Hyposmotically activated chloride channels in cultured rabbit non‐pigmented ciliary epithelial cells

1 We used whole‐cell patch‐clamp recording techniques and noise analysis of whole‐cell current to investigate the properties of hyposmotic shock (HOS)‐activated Cl− channels in SV40‐transformed rabbit non‐pigmented ciliary epithelial (NPCE) cells. 2 Under conditions designed to isolate Cl− currents, exposure of cells to hyposmotic external solution reversibly increased the whole‐cell conductance. 3 The whole‐cell current activated with a slow time course (> 15 min), exhibited outward rectification and was Cl− selective. 4 The disulphonic stilbene derivatives 4,4′‐diisothiocyanatostilbene‐2,2′‐disulfonic acid (DIDS, 0·5 mM), 4‐acetamido‐4′‐isothiocyanatostilbene‐2,2′‐disulfonic acid (SITS, 0·5 mM) and 4,4′‐dinitrostilbene‐2,2′‐disulfonic acid (DNDS, 0·5 mM) produced a voltage‐sensitive block of HOS‐activated Cl− current at depolarized potentials, whereas niflumic acid produced a voltage‐independent block of the current. 5 Under Ca2+‐free conditions, HOS stimulation still reversibly activated the Cl− current, but the amplitude of current was reduced and the time course of current activation was slower compared with control (P < 0·05). 6 The non‐specific kinase inhibitor H‐7 (100 μM), upregulated HOS‐activated Cl− current amplitude in all cells tested (P < 0·05). 7 Noise analysis of whole‐cell Cl− current indicated that cell swelling activated a high density of small conductance Cl− channels (< 1 pS). 8 We conclude that HOS primarily activates a high density of volume‐sensitive small conductance Cl− channels in rabbit NPCE cells, and that Ca2+ and phosphorylation are involved in channel regulation.