MECHANISM OF ION PERMEATION IN SKELETAL-MUSCLE CHLORIDE CHANNELS

Voltage-gated Cl- channels belonging to the CIC family exhibit unique properties of ion permeation and gating. We functionally probed the co nduction pathway of a recombinant human skeletal muscle Cl- channel (h ClC-1) expressed both in Xenopus oocytes and in a mammalian cell line by investigating block by extracellular or intracellular I- and relate d anions. Extracellular and intracellular T-exert blocking actions on hClC-1 currents that are both concentration and voltage dependent. Sim ilar actions were observed for a variety of other halide (Br-) and pol yatomic (SCN-, NO3-, CH3SO3-) anions. In addition, I-block is accompan ied by gating alterations that differ depending on which side of the m embrane the blocker is applied. External I- causes a shift in the volt age-dependent probability that channels exist in three definable kinet ic states (fast deactivating, slow deactivating, nondeactivating), whi le internal I- slows deactivation. These different effects on gating p roperties 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 mod ulated by the gating state of the channel. Furthermore, estimates of e lectrical distance for I- binding suggest that conformational changes affecting the two ion binding sites occur during gating transitions. T hese results have implications for understanding mechanisms of ion sel ectivity in hClC-1, and for defining the intimate relationship between gating and permeation in ClC channels.