Evidence That Outwardly Rectifying Cl − Channels Underlie Volume-Regulated Cl − Currents in Heart

Abstract

Swelling-induced Cl⁻ current (I_Cl.swell) is present in most cardiac tissues, but the unitary channel underlying I_Cl.swell is unknown. We used the cell-attached patch-clamp technique to assess the properties of single channels underlying I_Cl.swell and the basally active Cl⁻ current (I_Cl.b) in rabbit atrial myocytes. Under isotonic conditions, single outwardly rectifying Cl⁻ channels (ORCCs) with a slope conductance of 28±1 pS at the reversal potential were observed in 21 (5.7%) of 367 patches. Unconditional kinetic analysis revealed at least three open and four closed-channel states. Hypotonic superfusion-induced swelling resulted in the appearance of active channels in 41 (15.5%) of 265 patches without channel activity under isotonic conditions and caused a second active channel to appear in 3 of 14 patches showing a single channel under isotonic conditions. Overall, channels were seen in 54 of 336 patches under hypotonic conditions (16.1%, P<.001 versus isotonic conditions). The current-voltage relations, reversal potential–[Cl⁻]_o relations, open probability, and kinetics of swelling-induced channels were indistinguishable from those of ORCCs under isotonic conditions. Unitary ORCCs, I_Cl.b, and I_Cl.swell were strongly and similarly inhibited by tamoxifen. Swelling-induced increases in macroscopic Cl⁻ current were attributable to an increase in the number of active ORCCs with no significant effects on single-channel amplitude or open probability. Estimated macroscopic currents based on cell surface area, patch dimensions, single-channel ORCC current amplitude, open probability, and density were consistent with measured values of I_Cl.b and I_Cl.swell. We conclude that ORCCs underlie volume-regulated basal and swelling-induced Cl⁻ currents in isolated rabbit atrial myocytes.