Slow modal gating of single G protein-activated K+ channels expressed in Xenopus oocytes.
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.
Yakubovich
D
D
Department of Physiology and Pharmacology, Sackler School of Medicine, Tel Aviv University, Ramat Aviv 69978, Israel.
Pastushenko
V
V
Bitler
A
A
Dessauer
C W
CW
Dascal
N
N
D000818:Animals; D003001:Cloning, Molecular; D005260:Female; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000378:metabolism; D015870:Gene Expression / Q000502:physiology; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology*; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009206:Myocardium / Q000737:chemistry; D009865:Oocytes / Q000502:physiology; D018408:Patch-Clamp Techniques; D010957:Plasmids; D015221:Potassium Channels / Q000235:genetics* / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying; D012333:RNA, Messenger / Q000494:pharmacology; D011976:Receptors, Muscarinic / Q000235:genetics / Q000378:metabolism; D014982:Xenopus laevis
D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D012333:RNA, Messenger; D011976:Receptors, Muscarinic; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D019206:GTP-Binding Protein alpha Subunits, Gi-Go