June 2013
Volume 54, Issue 15
Free
ARVO Annual Meeting Abstract  |   June 2013
P2X channels contribute to a non-selective cation (NSC) conductance in isolated lens fiber cells that is activated by cell shrinkage
Author Affiliations & Notes
  • Paul Donaldson
    Optometry and Vision Science, University of Auckland, Auckland, New Zealand
    School of Medical Sciences, University of Auckland, Auckland, New Zealand
  • Simon Gunning
    Optometry and Vision Science, University of Auckland, Auckland, New Zealand
  • Footnotes
    Commercial Relationships Paul Donaldson, None; Simon Gunning, None
  • Footnotes
    Support None
Investigative Ophthalmology & Visual Science June 2013, Vol.54, 3681. doi:https://doi.org/
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      Paul Donaldson, Simon Gunning; P2X channels contribute to a non-selective cation (NSC) conductance in isolated lens fiber cells that is activated by cell shrinkage. Invest. Ophthalmol. Vis. Sci. 2013;54(15):3681. doi: https://doi.org/.

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

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Abstract

Purpose: To determine whether members of the P2X family of ATP gated non-selective cation (NSC) channels, that insert into the membranes of fiber cells in response to osmotic stress(1), contribute to a Gd3+-insensitive, La3+-sensitive NSC conductance that is activated by cell shrinkage in isolated fiber cells(2).

Methods: Immunohistochemistry was used to visualise the subcellular distribution of P2X channels in sections taken from rat lenses incubated in isotonic and hypertonic Artificial Aqueous Humor. Differentiating fiber cells were isolated from rat lenses in the presence of the NSC channel inhibitor Gd3+, and the changes in membrane currents following exposure of cells to P2X agonists (ATP; α,βMetATP) and/or antagonists (PPADS, NF449), before and after hypertonic challenge recorded by whole cell patch clamping(2).

Results: In whole lenses antibody labelling for P2X1 and P2X4 was predominately cytoplasmic under isotonic conditions, but became associated with the broad-side membranes of fiber cells following hypertonic challenge. In isolated fiber cells incubated in Gd3+ a residual NSC conductance persisted which was further enhanced by cell shrinkage and completely inhibited by La3+. Under isotonic conditions addition of modulators of P2X activity had no significant effect on the La3+-sensitive NSC conductance indicating that this basal conductance is not mediated by P2X channels, a result consistent with the cytoplasmic location of P2X channels observed in whole lenses. In contrast, after exposure of cells to hypertonic challenge, the addition of the P2X agonists ATP or α,βMetATP caused a significant additional increase in the La3+-sensitive NSC conductance. This additional ATP-sensitive component observed following hypertonic challenge was blocked by the general P2X inhibitor PPADS, and the P2X1-specific blocker NF449.

Conclusions: P2X1/4 receptors inserted into the broad side membranes following hypertonic stress are functionally active and distinct from the Gd3+-insensitive, but La3+-sensitive NSC conductance. The insertion of P2X channels in response to osmotic challenge suggests these channels are involved in modulating lens volume, a key requirement for the maintenance of lens transparency. (1) Suzuki-Kerr et al. Invest Ophthalmol Visual Sci 51:4156-63, 2010 (2) Gunning et al. Am J Physiol - Cell Physiol Oct 10, 2012. [Epub ahead of print]

Keywords: 569 ion channels • 599 microscopy: light/fluorescence/immunohistochemistry • 508 electrophysiology: non-clinical  
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