Modulation of reconstituted ATP-sensitive K(+)-channels by GTP-binding proteins in a mammalian cell line.
1. The action of GTP-binding proteins on ATP-sensitive potassium (KATP) channels was investigated. KATP channels were expressed in a mammalian cell line (COS-1 cells) by cotransfecting vectors carrying the sulphonylurea receptor (SUR1) and BIR (Kir6.2), a member of the inward rectifier K+ channel family. G proteins were also tested on KATP channels composed of an isoform of SUR1, SUR2A, in combination with Kir6.2. 2. The alpha and beta gamma subunits of the GTP binding protein G1 were tested separately in inside-out patches under continuous recording. G alpha-11 increases the activity of SUR1-Kir6.2 and SUR2A-Kir6.2 channels by 200 and by 30%, respectively. 3. G alpha-12 does not increase the activity of SUR1-Kir6.2 channels, but increase the activity of SUR2A-Kir6.2 channels by 30%. 4. Control experiments showed that GTP gamma S, a specific activator of G proteins, and heat-inactivated G alpha-11 do not increase the single channel activity. 5. No effects of the other subunits (beta gamma) from either G11 or G12 on the single channel activity were observed. 6. The protein kinase C inhibitors H7 and an inhibitory peptide (FARKGALRQKNV) had no effect on the modulatory action of G alpha-11 on SUR1-Kir6.2 channels. 7. We conclude that both types of reconstituted KATP channels are modulated by G proteins.
Sánchez
J A
JA
Research Centre for Pathogenic Fungi and Microbial Toxicoses, Chiba University, Japan.
Gonoi
T
T
Inagaki
N
N
Katada
T
T
Seino
S
S
D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000494:pharmacology*; D000465:Algorithms; D000818:Animals; D002417:Cattle; D002460:Cell Line; D004594:Electrophysiology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000302:isolation & purification / Q000378:metabolism*; D008564:Membrane Potentials / Q000502:physiology; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000187:drug effects* / Q000235:genetics / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism; D051381:Rats; D014162:Transfection
D004791:Enzyme Inhibitors; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000255:Adenosine Triphosphate; D011493:Protein Kinase C; D019204:GTP-Binding Proteins