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Departments of 1 Physiology and 2 Medicine, University of Pennsylvania School of Medicine, Philadelphia, Pennsylvania 19104-6085; and 3 Department of Ophthalmology, University of Arizona, Tucson, Arizona 85711-1824
The volume of
certain subpopulations of trabecular meshwork (TM) cells may modify
outflow resistance of aqueous humor, thereby altering intraocular
pressure. This study examines the contribution that
Na+/H+, Cl
/HCO
efflux mechanisms have on
the volume of TM cells. Volume, Cl currents, and
intracellular Ca2+ activity of cultured human TM cells were
studied with calcein fluorescence, whole cell patch clamping, and fura
2 fluorescence, respectively. At physiological bicarbonate
concentration, the selective Na+/H+ antiport
inhibitor dimethylamiloride reduced isotonic cell volume. Hypotonicity
triggered a regulatory volume decrease (RVD), which could be inhibited
by the Cl channel blocker
5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB), the K+
channel blockers Ba2+ and tetraethylammonium, and the
K+-Cl symport blocker
[(dihydroindenyl)oxy]alkanoic acid. The fluid uptake mechanism in
isotonic conditions was dependent on bicarbonate; at physiological
levels, the Na+/H+ exchange inhibitor
dimethylamiloride reduced cell volume, whereas at low levels the
Na+-K+-2Cl symport inhibitor
bumetanide had the predominant effect. Patch-clamp measurements showed
that hypotonicity activated an outwardly rectifying, NPPB-sensitive
Cl channel displaying the permeability ranking
Cl > methylsulfonate > aspartate.
2,3-Butanedione 2-monoxime antagonized actomyosin activity and both
increased baseline [Ca2+] and abolished
swelling-activated increase in [Ca2+], but it did not
affect RVD. Results indicate that human TM cells display a
Ca2+-independent RVD and that volume is regulated by
swelling-activated K+ and Cl channels,
Na+/H+ antiports, and possibly
K+-Cl symports in addition to
Na+-K+-2Cl symports.
outflow facility; calcein; chloride channels; potassium-chloride symport; sodium/hydrogen antiport; methylsulfonate; aspartate; intraocular pressure; [(dihydroindenyl)oxy]alkanoic acid
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