Mechanism of Ion Permeation in Skeletal Muscle Chloride Channels

Voltage-gated Cl(−) channels belonging to the ClC family exhibit unique properties of ion permeation and gating. We functionally probed the conduction pathway of a recombinant human skeletal muscle Cl(−) channel (hClC-1) expressed both in Xenopus oocytes and in a mammalian cell line by investigating block by extracellular or intracellular I(−) and related anions. Extracellular and intracellular I(−) exert blocking actions on hClC-1 currents that are both concentration and voltage dependent. Similar actions were observed for a variety of other halide (Br(−)) and polyatomic (SCN(−), NO(3) (−), CH(3)SO(3) (−)) anions. In addition, I(−) block is accompanied by gating alterations that differ depending on which side of the membrane the blocker is applied. External I(−) causes a shift in the voltage-dependent probability that channels exist in three definable kinetic states (fast deactivating, slow deactivating, nondeactivating), while internal I(−) slows deactivation. These different effects on gating properties can be used to distinguish two functional ion binding sites within the hClC-1 pore. We determined K (D) values for I(−) block in three distinct kinetic states and found that binding of I(−) to hClC-1 is modulated by the gating state of the channel. Furthermore, estimates of electrical distance for I(−) binding suggest that conformational changes affecting the two ion binding sites occur during gating transitions. These results have implications for understanding mechanisms of ion selectivity in hClC-1, and for defining the intimate relationship between gating and permeation in ClC channels.