INTERACTION OF PERMEANT AND BLOCKING LONE IN CLONED INWARD-RECTIFIER K+ CHANNELS

Blocking cloned inward-rectifier potassium (K-ir) channels from the cy toplasmic side was analyzed with a rapid application system exchanging the intracellular solution on giant inside-out patches from Xenopus o ocytes in <2 ms. Dependence of the pore-block on interaction of the bl ocking molecule with permeant and impermeant ions on either side of th e membrane was investigated in K(ir)1.1 (ROMK1) channels blocked by am monium derivatives and in K(ir)4.1 (BIR10) channels blocked by spermin e. The blocking reaction in both systems showed first-order kinetics a nd allowed separate determination of on-and off-rates. The off-rates o f block were strongly dependent on the concentration of internal and e xternal bulk ions, but almost independent of the ion species at the cy toplasmic side of the membrane. With K+ as the only cation on both sid es of the membrane, off-rates exhibited strong coupling to the K+ reve rsal potential (E-K) and increased and decreased with reduction in int ra and extracellular K+ concentration, respectively. The on-rates show ed significant dependence on concentration and species of internal bul k ions. This control of rate-constants by interaction of permeant and impermeant internal and external ions governs the steady-state current -voltage relation (I-V) of K-ir channels and determines their physiolo gical function under various conditions.