Three different Cl- channels in the bovine ciliary epithelium activated by hypotonic stress

1. Hypotonic solution induced transient Cl- channel activity in both pigmented and non-pigmented ciliary epithelial cells in cell-attached patches. 2. The activation time constants for these currents were 63 and 97 s for non-pigmented and pigmented ciliary epithelial (NPCE and PCE) cells, respectively. The currents inactivated during the exposure to hypotonic solution with time constants of 59 and 304 s for NPCE and PCE cells, respectively. 3. Single channel analysis revealed the presence of a low-conductance Cl- channel in both the NPCE (7.3 +/- 0.4 pS) and PCE (8.6 +/- 0.9 pS) cells. In addition, an intermediate-conductance (18.8 pS) channel was found alone in the NPCE cells and a maxi Cl- channel (105 pS) was found in the PCE cells. The similarities between the 18.8 pS Cl- channel and the volume-activated organic osmolyte-anion channel (VSOAC) are discussed. 4. The hypotonic activation of all three Cl- channels was prevented by the inclusion of 100 microM 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) in the patch pipette. 5. Inclusion of tamoxifen in the patch pipette inhibited only the intermediate-conductance channel in the NPCE cells. This is consistent with the identification of the intermediate 18 pS Cl- channel, found only in NPCE cells, as VSOAC. 6. Negative pressure also evoked single Cl- channel activity but the activation kinetics were quite different, suggesting that, even if the channels are the same, the second messenger pathways involved are different. 7. The activation/inactivation kinetics of these Cl- channels correlate well with the time course of regulatory volume decrease (RVD) in these cells, suggesting that these channels may well participate in the process of RVD. 8. The asymmetrical distribution of Cl- channels in the ciliary epithelium may be significant for aqueous humour secretion. For instance, the presence of the VSOAC-like channel in the NPCE cells may allow one-way traffic of anions and organic osmolytes into the eye and, if these solutes were loaded by the PCE cells, then a vectorial flow from the blood into the eye would occur.