde axonal transport were pulse labeled by intravitreous injection of (35)S-methionine plus or minus ET-1 (2 nmol) in HEPES buffer (pH 7.4). At appropriate time intervals, optic nerves were dissected, sectioned while frozen, and homogenized in denaturing buffer, and transported protein was quantitated by liquid scintillation counting. Counts corrected for efficiency, quench, background, and decay were statistically evaluated (ANOVA, n = 7). RESULTS: Effects of treatment with intravitreous ET-1 on anterograde axonal transport were significant, biphasic, and prolonged (4 hours to 21 days). The initial phase was a significant enhancement of transport at times normally associated with small, fast-moving tubulovesicles (4 and 24 hours), followed by significant impairments at times normally associated with transport of mitochondria (28-36 hours), cytoplasmic matrix (4 days), and cytoskeletal proteins (21 days). The most pronounced effect of ET-1 was decreased axonal transport at times associated with normal anterograde transport of mitochondrial proteins (28, 32, and 36 hours, P = < 0.001, P < 0.015, and P < 0.001, respectively). This was mimicked by ET-3 at 28 hours. CONCLUSIONS: Effects of intravitreous ET-1 are consistent with a receptor-mediated role for elevated ETs in pathologic misregulation(s) of anterograde axonal transport.
Authors: Dorota L Stankowska; Alena Z Minton; Margaret A Rutledge; Brett H Mueller; Nitasha R Phatak; Shaoqing He; Hai-Ying Ma; Michael J Forster; Thomas Yorio; Raghu R Krishnamoorthy Journal: Invest Ophthalmol Vis Sci Date: 2015-01-13 Impact factor: 4.799