Polyamine block completely inhibits the outward currents of Kir2.1 at 0 [K⁺]_out. (A) Outward K⁺ currents and their I–V relationships obtained from an inside-out patch membrane using K⁺-free (150 mM Na⁺) external solution ∼60 s after excising the patch in polyamine-free, 150 mM K⁺ solution. Currents were not observed immediately after patch excision. Test pulses were applied from a holding potential of −100 mV. Subsequent disappearance of the outward currents with K⁺-free (150 mM Na⁺) internal solution and their recovery are also shown. (B) Inhibition of the outward currents observed under the external K⁺-free (150 mM Na⁺) condition with subsequent application of 0.1–10 µM cytoplasmic spermine. Currents were recorded from an inside-out patch membrane using test pulses applied from a holding potential of −80 mV and followed by a step pulse to 0 mV. Left: Selected current traces obtained at test potentials between −100 and 20 mV in 10-mV steps. Top right: I–V relationships obtained at the end of test pulses. Bottom right: Amplitudes of tail currents during the subsequent step to 0 mV, reflecting fractional openings of Kir2.1 at the end of the test pulses. Dashed lines superimposed on current traces indicate the zero-current level. Data are representative of eight independent experiments.