Antagonistic analogs of growth hormone-releasing hormone (GHRH) are being synthesized in our laboratory for various clinical applications, including treatment of certain endocrine disorders and insulin-like growth factor I-dependent tumors. To evaluate the endocrine effect of these GHRH antagonists, a sensitive dynamic in vitro system has been developed. The concentration causing 50% inhibition (IC50) of the standard GHRH antagonist human [N-Ac-Tyr1,D-Arg2]GHRH-(1-29)-NH2 is 4.5 x 10(-8) M in our dispersed pituitary cell superfusion system. This value is 11 times less than that measured in earlier static pituitary cell cultures. This reliable dynamic system is simple, fast, and inexpensive and not only makes it possible to obtain quantitative data on the inhibitory capacity of the antagonists but also provides information about the intrinsic GHRH activity of the analog. The dynamic interactions of the GHRH antagonist, the GHRH receptors, and GH release can also be evaluated by this superfusion system. The pulsatile GH release induced by 10(-9) M human GHRH-(1-29)-NH2 was inhibited by two modes of application, preincubation and simultaneous administration of the GHRH antagonist (10(-9) to 10(-6) M). The reduction in GHRH-stimulated GH response was more pronounced when the cells were preincubated with the antagonist prior to GHRH infusion than for simultaneous application. The inhibitory effect of the antagonist was dose-dependent, temporary, and of the competitive type. GH release induced by nonspecific stimulus (100 mM potassium chloride) was not influenced by the GHRH antagonist. This sensitive dynamic in vitro system appears to be a suitable method for screening the biological activity of various GHRH antagonists and eliminates the drawbacks of static pituitary cell culture.