Different Roles of Carbonic Anhydrase in Human vs. Bovine Corneal Endothelial Transport

Abstract

Purpose: Regulation of fluid homeostasis in the cornea is critical for maintaining corneal clarity. Buffering by carbonic anhydrases plays a central role in rabbit and bovine models of corneal endothelial fluid transport. Topical carbonic anhydrase inhibitors (CAIs) in rabbits result in decreased fluid efflux measured by increases in corneal thickness. However, routine use of topical and systemic CAIs for glaucoma in humans does not increase corneal thickness. The hypothesis tested here is that there are species differences in the role of carbonic anhydrases in corneal endothelial transport.

Methods: Protocols were approved by the R&D Committee (VAMC, Buffalo, NY). Bovine eyes were obtained from local abattoirs. De-identified human corneas not suitable for transplant were obtained from the local eye bank. Corneal endothelial transport was measured using the short-circuit current (I_sc) technique. The recording solution was (in mM): 111.6 NaCl, 29 NaHCO₃, 4.8 KCl, 1.0 CaCl₂, 0.8 MgCl₂, 0.9 NaH₂PO₄, 10 HEPES, 5 glucose, bubbled with 5% CO₂ / 95% air, pH 7.5. CAIs were added (500 µM acetazolamide, 100 µM ethoxzolamide, 100 µM dorzolamide, or 100 µM brinzolamide) and I_sc measured. Water and DMSO were used as controls. Drug effect was the percentage change of total I_sc.

Results: All CAIs generated significant (p<0.0001) rapid declines in I_sc in bovine corneal endothelium compared to water or DMSO (mean % change ± S.D.: ethoxzolamide −35.5 ± 12.9, n = 8; dorzolamide −33.6 ± 7.2, n = 8; brinzolamide −35.5 ± 13.5, n = 9; acetazolamide −21 ± 9.5, n = 8; water −1.6 ± 4.3, n = 7; DMSO +1.3 ± 4.8, n = 8). In contrast, there was no rapid decline in human (mean % change ± S.D.: ethoxzolamide −4.8 ± 10.3, n = 2; dorzolamide +8.0 ± 20.4, n = 3; brinzolamide +6.7 ± 13.9, n = 3; acetazolamide +2.7 ± 38.9, n = 6). The I_sc differences between bovine and human were significant (p < 0.05) for all CAIs except acetazolamide (p = 0.12).

Conclusions: Carbonic anhydrases support one-third of active ion transport in bovine corneal endothelium but do not in human. This finding suggests distinct mechanisms of ion transport are operational in corneal endothelium in different species and supports the clinical observation that therapeutic use of CAIs does not disrupt corneal fluid homeostasis in humans. Such species differences will be instrumental in guiding future research and development in corneal endothelial disease.

Keywords: 481 cornea: endothelium • 443 carbonic anhydrase • 503 drug toxicity/drug effects