INVITRO MODULATION OF GROWTH-HORMONE (GH) SECRETION FROM EARLY TO MIDGESTATION HUMAN FETAL PITUITARIES BY GH-RELEASING FACTOR AND SOMATOSTATIN - ROLE OF G(S)-ADENYLATE CYCLASE-G(I) COMPLEX AND CA(2+) CHANNELS

Using explant cultures of human fetal anterior pituitary glands (9-19 weeks fetal ap) and an acute (3-h) test protocol, we investigated the role of two signal transduction pathways (G(s)-adenylate cyclase-G(i), Ca2+ channels) in GH-releasing factor (GRF)/somatostatin (SRIF) regulation of GH secretion during the first half of gestation. Data have been analyzed for ontogenic changes using three age groups: 9-10, 12-13, and 15-19 weeks fetal age. The fetal somatotrope shows dose-related responses to forskolin (0.1-10 mum), dibutyryl cAMP (dBcAMP; 0.1-1 mm), and theophylline (0.01-1 mm), all factors that increase intracellular concentrations of cAMP; there is a significant age-related increase in the stimulatory effects of 1 mm dBcAMP and 1 mm theophylline. When any one of these three factors is added with 10 nm GRF, there are no significant additive or synergistic effects on GH secretion. Although 10 nm SRIF has no effect at 9-10 weeks, it is inhibitory in the 12-13 and 15-19 week groups, significantly suppressing the stimulatory effect of 1 mum forskolin and completely blocking the effects of 1 mm dBcAMP or theophylline. Pretreatment of cultures with pertussis toxin completely blocks SRIF inhibition of both basal and GRF-stimulated GH release. KCl (5-50 mm) and Bay K 8644 (0.1-10 mum), both activators of Ca2+ channels, have dose-related stimulatory effects on GH release; 50 mm KCl shows an age-related increase in stimulatory activity. If 10 nM GRF, 1 mum forskolin, 1 mm theophylline or 1 mm dBcAMP is added with either 50 mm KCl or 1 mum Bay K 8644, there is an additive response. SRIF (10 nm) completely blocks the stimulatory effects of 1 mum Bay K 8644 and markedly inhibits the effects of 50 mm KCI from as early as the ninth week of fetal age. The Ca2+ channel blocker nifedipine (1-10 mum) significantly inhibits basal as well as stimulated (GRF, forskolin, dBcAMP, theophylline, KCI, and Bay K) GH release from as early as 9 weeks fetal age; in contrast, the calmodulin blocker trifluoperazine (0.01-10 mum) has no effects on basal GH secretion and only slightly inhibits the stimulatory effects of KCI. Pretreatment with 10 nm GRF for 24 h significantly decreases a subsequent 3-h response to 10 nm GRF, but does not alter the subsequent response to 1 mm theophylline or 50 mm KCl. These data suggest that between 9 and 19 weeks of fetal age: 1) there are age-related increases in the activity of cAMP-dependent (and, to a lesser extent, Ca2+-dependent) signal transduction pathways in the human somatotrope; 2) Ca2+ influx (but not calmodulin) plays a critical role in basal GH release; 3) GRF regulates fetal somatotrope activity through Ca2+- as well as cAMP-dependent mechanisms; 4) SRIF acts through a pertussis toxin-sensitive G-protein to inhibit basal as well as GRF-stimulated GH secretion; 5) the sites at which SRIF influences the fetal somatotrope are linked first to Ca2+ influx and, after 10 weeks of fetal life, to cAMP generation as well; and 6) GRF desensitization may be due to a change at the GRF receptor level.