Extracellular Zinc Ion Inhibits ClC-0 Chloride Channels by Facilitating Slow Gating

Extracellular Zn(2+) was found to reversibly inhibit the ClC-0 Cl(−) channel. The apparent on and off rates of the inhibition were highly temperature sensitive, suggesting an effect of Zn(2+) on the slow gating (or inactivation) of ClC-0. In the absence of Zn(2+), the rate of the slow-gating relaxation increased with temperature, with a Q(10) of ∼37. Extracellular Zn(2+) facilitated the slow-gating process at all temperatures, but the Q(10) did not change. Further analysis of the rate constants of the slow-gating process indicates that the effect of Zn(2+) is mostly on the forward rate (the rate of inactivation) rather than the backward rate (the rate of recovery from inactivation) of the slow gating. When ClC-0 is bound with Zn(2+), the equilibrium constant of the slow-gating process is increased by ∼30-fold, reflecting a 30-fold higher Zn(2+) affinity in the inactivated channel than in the open-state channel. As examined through a wide range of membrane potentials, Zn(2+) inhibits the opening of the slow gate with equal potency at all voltages, suggesting that a two-state model is inadequate to describe the slow-gating transition. Following a model originally proposed by Pusch and co-workers (Pusch, M., U. Ludewig, and T.J. Jentsch. 1997. J. Gen. Physiol. 109:105–116), the effect of Zn(2+) on the activation curve of the slow gate can be well described by adding two constraints: (a) the dissociation constant for Zn(2+) binding to the open channel is 30 μM, and (b) the difference in entropy between the open state and the transition state of the slow-gating process is increased by 27 J/ mol/°K for the Zn(2+)-bound channel. These results together indicate that extracellular Zn(2+) inhibits ClC-0 by facilitating the slow-gating process.