Bicarbonate transport mechanisms in rabbit ciliary body epithelium

Abstract Sections of whole ciliary body dissected from Dutch belted rabbits were incubated with the cell entrappable pH probe BCECEF-AM. This led to a highly specific localization of epifluorescence emission at the exposed, non-pigmented cell layer (npe) of the dual layered epithelium that covers this organ. The BCECF-loaded tissue sections were superfused in a flow-through chamber and the intracellular pH (pH i ) of small groups (10–20) of cells was derived from the ratio of the emission intensities derived from excitations at 490 and 440 nm. In CO 2 /HCO 3 − Ringer's, npe pH i = 7·09±0·11. Replacement of CO 2 /HCO 3 − by Hepes increased pH i by 0·22±0·02, indicating alkali secretory activity under the bicarbonate-rich conditions. Replacement of Cl − by gluconate elicited a rapid, 0·6-U increase in pH i . This effect exhibited little dependence on Na + and was inhibited by 0·5 m m dihydro-4,4′-diisothiocyanatostilbene -2,2′-disulfonate (H 2 DIDS). These results indicate the presence of an electroneutral Cl − /base exchange activity. Elevation of [K + ] (by partial replacement of Na + ) also elicited increases in pH i . In Cl − -free media pH i reached 7·8–8·0, a condition under which intracellular [HCO 3 − ] is at least twice as high as its extracellular value. This effect did not occur in the absence of Na + . The Na + -dependent high [K + ]-induced pH i increase was inhibited by H 2 DIDS. The effects of Ba 2+ on pH i , alone and in combination with high [K + ], as well as that of full K + removal, suggested that the link between high [K + ] and pH i increase was mainly due to the effect of cell depolarization on an electronegative Na + dependent HCO 3 − transporter. Under normal physiological conditions, the two acid/base transport systems are the main determinants of npe pH i .