Residues and mechanisms for slow activation and Ba2+ block of the cardiac muscarinic K+ channel, Kir3.1/Kir3.4

Mechanisms and residues responsible for slow activation and Ba2+ block of t he cardiac muscarinic K+ channel, Kir3.1/Kir3.4, were investigated using si te-directed mutagenesis. Mutagenesis of negatively charged residues located throughout the pore of the channel (in H5, M2, and proximal C terminus) re duced or abolished slow activation. The strongest effects resulted from mut agenesis of residues in H5 close to the selectivity filter; mutagenesis of residues in M2 and proximal C terminus equivalent to those identified as im portant determinants of the activation kinetics of Kir2.1 was less effectiv e. In giant patches, slow activation was present in cell-attached patches, lost on excision of the patch, and restored on perfusion with polyamine, Mu tagenesis of residues in H5 and M2 close to the selectivity filter also dec reased Ba2+ block of the channel. A critical residue for Ba2+ block was ide ntified in Kir3.4. Mutagenesis of the equivalent residue in Kir3.1 failed t o have as pronounced an effect on Ba2+ block, suggesting an asymmetry of th e channel pore. It is concluded that slow activation is principally the res ult of unbinding of polyamines from negatively charged residues close to th e selectivity filter of the channel and not an intrinsic gating mechanism. Ba2+ block involves an interaction with the same residues.