Abstract
The slow kinetics of G protein-activated K+ (GIRK) channels expressed in Xenopus oocytes were studied in single-channel, inside-out membrane patches. Channels formed by GIRK1 plus GIRK4 subunits, which are known to form the cardiac acetylcholine (ACh)-activated GIRK channel (KACh), were activated by a near-saturating dose of G protein betagamma subunits (Gbetagamma; 20 nM). The kinetic parameters of the expressed GIRK1/4 channels were similar to those of cardiac KACh. GIRK1/4 channels differed significantly from channels formed by GIRK1 with the endogenous oocyte subunit GIRK5 (GIRK1/5) in some of their kinetic parameters and in a 3-fold lower open probability, Po. The unexpectedly low Po (0.025) of GIRK1/4 was due to the presence of closures of hundreds of milliseconds; the channel spent approximately 90 % of the time in the long closed states. GIRK1/4 channels displayed a clear modal behaviour: on a time scale of tens of seconds, the Gbetagamma-activated channels cycled between a low-Po mode (Po of about 0.0034) and a bursting mode characterized by an approximately 30-fold higher Po and a different set of kinetic constants (and, therefore, a different set of channel conformations). The available evidence indicates that the slow modal transitions are not driven by binding and unbinding of Gbetagamma. The GTPgammaS-activated Galphai1 subunit, previously shown to inhibit GIRK channels, substantially increased the time spent in closed states and apparently shifted the channel to a mode similar, but not identical, to the low-Po mode. This is the first demonstration of slow modal transitions in GIRK channels. The detailed description of the slow gating kinetics of GIRK1/4 may help in future analysis of mechanisms of GIRK gating.