Specificity of action of guanine nucleotide-binding regulatory protein subunits on the cardiac muscarinic K+ channel.

The cardiac muscarinic receptor stimulates a potassium-selective ionic current (IK.ACh) through activation of a guanine nucleotide-binding regulatory protein. Purified alpha and beta gamma subunits of the guanine nucleotide-binding regulatory protein have each been reported to open the K+ channel. We have reported that nanomolar concentrations of purified brain beta gamma subunits activated IK.ACh in chicken embryonic atrial patches. In contrast, J. Codina, A. Yatani, D. Grenet, A.M. Brown, and L. Birnbaumer [(1987) Science 236, 442-445] subsequently reported that picomolar concentrations of activated erythrocyte alpha subunits (i.e., the 40-kDa alpha subunit that the authors call alpha K) opened K+ channels in guinea pig atrial patches. In this paper, we further explore the specificity of various beta gamma and alpha subunits in embryonic chicken and neonatal rat atrial patches. Beta gamma subunits from either human placenta (beta 35 gamma) or bovine brain (beta 35,36 gamma) activated IK.ACh whereas transducin beta gamma (beta 36 gamma) did not. The beta gamma activation was consistent in rat and chicken patches [118 of 123 patches (97%)]. Beta gamma subunits opened K+ channels at concentrations greater than or equal to 200 pM and maximally activated the channel at 10 nM. Beta gamma or guanosine 5'-[gamma-thio]triphosphate (GTP[gamma-S]) channel activation could be reversed by alpha 41-GDP. The purified brain beta gamma preparation was contaminated with less than 0.01% unactivated alpha. The detergent (3-[(3-cholamidopropyl)dimethylammonio]-1-propanesulfonate; CHAPS), used to suspend the hydrophobic beta gamma, did not activate IK.ACh alone, with buffer, with heat-inactivated beta gamma, or with transducin beta gamma. Unactivated alpha subunits did not open K+ channels. Activated, alpha subunits purified from human erythrocytes (alpha 40-GTP[gamma-S]) or bovine brain (alpha 39-GTP[gamma-S]) at concentrations of 10 pM or higher (up to 1 nM) opened K+ channels less frequently in chicken atrial patches [5 of 27 patches (19%) and 9 of 35 patches (26%), respectively] than in rat atrial patches [5 of 11 patches (45%) and 11 of 19 patches (58%), respectively]. Negative results were not due to patch vesicle formation. Other experiments indicated that alpha and beta gamma activated the same population of channels. Activation of the channel by both beta gamma and alpha subunits implies a more complicated scheme for guanine nucleotide-binding regulatory protein action than previously proposed.