Characterization of the G protein coupling of SRIF and beta-adrenergic receptors to the maxi KCa channel in insulin-secreting cells.
Modulation of the Ca- and voltage-dependent K channel--KCa--by receptors coupled to the G proteins G(i)/G(o) and Gs has been studied in insulin-secreting cells using the patch clamp technique. In excised outside-out patches somatostatin (somatotropin-releasing inhibitory factor; SRIF) caused concentration-dependent inhibition of the KCa channel, an effect that was prevented by pertussis toxin (PTX). In inside-out patches, exogenous alpha subunits of either G(i)- or G(o)-type G proteins also inhibited the KCa channel (IC50 5.9 and 5.7 pM, respectively). These data indicate that SRIF suppresses KCa channel activity via a membrane-delimited pathway that involves the alpha subunits of PTX-sensitive G proteins G(i) and/or G(o). In outside-out patches, activation of Gs either by beta-agonists or with cholera toxin (CTX) increased KCa channel activity, consistent with a membrane-delimited stimulatory pathway linking the beta-adrenergic receptor to the KCa channel via Gs. In outside-out patches, channel inhibition by SRIF suppressed the stimulatory effect of beta-agonists but not that of CTX, while in inside-out patches CTX reversed channel inhibition induced by exogenous alpha i or alpha o. Taken together these data suggest that KCa channel activity is enhanced by activation of Gs and blocked by activated G(i) and/or G(o). Further, KCa channel stimulation by activated Gs may be "direct," while inhibition by G(i)/G alpha may involve deactivation of Gs. In inside-out patches KCa channel activity was reduced by an activator of protein kinase C (PKC) and enhanced by inhibitors of PKC, indicating that PKC also acts to inhibit the KCa channel via a membrane delimited pathway. In outside-out patches, chelerythrine, a membrane permeant inhibitor of PKC prevented the inhibitory effect of SRIF, and in inside-out patches PKC inhibitors prevented the inhibitory effect of exogenous alpha i or alpha o. These data indicate that PKC facilitates the inhibitory effect of the PTX-sensitive G proteins which are activated by coupling to SRIF receptors. To account for these results a mechanism is proposed whereby PKC may be involved in G(i)/G(o)-induced deactivation of Gs.
Ribalet
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Department of Physiology, University of California Los Angeles 90024, USA.
Eddlestone
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D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D002772:Cholera Toxin / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism*; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D007545:Isoproterenol / Q000494:pharmacology; D009638:Norepinephrine / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D037342:Pertussis Toxin; D015221:Potassium Channels / Q000187:drug effects / Q000378:metabolism*; D011493:Protein Kinase C / Q000494:pharmacology; D011943:Receptors, Adrenergic, beta / Q000378:metabolism*; D013004:Somatostatin / Q000031:analogs & derivatives / Q000494:pharmacology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D007328:Insulin; D015221:Potassium Channels; D011943:Receptors, Adrenergic, beta; D010566:Virulence Factors, Bordetella; D013004:Somatostatin; D002772:Cholera Toxin; D037342:Pertussis Toxin; D011493:Protein Kinase C; D019204:GTP-Binding Proteins; D007545:Isoproterenol; D002118:Calcium; D009638:Norepinephrine