We found that roxatidine stimulates mucus secretion and synthesis by cultured rabbit gastric mucosal cells. In this study, we examined the roles of the extracellular Ca++ and calmodulin in these effects of roxatidine. Reduction of the extracellular Ca++ concentration decreased the roxatidine-induced increases in mucus secretion and synthesis by gastric mucosal cells. Roxatidine concentration-dependently promoted Ca++ influx and caused an increases in intracellular Ca++. After the addition of roxatidine, the increases in the secretion and synthesis reflected those in Ca++ influx and intracellular Ca++ concentration and then disappeared as Ca++ influx and intracellular Ca++ concentration returned to the control level. The roxatidine-stimulated Ca++ influx and intracellular Ca++ mobilization were abolished by reduction of the extracellular Ca++ concentration. Nifedipine and diltiazem inhibited both the effects of roxatidine, but even at 10 microM, the inhibition was partial. Furthermore, W-7 (a calmodulin antagonist) completely abolished the effects of roxatidine on mucus secretion and synthesis without causing a reduction of the stimulated Ca++ influx. Taken together, these results suggest that roxatidine promotes Ca++ influx through both voltage-sensitive Ca++ channels and other Ca++ entry gates and the subsequent intracellular Ca++ mobilization, leading to potentiation of mucus secretion and synthesis by rabbit gastric mucosal cells. In addition, Ca(++)-activated calmodulin may play a pivotal role in these stimulatory effects of roxatidine.