title: Involvement of GABA transporters in atropine-treated myopic retina as revealed by iTRAQ quantitative proteomics.
abstract: Atropine, a muscarinic antagonist, is known to inhibit myopia progression in several animal models and humans. However, the mode of action is not established yet. In this study, we compared quantitative iTRAQ proteomic analysis in the retinas collected from control and lens-induced myopic (LIM) mouse eyes treated with atropine. The myopic group received a (-15D) spectacle lens over the right eye on postnatal day 10 with or without atropine eye drops starting on postnatal day 24. Axial length was measured by optical low coherence interferometry (OLCI), AC-Master, and refraction was measured by automated infrared photorefractor at postnatal 24, 38, and 52 days. Retinal tissue samples were pooled from six eyes for each group. The experiments were repeated twice, and technical replicates were also performed for liquid chromatography-tandem mass spectrometry (LC-MS/MS) analysis. MetaCore was used to perform protein profiling for pathway analysis. We identified a total of 3882 unique proteins with <1% FDR by analyzing the samples in replicates for two independent experiments. This is the largest number of mouse retina proteome reported to date. Thirty proteins were found to be up-regulated (ratio for myopia/control > global mean ratio + 1 standard deviation), and 28 proteins were down-regulated (ratio for myopia/control < global mean ratio - 1 standard deviation) in myopic eyes as compared with control retinas. Pathway analysis using MetaCore revealed regulation of γ-aminobutyric acid (GABA) levels in the myopic eyes. Detailed analysis of the quantitative proteomics data showed that the levels of GABA transporter 1 (GAT-1) were elevated in myopic retina and significantly reduced after atropine treatment. These results were further validated with immunohistochemistry and Western blot analysis. In conclusion, this study provides a comprehensive quantitative proteomic analysis of atropine-treated mouse retina and suggests the involvement of GABAergic signaling in the antimyopic effects of atropine in mouse eyes. The GABAergic transmission in the neural retina plays a pivotal role in the maintenance of axial eye growth in mammals.
authors.affiliation: Singapore Eye Research Institute , 11 Third Hospital Avenue, Singapore 168751, Singapore.
authors.forename: Veluchamy A
authors.initials: VA
authors.lastname: Barathi
authors.forename: Shyam S
authors.initials: SS
authors.lastname: Chaurasia
authors.forename: Michael
authors.initials: M
authors.lastname: Poidinger
authors.forename: Siew Kwan
authors.initials: SK
authors.lastname: Koh
authors.forename: Dechao
authors.initials: D
authors.lastname: Tian
authors.forename: Candice
authors.initials: C
authors.lastname: Ho
authors.forename: P Michael
authors.initials: PM
authors.lastname: Iuvone
authors.forename: Roger W
authors.initials: RW
authors.lastname: Beuerman
authors.forename: Lei
authors.initials: L
authors.lastname: Zhou
mesh_terms: D000818:Animals; D001285:Atropine / Q000494:pharmacology*; D015153:Blotting, Western; D002853:Chromatography, Liquid; D050485:GABA Plasma Membrane Transport Proteins / Q000378:metabolism*; D005786:Gene Expression Regulation / Q000187:drug effects*; D007368:Interferometry; D051379:Mice; D009216:Myopia / Q000188:drug therapy / Q000378:metabolism*; D040901:Proteomics / Q000379:methods*; D012029:Refraction, Ocular / Q000187:drug effects / Q000502:physiology; D012160:Retina / Q000187:drug effects / Q000378:metabolism*; D053719:Tandem Mass Spectrometry
chemical_list: D050485:GABA Plasma Membrane Transport Proteins; D001285:Atropine
keywords: iTRAQ; lens-induced myopia mouse model; myopia; quantitative proteomics; retina proteomics