CHARACTERIZATION OF THE G-PROTEIN COUPLING OF SRIF AND BETA-ADRENERGIC RECEPTORS TO THE MAXI K-CA CHANNEL IN INSULIN-SECRETING CELLS

Modulation of the Ca- and voltage-dependent K channel-K-Ca-by receptor s coupled to the G proteins G(i)/G(o) and G(s) has been studied in ins ulin-secreting cells using the patch clamp technique. In excised outsi de-out patches somatostatin (somatotropin-releasing inhibitory factor; SRIF) caused concentration-dependent inhibition of the K-Ca channel, an effect that was prevented by pertussis toxin (PTX), In inside-out p atches, exogenous alpha subunits of either G(i)- or G(o)-type G protei ns also inhibited the K-Ca channel (IC50 5.9 and 5.7 pM, respectively) . These data indicate that SRIF suppresses K-Ca channel activity via a membrane-delimited pathway that involves the alpha subunits of PTX-se nsitive G proteins G(i) and/or G(o). In outside-out patches, activatio n of G(s) either by beta-agonists or with cholera toxin (CTX) increase d K-Ca channel activity, consistent with a membrane-delimited stimulat ory pathway linking the beta-adrenergic receptor to the K-Ca channel v ia G(s). 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 exogeno us alpha(i) or alpha(o). Taken together these data suggest that K-Ca c hannel activity is enhanced by activation of G(2) and blocked by activ ated G(i) and/or G(o). Further, K-Ca channel stimulation by activated G(s) may be ''direct,'' while inhibition by G(i)/G(o) may involve deac tivation of G(s). In inside-out patches K-Ca channel activity was redu ced by an activator of protein kinase C (PKC) and enhanced by inhibito rs of PKC, indicating that PKC also acts to inhibit the K-Ca channel v ia 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 inhib itory effect of exogenous alpha(i) or alpha(o). These data indicate th at PKC facilitates the inhibitory effect of the PTX-sensitive G protei ns which are activated by coupling to SRIF receptors. To account for t hese results a mechanism is proposed whereby PKC may be involved in G( i)/G(o)-induced deactivation of G(s).