Slow modal gating of single G protein-activated K+ channels expressed in Xenopus oocytes

摘要

class="enumerated" style="list-style-type:decimal">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 βγ subunits (Gβγ; 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 ∼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 Gβγ-activated channels cycled between a low-Po mode (Po of about 0.0034) and a bursting mode characterized by an ∼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 Gβγ.The GTPγS-activated Gαi1 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.