Uncoupling of Gi proteins with PTX prevents counteraction of GLP-1 potentiation of glucose-stimulated insulin secretion by clonidine. Cells pretreated with 100 ng/ml PTX for 18 h and control cells were subjected to analysis of static insulin secretion induced by 11 mM glucose for 30 min in the absence or presence of the subthreshold concentration and ED50 concentration of clonidine, the ED50 concentration of GLP-1, and their combinations. Insulin secretion was evaluated by a standard insulin radioimmunoassay. Both control and PTX-pretreated cells well responded to 30 min stimulation with 11 mM glucose (, versus corresponding 3 mM glucose-treated groups). PTX-pretreated cells released significantly more insulin than control cells following 11 mM glucose stimulation (, ). Neither the subthreshold concentration (3 nM) nor the ED50 concentration (4 μM) of clonidine altered glucose-stimulated insulin secretion in PTX-pretreated cells (, versus PTX-pretreated group subjected to only 11 mM glucose stimulation). In contrast, GLP-1 at the ED50 concentration of 0.1 nM induced a significant potentiation of glucose-induced insulin release from PTX-pretreated cells (, versus PTX-pretreated group subjected to only 11 mM glucose stimulation). Intriguingly, glucose-stimulated insulin secretion from PTX-pretreated cells incubated with the ED50 concentration of GLP-1 alone did not significantly differ from that from those subjected to incubation with the ED50 concentration of GLP-1 plus the subthreshold concentration of clonidine (, ). Clonidine at the subthreshold concentration could no longer influence potentiation of glucose-stimulated insulin secretion by the ED50 concentration of GLP-1 in PTX-pretreated cells. versus 3 mM glucose-treated group without PTX pretreatment, versus 11 mM glucose-treated group without PTX pretreatment or PTX-pretreated group subjected to 3 mM glucose incubation, versus PTX-pretreated group subjected to 11 mM glucose stimulation.
