K⁺-, Cl^–- and Nonselective Channels in Opossum Kidney Cells

Abstract

The opossum kidney (OK) cell line serves as a model for proximal tubular cells in many respects. The aim of the present study was to characterize the ion channels found in this cell line and to examine their regulation. OK cells of passages 85–92 were grown on glass coverslips in Dulbecco’s modified Eagle’s medium. Cells were used in a subconfiuent state. Standard patch-clamp experiments were carried out predominantly in the inside/out configuration. Four types of ion channels were found: large-conductance K⁺ channels (bK⁺), intermediate-conductance K⁺ channels (iK⁺), Cl^– channels and nonselective (NS) cation channels. The bK⁺ channel was Ca²⁺-dependent. It had a mean conductance of 174 ± 45 pS (n = 21) at a clamp voltage (V_c) = 0 mV (pipette KCl/bath NaCl). The channel was inhibited by Ba²⁺ (10^–3 mol/l, n = 7) in a voltage-dependent manner; the half-maximal inhibition (IC₅₀) was ≈ 10^–4 mol/l at V_c = 0 mV. Quinidine (10^–5 mol/l, n ≈ 4) as well as verapamil (≧ 10^–5 mol/l, n = 4) and diltiazem (5 × 10^-4 mol/l, n = 4) induced fast flickering and a reduction in mean open probability. Tetraethylammonium+ (10^–2 mol/l) blocked from the outside of the membrane only. The less frequently observed iK⁺ channel had a mean conductance of 62 ± 24 pS (n = 10). The channel showed no Ca²⁺ dependence but inactivation by Mg²⁺ at 10^–3 mol/l. The channel could be blocked reversibly by diltiazem (5 × 10^–4 mol/l, n = 1) and verapamil (10^–4 mol/l, n = 1). The Cl^– channel was outwardly rectifying and had an outward conductance of 51 ± 29 pS (g+) and an inward conductance of 36 ± 17pS (g; 5-Nitro-2-(3-phenylpropylamino)-benzoate at 10^–5 mol/l (n = 3) induced a reversible flicker block. The NS channel had a mean conductance of 27 ± 6 pS (n = 5) and did not rectify.