July 2018
Volume 59, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2018
Tandem-pore domain potassium channels are components of intracellular signaling pathways, determinants of pH sensitivity and mechanosensitivity of human trabecular meshwork cells.
Author Affiliations & Notes
  • Oleg Yarishkin
    Department of Ophthalmology and Visual Sciences, University of Utah School of Medicine, Salt Lake City, Utah, United States
  • Tam Phuong
    Department of Ophthalmology and Visual Sciences, University of Utah School of Medicine, Salt Lake City, Utah, United States
  • Jackson M Baumann
    Department of Bioenginering, University of Utah, Salt Lake City, Utah, United States
    Bioengineering Graduate Program, University of Utah, Salt Lake City, Utah, United States
  • Alan S Crandall
    Department of Ophthalmology and Visual Sciences, University of Utah School of Medicine, Salt Lake City, Utah, United States
  • Feryan Ahmed
    Glauconix, Rochester, New York, United States
  • Karen Yud Torrejon
    Glauconix, Rochester, New York, United States
  • David Krizaj
    Department of Ophthalmology and Visual Sciences, University of Utah School of Medicine, Salt Lake City, Utah, United States
    Department of Neurobiology & Anatomy, University of Utah School of Medicine, Salt Lake City, Utah, United States
  • Footnotes
    Commercial Relationships   Oleg Yarishkin, None; Tam Phuong, None; Jackson Baumann, None; Alan Crandall, None; Feryan Ahmed, None; Karen Torrejon, None; David Krizaj, None
  • Footnotes
    Support  EY022076, EY027920, EY014800 Core, the Diabetes and Metabolism Center at the University of Utah, the Willard L. Eccles Foundation, the Moran Eye Institute and unrestricted funds from RPB to the Department of Ophthalmology
Investigative Ophthalmology & Visual Science July 2018, Vol.59, 1660. doi:https://doi.org/
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    • Get Citation

      Oleg Yarishkin, Tam Phuong, Jackson M Baumann, Alan S Crandall, Feryan Ahmed, Karen Yud Torrejon, David Krizaj; Tandem-pore domain potassium channels are components of intracellular signaling pathways, determinants of pH sensitivity and mechanosensitivity of human trabecular meshwork cells.. Invest. Ophthalmol. Vis. Sci. 2018;59(9):1660. doi: https://doi.org/.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Our goal is to identify and characterize the mechanotransducers in the trabecular meshwork (TM), a pressure-sensitive tissue that regulates the dynamics of aqueous fluid outflow in the eye. Here, we studied ion channels that regulate the membrane potential, excitability and pressure sensitivity in TM cells. We sought to establish the molecular link between mechanotransduction, calcium homeostasis and TM contractility that underpin the dynamic regulation of intraocular pressure (IOP).

Methods : Transcript and protein levels in primary human TM cells were assessed with PCR, Western blot and immunocytochemistry. Patch-clamp, high-speed pressure clamp and calcium imaging were used to characterize the electrophysiological features of the mechanotransducers. Outflow facility of human TM was assessed in nano-scaffold model of TM.

Results : TM cells express multiple putative mechanogated channels, among which TRPV4, TREK-1 and TASK-1 mRNA and protein predominate. The membrane potential and steady-state whole-cell current were sensitive to K2P blockers, external pH and short hairpin RNA (shRNA) that target specific K2P isoforms. shRNAs attenuated the whole-cell conductance whereas K2P channel agonists hyperpolarized the membrane potential and increased the transmembrane conductance. Patch clamp and HSPC experiments conducted in the presence of TRPV4 antagonists showed robust activation of outward currents in response to physiological steps of positive pressure. These responses were suppressed by quinine and reduced in cells that overexpressed K2P-specific, but not scrambled, shRNAs. K2P inhibition elevated intracellular Ca2+ levels, an effect that was attenuated by high-K+. The outflow facility in the nano-scaffold model of the healthy human TM was, however, unaffected by quinine.

Conclusions : This study suggests that tensile homeostasis in human TM cells is maintained by the opposing dynamic activation of pressure-sensitive TRPV4 and K2P channels. Mechanical stretch and/or pressure evoke hyperpolarizing shifts that are linked to K2P activation, intracellular Ca2+ homeostasis and thus TM contractility. The pressure sensitivity of the conventional outflow pathway may be, under certain conditions, negatively regulated by K2P channels.

This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.

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