Cardiac m2 muscarinic acetylcholine receptors reduce heart rate by coupling to heterotrimeric (alpha beta gamma) guanine nucleotide-binding (G) proteins that activate IKACh, an inward rectifier K+ channel (IRK). Activation of the GIRK subunit of IKACh requires G beta gamma subunits; however, the structural basis of channel regulation is unknown. To determine which sequences confer G beta gamma regulation upon IRKs, we generated chimeric proteins composed of GIRK and RB-IRK2, a related, G protein-insensitive channel. Importantly, a chimeric channel containing the hydrophobic pore region of RB-IRK2 joined to the amino and carboxyl termini of GIRK exhibited voltage- and receptor-dependent activation in Xenopus oocytes. Furthermore, carboxy-terminal sequences specific to this chimera and GIRK bound G beta gamma subunits in vitro. Thus, G beta gamma may regulate IRKs by interacting with sequences adjacent to the putative channel pore.