The Kv1.3 potassium channel lacks N-type inactivation, but during prolonged depolarized periods it inactivates via the slow (P/C-type) mechanism. It bears a titratable histidine residue in position 399 (equivalent of Shaker 449), a site known to influence the rate of slow inactivation. As opposed to several other voltage-gated K⁺ channels, slow inactivation of Kv1.3 is slowed when external pH (pH_o) is lowered in physiological conditions. We have found that: 1. When H399 was mutated to a lysine, arginine, leucine, valine or tyrosine, extracellular acidification (pH 5.5) accelerated inactivation reminiscent of other Kv channels. 2. Inactivation of the wild type channel was accelerated by low pH_o when the ionic strength of the external solution was raised. Inactivation of H399K was also accelerated by high ionic strength at pH 7.35, but not that of H399L. 3. Following the external application of blocking Ba²⁺ ions, recovery of the wild type current during washout was slower in low pH_o. 4. The dissociation rate of Ba²⁺ was pH-insensitive for both H399K and H399L. Furthermore, Ba²⁺ dissociation rates were equal for H399K and the wild type at pH 5.5, and also for H399L and the wild type at pH 7.35. These observations support a model in which the electric field of the protonated histidines creates a potential barrier for K⁺ ions just outside the external mouth of the pore that hinders their exit from the binding site controlling inactivation. In Kv1.3, this effect overrides the generally observed speeding of slow inactivation when pH_o is reduced.