Phospholipase C is involved in the insulinotropic effect of carbachol (CCh) and cholecystokinin octapeptide (CCK-8). The involvement of the type of G protein was investigated in rat pancreatic islets. Guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S; a nonhydrolyzable GTP analogue) increased insulin release in electrically permeabilized islets. Both CCh and CCK-8 increased the GTP gamma S effect indicative of an involvement of G proteins. Pretreatment of the islets with pertussis toxin (PT) impaired the CCh-induced insulin secretion in the presence of 3.0 mM glucose and inhibited the stimulatory CCh effect on inositol 1,4,5-trisphosphate (IP3) levels at low and high glucose. In contrast to CCh, the CCK-8 effect on both insulin release and IP3 levels of islets was not modified by a PT pretreatment at various glucose concentrations. Two types of experiments indicate the type of G protein involved: first, long-term agonistic stimulation by either CCh or CCK-8 led to a downregulation of alpha o and alpha q/11, respectively; second, introduction of specific anti-alpha o or -alpha q/11 antibodies into electrically permeabilized islets nearly completely abolished the effects of CCh and CCK-8, respectively. The data indicate that both CCh and CCK-8 act as insulinotropic agents via the phospholipase C system; in the effect of CCh the PT-sensitive alpha o and in the effect of CCK-8 the PT-insensitive alpha q/11 is involved.