Rapid desensitization of G protein-gated inwardly rectifying K(+) currents is determined by G protein cycle.
Activation of G protein-gated inwardly rectifying K(+) (GIRK) channels, found in the brain, heart, and endocrine tissue, leads to membrane hyperpolarization that generates neuronal inhibitory postsynaptic potentials, slows the heart rate, and inhibits hormone release. During stimulation of G(i/o)-coupled receptors and subsequent channel activation, it has been observed that the current desensitizes. In this study we examined mechanisms underlying fast desensitization of cloned heteromeric neuronal Kir3.1+3.2A and atrial Kir3.1+3.4 channels and also homomeric Kir3.0 currents in response to stimulation of several G(i/o) G protein-coupled receptors (GPCRs) expressed in HEK-293 cells (adenosine A(1), adrenergic alpha(2A), dopamine D(2S), M(4) muscarinic, and GABA(B1b/2) receptors). We found that all agonist-induced currents displayed a similar degree of desensitization except the adenosine A(1) receptor, which exhibits an additional desensitizing component. Using the nonhydrolyzable GTP analog guanosine 5'-O-(3-thiotriphosphate) (GTPgammaS), we found that this is due to a receptor-dependent, G protein-independent process. Using Ca(2+) imaging we showed that desensitization is unlikely to be accounted for solely by phospholipase C activation and phosphatidylinositol 4,5-bisphosphate (PIP(2)) hydrolysis. We examined the contribution of the G protein cycle and found the following. First, agonist concentration is strongly correlated with degree of desensitization. Second, competitive inhibition of GDP/GTP exchange by using nonhydrolyzable guanosine 5'-O-(2-thiodiphosphate) (GDPbetaS) has two effects, a slowing of channel activation and an attenuation of the fast desensitization phenomenon. Finally, using specific Galpha subunits we showed that ternary complexes with fast activation rates display more prominent desensitization than those with slower activation kinetics. Together our data suggest that fast desensitization of GIRK currents is accounted for by the fundamental properties of the G protein cycle.
Leaney
Joanne L
JL
Departmentt of Medicine, University College London, London WC1E 6JJ, UK.
Benians
Amy
A
Brown
Sean
S
Nobles
Muriel
M
Kelly
David
D
Tinker
Andrew
A
D002460:Cell Line; D004553:Electric Conductivity; D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D043802:GTP-Binding Protein alpha Subunits, Gq-G11 / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006801:Humans; D015640:Ion Channel Gating; D015221:Potassium Channels / Q000378:metabolism; D024661:Potassium Channels, Inwardly Rectifying / Q000378:metabolism* / Q000502:physiology; D020033:Protein Isoforms / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism
D051676:G Protein-Coupled Inwardly-Rectifying Potassium Channels; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D020033:Protein Isoforms; D011956:Receptors, Cell Surface; D019204:GTP-Binding Proteins; D043802:GTP-Binding Protein alpha Subunits, Gq-G11