1. The whole‐cell patch clamp technique was used to investigate the swelling‐activated currents in bovine non‐pigmented ciliary epithelial (NPCE) cells. 2. Exposure to hypotonic solution activated a current that was blocked by 5‐nitro‐2‐(3‐phenylpropylamino)benzoic acid (NPPB). The I‐V relationship was shifted in the direction expected for a Cl‐ current when the external Cl‐ was replaced by gluconate (permeability ratio P(gluconate)/PCl = 0.17). The inhibition of the current evoked by voltage clamp steps of +80 mV yielded an IC50 for NPPB of 13.4 microM. 3. The current was found to be dependent on ATP. With ATP in the patch pipette the current could be repeatedly activated by exposure to hypotonic solution but when ATP was omitted the current ran down with time. 4. The development of this current was associated with visible cell swelling and inhibitors of regulatory volume decrease in these cells, e.g. tamoxifen, 4‐acetamido‐4'‐isothiocyanatostilbene‐2,2'‐disulphonic acid (SITS) and 4,4'‐diisothiocyanostilbene‐2,2'‐disulphonic acid (DIDS), also inhibited this current. 5. The volume‐activated current was additionally blocked by NPPB, verapamil, quinidine and dideoxyforskolin. 6. The current was independent of external calcium and exhibited slight outward rectification and time‐dependent inactivation at strong depolarizing potentials. 7. Disrupting the cytoskeleton and microtubules with cytochalasin B and colchicine had no effect on the activation of the Cl‐ current. 8. An antibody (C219) to the MDR1 gene product, P‐glycoprotein, caused a functional block of the swelling‐activated Cl‐ current when added to the patch pipette. 9. Immunofluorescence studies using the monoclonal antibodies C219 and JSB‐1 demonstrated the presence of P‐glycoprotein in the ciliary epithelial cells. The immunofluorescence was stronger on the non‐pigmented than on the pigmented cells. 10. It is concluded that swelling in NPCE cells activates a Ca(2+)‐independent, ATP‐dependent Cl‐ current and that the activity of this current is associated with P‐glycoprotein. 11. It is suggested that this Cl‐ current contributes to regulatory volume decrease and may participate in the secretory activity of these cells.
In this report, we present the characteristics of a Cl- channel found in lens fiber cells. The single channel has a conductance of 17 pS, a linear current-voltage curve, is activated by ATP or strong depolarization and is blocked by verapamil, quinidine, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, 5-nitro-2-(3- phenylpropylamino)benzoate, dideoxyforskolin, and tamoxifen. These properties are similar to those reported for a volume-activated Cl- channel associated with the multidrug resistance (MDR) gene product, P glycoprotein (24). Confirming this connection, we demonstrate that our lens Cl- channel is inhibited by an antibody to P glycoprotein. The data we present here may, therefore, be the first characterization of the single channel activity of the Cl- channel associated with P glycoprotein.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.