PURPOSE. To investigate the role of active chloride secretion by the ciliary epithelium in the formation of aqueous humor (AH), by using the in vitro perfused eye. METHODS. Bovine eyes collected from an abattoir were cannulated through the ophthalmic artery and perfused with oxygenated Krebs' solution at 37°C. Aqueous humor formation (AHF) was measured by the fluorescein-dilution technique. Drugs were added to the perfusate and/or to the anterior chamber. RESULTS. NaK-adenosine triphosphatase (ATPase) inhibitor, ouabain (1.0 mM), produced a significant reduction in AHF by 46% and 42% when added to the stromal or aqueous side, respectively. When added to both sides (1.0 mM), it produced a reduction of 61%. Bumetanide (0.1 mM), a specific inhibitor of Na-K-2Cl cotransport, and furosemide (0.1 mM), a nonspecific anion transport inhibitor, produced 35% and 45% reductions when applied to the stromal side. DIDS (0.001-0.1 mM), which is believed to inhibit the Cl-HCO3 exchanger, Na-HCO3 cotransporter, and chloride channel, produced a dose-dependent reduction when added to the stromal side. The inhibition was 55% by the highest concentration used. 5-Nitro-2-(3-phenylpropylamino)-benzoic acid (NPPB; 0.1 mM, a chloride channel blocker in the nonpigmented cells, produced a 25% reduction when applied to the aqueous side. Acetazolamide (0.1 mM), a carbonic anhydrase inhibitor, applied to the stromal side, produced 31% reduction. CONCLUSIONS. At least 60% of the AH is formed by active secretion in bovine eyes. Transport of anions through the ciliary epithelium (CE), particularly the chloride ion, plays a crucial role in AHF.
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