Effect of prolonged depolarization on the current amplitude. Panels show representative current recordings for WT hKv2.1 (a), WT hKv3.1 (b), and their corresponding hKv2.1-T377A and hKv3.1-T400A mutants. Current recordings were elicited with the pulse protocols shown on top. (a) A prolonged depolarization induces U-type inactivation in hKv2.1, which manifests as a time-dependent reduction in current amplitude (black trace). This decrease in current amplitude can be quantified by the current elicited during a subsequent test pulse at +60 mV, after a conditioning test pulse to +40 mV (black trace) or to −80 mV (gray trace). Inset on the right shows an enlarged view of the currents elicited during the test pulse to +60 mV. The magnitude of the current reduction during the test pulse at +60 mV is proportional to the size of the black arrow. For the mutant hKv2.1-T377A, the current amplitude slowly increased during membrane depolarization at +40 mV (red trace), noticeable by the increase in current amplitude (red arrow) during the +60 mV test pulse between a conditioning prepulse to −80 (blue) or +40 mV (red). (b) The current of WT hKv3.1, alike to hKv2.1, decreased in a time-dependent fashion upon prolonged depolarization at +40 mV (black trace). In contrast, hKv3.1-T400A did not display current reduction (red trace). Black arrow highlights, again, the amount of current reduction for WT hKv3.1 between a prepulse to −80 mV (gray trace) and +40 mV (black trace). The absence of current inactivation for hKv3.1-T400A between a prepulse to −80 mV (blue) and +40 mV (red) is indicated with the red arrow.