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Phi Ho, Vu Tran, Lais Cabrera, Sanjoy Bhattacharya; Localization of mechanotransduction channels in the trabecular meshwork of glaucomatous DBA/2J mice. Invest. Ophthalmol. Vis. Sci. 2013;54(15):4593.
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© ARVO (1962-2015); The Authors (2016-present)
To determine whether selected mechanotransduction channels (TREK-1, TRPM2, TRPA1 and TRPC1-6) localize in the trabecular meshwork (TM) of glaucomatous DBA/2J mice. To determine whether exposure of selected lipids results in modulation of level of any of these channels on TM cells. Our hypothesis is that select lipids modulate mechanotransduction channels leading to alteration of TM cell shape and behavior.
All studies were performed adhering to the ARVO statement for use of animals in vision research under IACUC approved protocols. DBA/2J mice of age 3-8 months (n=40 at each point) was used for detection of mechanotransduction channels. Immunohistochemistry and Western Blot analyses were carried out using commercial antibodies to mechanotransduction channels (TREK-1, TRPM2, TRPA1 and TRPC1-4 and 6) that were found in the DBA/2J TM by genomic analyses (from GEO database confirmed with RT-PCR analyses). Two human TM cell lines maintained in the laboratory were subjected to treatment with select phospholipids and determination of changes in the level using polyclonal antibodies to TREK-1. The phospholipids for these experiments were selected based on their prior differential identification in normal but not glaucomatous TM using shotgun lipidomics approach in our laboratory.
We detected TREK-1, TRPM2, TRPC3 and TRPC6 in the TM of DBA/2J mice at 3-8 months of age. We detected some changes in the level of these channels with aging in DBA/2J mice. We found level changes in TREK-1 channels (normalized with GAPDH or actin) determined by Western Blot analyses in human TM cells in response to exposure to select exogenous added phosphatidylserines and sphingomyelins in cell culture.
Our investigation confirms presence of TREK-1, TRPM2, TRPC1-4 and TRPC6 mechanotransduction channels at protein levels in the DBA/2J mice TM. This is in parallel to observation in the human TM. Levels of TREK-1 was modulated in human TM cells in response to exposure to select exogenously added lipids.
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