MODULATION OF RECONSTITUTED ATP-SENSITIVE K-BINDING PROTEINS IN A MAMMALIAN-CELL LINE( CHANNELS BY GTP)

1. The action of GTP-binding proteins on ATP-sensitive potassium (K-AT P) channels was investigated. K-ATP channels were expressed in a mamma lian cell line (COS-1 cells) by cotransfecting vectors carrying the su lphonylurea receptor (SUR1) and BIR (Kir6.2), a member of the inward r ectifier K+ channel family. G proteins were also tested on K-ATP chann els composed of an isoform of SUR1, SUR2A, in combination with Kir6.2. 2. The alpha and beta gamma subunits of the GTP binding protein G(i) were tested separately in inside-out patches under continuous recordin g. G(alpha-i1) increases the activity of SUR.1-Kir6.2 and SUR2A-Kir6.2 channels by 200 and by 30%, respectively. 3. G(alpha-i2) does not inc rease the activity of SUR1-Kir6.2 channels, but increases the activity of SUR2A-Kir6.2 channels by 30%. 4. Control experiments showed that G TP gamma S, a specific activator of a proteins, and heat-inactivated G (alpha-i1) do not increase the single channel activity. 5. No effects of the other subunits (beta gamma) from either G(i1) or G(i2) on the s ingle channel activity were observed. 6. The protein kinase C inhibito rs H7 and an inhibitory peptide (FARKGALRQKNV)had no effect on the mod ulatory action of G(alpha-i1) on SUR1-Kir6.2 channels.7. We conclude t hat both types of reconstituted K-ATP channels are modulated by G prot eins.