The somatodendritic A-type current (ISA ) in hippocampal CA1 pyramidal neurons is mediated by Kv4.2 channels. Subthreshold activation of these channels dampens backpropagating action potentials. Their inactivation allows coincidence detection of pre- and postsynaptic activity. Closed-state inactivation of Kv4.2 channels in response to weak synaptic input makes coincidence detection extremely sensitive. However, the mechanism underlying closed-state inactivation has remained unresolved. We tested if uncoupling of the voltage sensor from the activation gate may be involved in Kv4.2 closed-state inactivation. We performed an alanine-scanning mutagenesis within the S4-S5 linker region and the distal S6 segment and functionally characterized mutant Kv4.2 channels under two-electrode voltage- clamp in Xenopus oocytes. Double-mutant cycle analysis showed that pairs of residues involved in gate opening also play a role in closed-state inactivation of Kv4.2 channels. A corresponding double-cysteine mutant (Kv4.2E323C,V404C) was studied under reducing (DTT) and oxidizing conditions (tbHO2). We observed slowed inactivation rates under oxidizing conditions which favor the formation of disulfide bonds. Our findings support the idea that temporary uncoupling of voltage sensor and activation gate underlies Kv4.2 closed-state inactivation. Supported by the Deutsche Forschungsgemsinschaft (Ba2055/1-1,2; Ba2055/1-3)