Pepsinogen secretion (PS) is modulated at the intracellular level by both cAMP and calcium ion. Cholecystokinin octapeptide (CCK-8), a potent stimulus for PS, is believed to act through calcium. The most extensively studied pathway for calcium-mediated modulation involves the formation of calcium/calmodulin complexes, leading to activation of calmodulin. We have therefore examined the hypothesis that an inhibitor of calmodulin might inhibit PS stimulated by CCK-8. The phenothiazine derivative trifluoperazine (TFP) was chosen as a calmodulin antagonist. We measured in vitro secretion of pepsinogen by isolated gastric glands as a function of TFP concentration 10(-6) M-5 X 10(-4) M), in the presence and absence of a maximal concentration of CCK-8 (10(-7) M). Cellular viability was determined by measurement of release of the enzyme lactate dehydrogenase (LDH) into the medium. TFP did not significantly inhibit PS stimulation by CCK-8 at any concentration (P greater than 0.05). At 10(-4) M, TFP actually augmented PS stimulation by CCK-8 (P less than 0.05). TFP alone significantly stimulated PS (P less than 0.05) at 5 X 10(-5) M and above. TFP did not raise cAMP levels at any concentration tested (P less than 0.05), in contrast to the adenylate cyclase activator forskolin, 10(-5) M, which caused a 6- to 37-fold increase (P less than 0.05). TFP, 2 X 10(-4) did not increase LDH levels significantly (P less than 0.05). Thus a calmodulin inhibitor, TFP, paradoxically stimulates PS. This stimulatory effect of TFP is not cAMP-dependent and is not accompanied by a nonspecific release of LDH into the medium.(ABSTRACT TRUNCATED AT 250 WORDS)