Vacuolar H(+)-ATPase in ocular ciliary epithelium

The mechanisms controlling the production of aqueous humor and the regulation of intraocular pressure are poorly understood. Here, we provide evidence that a vacuolar H(+)-ATPase (V-ATPase) in the ocular ciliary epithelium is a key component of this process. In intracellular pH (pH(i)) measurements of isolated ciliary epithelium performed with 2′,7-biscarboxyethyl-5(6)-carboxyfluorescein (BCECF), the selective V-ATPase inhibitor bafilomycin A(1) slowed the recovery of pH(i) in response to acute intracellular acidification, demonstrating the presence of V-ATPase in the plasma membrane. In isolated rabbit ciliary body preparations examined under voltage-clamped conditions, bafilomycin A(1) produced a concentration-dependent decrease in short-circuit current, and topical application of bafilomycin A(1) reduced intraocular pressure in rabbits, indicating an essential role of the V-ATPase in ciliary epithelial ion transport. Immunocytochemistry utilizing antibodies specific for the B1 isoform of the V-ATPase 56-kDa subunit revealed localization of V-ATPase in both the plasma membrane and cytoplasm of the native ciliary epithelium in both rabbit and rat eye. The regional and subcellular distribution of V-ATPase in specific regions of the ciliary process was altered profoundly by isoproterenol and phorbol esters, suggesting that change in the intracellular distribution of the enzyme is a mechanism by which drugs, hormones, and neurotransmitters modify aqueous humor production.