External Ba 2+ Block of Human Kv1.5 at Neutral and Acidic pH: Evidence for H o+-Induced Constriction of the Outer Pore Mouth at Rest

Abstract

Previous studies have shown that low pH o accelerates depolarization-induced inactivation and decreases the macroscopic conductance by reducing channel availability. To test the hypothesis that outer pore constriction underlies the decreased conductance at low pH o, external Ba 2+ was used to examine the accessibility of the channel pore at rest under neutral and acidic conditions. At pH o 7.4, Ba 2+ block of closed channels follows a monoexponential time course and involves a low-affinity superficial site ( K D ≅ 1 mM, −80 mV, 0 mM K o + ) and a high-affinity site ( K D ≅ 4 μM) deeper in the pore. Depolarization promotes Ba 2+ dissociation and an analytical model incorporating state-dependent changes of Ba 2+ affinity is presented that replicates the frequency dependence of the time course and the extent of block. Open-channel block by Ba 2+ is weak. At pH o 5.5, both the access to and exit from the deep site is inhibited. These results are consistent with a low-pH o-induced conformational change in the outer pore that prevents Ba 2+ binding at rest or unbinding during depolarization. If a pore constriction is involved, similar to that proposed to occur during P/C-type inactivation, this would imply that closed-state inactivation in Kv1.5 occurs under acidic conditions.