1. Mn(2+) evoked an atropine-resistant secretion of amylase from the isolated pancreas of the young rat. The lowest effective concentration of Mn(2+) was 10(-3)m. The response to 10(-2)m-Mn(2+) was biphasic, an initial peak being followed by a slow sustained rise in amylase output. The maximal effect of 10(-2)m-Mn(2+) was to double the basal rate of amylase secretion after 70 min incubation.2. Co(2+) (10(-2)m) also stimulated amylase secretion. The maximal rate, about three times the basal value, was attained after 20 min incubation. Atropine partially inhibited this effect.3. Ca(2+) (10(-2)m) evoked an atropine-resistant amylase secretion similar in both magnitude and time course to the sustained phase observed with 10(-2)m-Mn(2+).4. Mn(2+) (10(-4)-10(-2)m) also increased the rate of (45)Ca efflux from the gland. Maximal efflux rates were attained after 30 min incubation and thereafter declined to basal values. A small increase was also observed with 10(-2)m-Co(2+), but not with 10(-2)m-Ca(2+). The effect of Co(2+) was almost completely abolished by atropine.5. Reducing the extracellular Ca(2+) concentration from 2.5 x 10(-3) to 10(-5)m did not reduce amylase secretion in response to 10(-2)m-Mn(2+), but secretion was abolished in a Ca(2+)-free medium containing EGTA. The increase in (45)Ca efflux rate evoked by Mn(2+) was inversely related to the extracellular Ca(2+) concentration.6. Mn(2+) (10(-2)m) increased the concentration of cyclic 3',5'-guanosine monophosphate (cyclic GMP) within the pancreas. Also, Mn(2+) accumulated within the cellular pool of the gland. The time course of both these effects was similar to the time course of (45)Ca efflux.7. Mn(2+) displaced Ca(2+) bound to isolated pancreatic microsomal membranes. The cation-binding sites on these membranes probably have a higher affinity for Mn(2+) than Ca(2+).8. We conclude that Mn(2+) stimulates enzyme secretion by displacing membrane-bound Ca(2+), the resulting increase in cytosolic Ca(2+) concentration activating the secretory mechanism.9. Mn(2+) partially inhibited amylase secretion stimulated by optimal doses of either acetylcholine (ACh) or caerulein. Maximal inhibition (about 60%) occurred with 10(-3)m-Mn(2+) (i.e. the lowest concentration required to stimulate secretion in the absence of secretagogues). Decreasing the extracellular Ca(2+) concentration reduced the inhibitory effect of Mn(2+).10. When glands were exposed to ACh and Mn(2+) simultaneously, the time required for inhibitory effects to develop was inversely related to the dose of ACh and the concentration of Mn(2+).11. Mn(2+) did not alter the acceleration of (45)Ca efflux evoked by ACh or by caerulein in a medium containing 2.5 x 10(-3)m-Ca(2+). However, under conditions of Ca(2+) deprivation ACh-stimulated (45)Ca efflux was greatly enhanced.12. Mn(2+) reduced the total amount of Ca(2+) accumulated into the cellular pool of the pancreas after 60 min incubation, but had no effect on the initial, rapid phase of Ca(2+) uptake.13. The effects of Mn(2+) on the relationship between ACh dose, amylase release and the extracellular Ca(2+) concentration suggest that the inhibitory actions of Mn(2+) cannot be explained by a simple, competitive interaction with the stimulant or with extracellular Ca(2+). However, the time course of inhibition is consistent with a requirement for Mn(2+) to accumulate within the acinar cells.14. Mn(2+) partially inhibited amylase secretion stimulated by hyperosmolarity and also increased the (45)Ca efflux rate under these conditions.15. Our results are not consistent with Mn(2+) exerting its inhibitory effect on secretagogue-stimulated enzyme secretion solely by blocking Ca(2+) influx from the extracellular space. We conclude that inhibition probably depends on the ability of Mn(2+) to displace Ca(2+) from binding sites involved in secretion, presumably coupled with a reduced ability of Mn(2+) to replace Ca(2+) in the secretory process.