We previously demonstrated that somatostatin (SRIF)-induced inhibition of GH secretion from human pituitary cells is mediated through both the SRIF receptor (SSTR) 2 and 5 subtypes. The interplay between these two SSTR subtypes in regulating GH was therefore tested in primary human fetal pituitary cultures (18–30 wk gestation). GHRH (10 nm)-stimulated GH secretion (51% increase, P < 0.05) was suppressed equally by either SSTR2 or SSTR5-selective agonists (10 nm). GH suppression correlated with agonist affinity for their respective receptor subtypes. Combined addition of SSTR2- and SSTR5-specific agonists was synergistic for GH suppression, achieving 73% (P < 0.05) inhibition as compared with inhibition attained with SSTR2 (32%) and SSTR5 (34%) agonists used alone (P < 0.05). The SSTR2 selective antagonist BIM-23454 dose-dependently blocked SSTR2 but not SSTR5-induced suppression of GH secretion. BIM-23454 also completely reversed GH suppression in response to the combined activation of SSTR2 and SSTR5. The IC₅₀ for BIM-23454 reversal of agonist-induced GH suppression was 55 nm and 33 nm for two SSTR2 agonists, 45 nm and 40 nm for the combination of SSTR2 and SSTR5 agonists, respectively, and 45 nm for the SSTR2/SSTR5 agonist BIM-23244, all of which were similar to the affinity of BIM-23454 for SSTR2 (32 nm). These results suggest the following: 1) activation of both SSTR2 and SSTR5 induces a functional association of receptor subtypes, resulting in synergistic GH suppression; 2) BIM-23454 is a potent SSTR2-selective antagonist capable of reversing SRIF-induced GH suppression; and 3) the ability of a selective SSTR2 antagonist to inhibit the GH suppressing action of SSTR2 agonist alone, SSTR2/SSTR5 biselective agonists, or SSTR2 and SSTR5 agonists in combination support the concept of a functional interaction between somatotroph SSTR2 and SSTR5 subtypes in primary human fetal pituitary cells.