Studies on bicarbonate transporters and carbonic anhydrase in porcine nonpigmented ciliary epithelium.
OBJECTIVE
Bicarbonate transport plays a role in aqueous humor (AH) secretion. The authors examined bicarbonate transport mechanisms and carbonic anhydrase (CA) in porcine nonpigmented ciliary epithelium (NPE).


METHODS
Cytoplasmic pH (pH(i)) was measured in cultured porcine NPE loaded with BCECF. Anion exchanger (AE), sodium bicarbonate cotransporter (NBC), and CA were examined by RT-PCR and immunolocalization. AH secretion was measured in the intact porcine eye using a fluorescein dilution technique.


RESULTS
Anion exchanger AE2, CAII, and CAIV were abundant in the NPE layer. In cultured NPE superfused with a CO(2)/HCO(3)(-)-free HEPES buffer, exposure to a CO(2)/HCO(3)(-)-containing buffer caused rapid acidification followed by a gradual increase in pH(i). Subsequent removal of CO(2)/HCO(3)(-) with HEPES buffer caused rapid alkalinization followed by a gradual decrease in pH(i). The rate of gradual alkalinization after the addition of HCO(3)(-)/CO(2) was inhibited by sodium-free conditions, DIDS, and the CA inhibitors acetazolamide and methazolamide but not by the Na-H exchange inhibitor dimethylamiloride or low-chloride buffer. The phase of gradual acidification after removal of HCO(3)(-)/CO(2) was inhibited by DIDS, acetazolamide, methazolamide, and low-chloride buffer. DIDS reduced baseline pH(i). In the intact eye, DIDS and acetazolamide reduced AH secretion by 25% and 44%, respectively.


CONCLUSIONS
The results suggest the NPE uses a Na(+)-HCO(3)(-) cotransporter to import bicarbonate and a Cl(-)/HCO(3)(-) exchanger to export bicarbonate. CA influences the rate of bicarbonate transport. AE2, CAII, and CAIV are enriched in the NPE layer of the ciliary body, and their coordinated function may contribute to AH secretion by effecting bicarbonate transport into the eye.
Shahidullah
Mohammad
M
Department of Physiology, University of Arizona, Tucson, Arizona 85724, USA. shahidua@email.arizona.edu
To
Chi-Ho
CH
Pelis
Ryan M
RM
Delamere
Nicholas A
NA
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000494:pharmacology; D000086:Acetazolamide / Q000494:pharmacology; D000818:Animals; D001082:Aqueous Humor / Q000378:metabolism*; D001639:Bicarbonates / Q000378:metabolism*; D001693:Biological Transport, Active / Q000502:physiology; D002256:Carbonic Anhydrases / Q000502:physiology*; D002478:Cells, Cultured; D027963:Chloride-Bicarbonate Antiporters / Q000502:physiology*; D002924:Ciliary Body / Q000187:drug effects / Q000378:metabolism*; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D005452:Fluoresceins / Q000378:metabolism; D005456:Fluorescent Dyes / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D020133:Reverse Transcriptase Polymerase Chain Reaction; D027982:Sodium-Bicarbonate Symporters / Q000502:physiology*; D013552:Swine
D001639:Bicarbonates; D027963:Chloride-Bicarbonate Antiporters; D005452:Fluoresceins; D005456:Fluorescent Dyes; D027982:Sodium-Bicarbonate Symporters; C043829:2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein; D002256:Carbonic Anhydrases; D000086:Acetazolamide; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid