The effects of low intracellular pH (pH(i) 6.4) on cloned small-conductance Ca2+-activated K+ channel currents of all three subtypes (SK1, SK2, and SK3) were investigated in HEK293 cells using the patch-clamp technique. In 400 nM internal Ca2+ [Ca2+](i), all subtypes were inhibited by pH(i) 6.4 in the order of sensitivity: SK1 > SK3 > SK2. The inhibition increased with the transmembrane voltage. In saturating internal Ca2+, the inhibition was abolished for SK1-3 channels at negative potentials, indicating a [Ca2+](i)-dependent mode of inhibition. Application of 50 mu M 1-ethyl-2-benzimidazolone was able to potentiate SK3 current to the same extent as at neutral pHi. We conclude that SK1-3 all are inhibited by low pHi. We suggest two components of inhibition: a [Ca2+](i)-dependent component, likely involving the SK beta-subunits calmodulin, and a voltage-dependent component, consistent with a pore-blocking effect. This pH(i)-dependent inhibition can be reversed pharmacologically. (c) 2006 Elsevier Inc. All rights reserved.