NA-CL--DEPENDENT AND HCO3--DEPENDENT BASE UPTAKE IN THE CILIARY BODY PIGMENT-EPITHELIUM()

Segments of whole ciliary body dissected from Dutch belted rabbits wer e incubated for 60 min at 36 degrees C in a 30 mu M Ca2+ Ringer's. The inner limiting membrane with its adherent non-pigmented epithelium th en was mechanically removed from the surface. The newly-exposed viable layer of pigmented cells was then loaded with the fluorescent probe 2 '-7'-bis (carboxymethyl)-5(6) carboxyfluorescein by incubating the seg ments for 45 min at RT with the cell permeable acetoxymethoxy form of the dye. These loaded tissues were perfused in a now-through chamber. mounted on the heated stage of a microscope equipped for quantitative epifluorescence, and the intracellular pH (pH(i)) of small groups of c ells was derived from the ratio of emission intensities generated by e xcitations at 490 and 440 nm, respectively. In N[2-hydroxyethyl] piper azine-N''-[2 ethane sulfonic acid] (Hepes)-buffered Ringer's the intra cellular pH was 7.23 +/- 0.21 (+/- S.D., n = 20). Replacement of 28 mM Hepes by 28 mM HCO3-/5% CO2 led to a 0.14 +/- 0.04 increase in pH(i). This increase required the presence of Na+ and Cl- and was inhibited by 0.2 mM di-isothiocyanatostilbene-2-2'-disulfonic acid. These observ ations as well as characteristic pH(i) responses to the removal or int roduction of Na+ or Cl- indicated the presence in the pigmented cells of a Na+- and Cl--dependentHCO(3)(-) transporter responsible for base uptake.