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Abstract #8257 Published in IGR 4-3
Inhibition of carbonic anhydrase activity in cultured bovine corneal endothelial cells by dorzolamide
Srinivas SP; Ong A; Zhai C; Bonanno JAInvestigative Ophthalmology and Visual Science 2002; 43: 3273-3278
PURPOSE: Fluid transport by the corneal endothelium is dependent on the presence of HCO3- and the activity of carbonic anhydrase (CA)-II and -IV, the cytoplasmic and membrane-bound CAs, respectively. This study was conducted to examine the inhibition of CA activity in cultured bovine corneal endothelial cells (BCECs) by dorzolamide, a topical CA inhibitor used in glaucoma therapy. METHODS: BCECs were grown on glass coverslips and then perfused with HCO3- -free Ringer's. The inward flux of CO2 was induced by exposure to CO2-HCO3- Ringer's and the opposing outward flux by returning to HCO3- -free Ringer's. Consequent transients in intracellular pH (pHi) were measured using the pH-sensitive fluorescent dye 2',7'-bis-(2-carboxyethyl)-5-(and-6-_-carboxyfluorescein (BCECF). During the inward flux of CO2, the maximum rate of change of pHi was taken as a quantitative measure of the overall CA activity in BCECs. RESULTS: Exposure to CO2-HCO3- ringer's led to a transient decrease in pHi (component A), followed by a rapid increase to a new steady state (component B). However, when the CO2-HCO3- Ringer's was removed, the pHi increased transiently (component C) and then rapidly returned to the original pHi (component D). Component A, caused by an inward flux of CO2 and its subsequent hydration by CA-II, was blocked by dorzolamide in a dose-dependent manner with a 50% inhibitory concentration (IC)50 of 2.4 μm (95% confidence interval: 0.5-10.85 μm). However, the inhibition of the outward flux of CO2, inward flux of HCO3-, and outward flux of HCO3- (associated with components C, B, and D, respectively) was not dose dependent. Cells that were exposed to 500 nm of the drug for longer than 30 minutes did not show a significantly greater inhibition of any of the components. Dorzolamide and acetazolamide (500 μm) did not show additive inhibition of any of the components (p = 0.13; n = 6). CONCLUSIONS: Dorzolamide significantly inhibits CA activity in BCECs at micromolar levels. Because these levels are encountered in the cornea and aqueous humor after topical administration, dorzolamide may compromise corneal hydration control, especially when the functional reserve of corneal endothelium is low. Dorzolamide does not appear to accumulate in the cells, because the inhibition of CA-II did not increase after prolonged exposure to the drug.
S.P. Srinivas, MD, 800 East Atwater Avenue, Indiana University, School of Optometry, Bloomington, IN 47405, USA. srinivas@indiana.edu
Classification:
11.5.2 Topical (Part of: 11 Medical treatment > 11.5 Carbonic anhydrase inhibitors)