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The actions of somatostatin (SST) in the nervous system are mediated by specific high affinity SST receptors (SSTR1-5). However, the role of this hormone and the distribution of its receptor subtypes have not yet been defined in neural structures of the human fetus. We have analyzed four neural tissues (CNS, hypothalamus, pituitary and spinal cord) from early to midgestation for the expression of five human SSTR mRNAs, using a reverse transcription-polymerase chain reaction and Southern blot approach. These fetal neural tissues all express mRNA for multiple SSTR subtypes from as early as 16 weeks of fetal life but the developmental patterns of expression vary considerably. Transcripts for SSTR1 and SSTR2A are the most widely distributed, being expressed in all four neural tissues. SSTR2A is often the earliest transcript to be detected (7.5 weeks in CNS). SSTR3 mRNA is confined to the pituitary, hypothalamus, and spinal cord. SSTR4 is expressed in fetal brain, hypothalamus and spinal cord but not pituitary. SSTR5 mRNA is detectable in the pituitary and spinal cord by 14-16 weeks of fetal life. This mapping of SSTR mRNA expression patterns in human fetal neural tissues is an important first step toward our goal of determining the role of SST in the nervous system during early stages in human development.

Using explant cultures of human anterior pituitary glands (9-19 weeks fetal age) and an acute (3-h) test protocol, we investigated fetal somatotrope responsiveness to human GH-releasing factor [hGRF-(1-44)] and somatostatin [SRIF-(1-14) and SRIF-(1-28)] as a function of age. Ontogenic data were analyzed using three age groups: 9-10, 12-13, and 15-19 weeks fetal age. Both daily (24-h) and acute test (3-h) basal GH secretion increases as a function of fetal age, with the greatest increase occurring after 12-13 weeks; however, the 3 h/24 h secretion ratio remains unchanged, at approximately 12%. GRF (0.1-10 nM) stimulates GH release in a dose-related fashion, regardless of fetal age; there is a significant increase in the response to both 1 nM (P < 0.05) and 10 nM (P < 0.01) GRF between 9-10 and 15-19 weeks fetal age and to 10 nM GRF (P < 0.05) between 12-13 and 15-19 weeks. Pretreatment of cultures (9-19 weeks) with 1 or 10 nM GRF for 24 h does not alter basal 3-h GH secretion, but significantly decreases subsequent responses to 1 or 10 nM GRF, respectively (P < 0.01). Pretreatment with 1 nM GRF does not alter a subsequent 3-h response to 10 nM GRF. SRIF-(1-14) (1-100 nM) causes a dose-related inhibition of basal GH secretion from as early as the ninth week of fetal life; there is a small age-related increase in the somatotrope response to 100 nM SRIF-(1-14) between 12-13 and 15-19 weeks fetal age (P < 0.05). In a group of 11- to 14-week-old fetal pituitaries, SRIF-(1-28) had a significantly (P < 0.05) greater inhibitory effect than SRIF-(1-14) at both 1 and 10 nM; the two peptides decreased basal GH secretion to a similar extent at 100 nM (52.8 +/- 4.0% of control value; P < 0.01). SRIF-(1-14) (10 and 100 nM) does not significantly alter 10 nM GRF-stimulated GH release from 9- to 10-week-old fetal pituitaries. However, by 12-13 weeks, 10 nM SRIF-(1-14) reduces GRF-stimulated GH secretion by 60% (P < 0.01), while 100 nM SRIF-(1-14) decreases it by 80% (P < 0.01); similar inhibitory effects are observed with 15- to 19-week-old fetal somatotropes.(ABSTRACT TRUNCATED AT 400 WORDS)

Using explant cultures of human fetal anterior pituitary glands (9-19 weeks fetal age) and an acute (3-h) test protocol, we investigated the role of two signal transduction pathways (Gs-adenylate cyclase-Gi, 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 microM), 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 microM 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 microM), 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 microM forskolin, 1 mM theophylline or 1 mM dBcAMP is added with either 50 mM KCl or 1 microM Bay K 8644, there is an additive response. SRIF (10 nM) completely blocks the stimulatory effects of 1 microM Bay K 8644 and markedly inhibits the effects of 50 mM KCl from as early as the ninth week of fetal age. The Ca2+ channel blocker nifedipine (1-10 microM) significantly inhibits basal as well as stimulated (GRF, forskolin, dBcAMP, theophylline, KCl, and Bay K) GH release from as early as 9 weeks fetal age; in contrast, the calmodulin blocker trifluoperazine (0.01-10 microM) has no effects on basal GH secretion and only slightly inhibits the stimulatory effects of KCl. 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.(ABSTRACT TRUNCATED AT 400 WORDS)

Adrenals obtained from human abortices at midpregnancy were kept under conditions of tissue culture and the production of cortisol and dehydroepiandrosterone sulfate (3beta-hydroxy-5-androsten-17-one, 3-sulfate; DHA-S) monitored by radioimmunoassays for up to 2 weeks. Basal production of dehydroepiandrosterone sulfate was considerably higher than that of cortisol. alpha1-24corticotrophin, alpha1-39corticotrophin, (a long acting porcine corticotrophin) at the concentration of 1 mU/ml of culture medium in both instances, and dibutyryl cyclic AMP (1mM) enhanced the production of both steroids some 3 to 30 times above control values. Medium harvested from homologous pituitary cultures had comparable corticotrophic activity. It is concluded that at midpregnancy the regulation of corticoidogenesis implies the existence of a corticotrophic factor either identical or closely related to ACTH.

Using an explant culture system, we have demonstrated that human somatotropes respond to growth hormone releasing factor (GRF) and somatostatin (SRIF) from as early as 9.5 weeks of fetal age. Responsiveness to GRF increases significantly as a function of age up to midgestation while SRIF inhibition of basal growth hormone (GH) release does not change. SRIF has little effect on GRF-stimulated GH secretion from early gestation pituitaries, but its ability to block GRF stimulation gradually increases with fetal age from 9.5 to 16 weeks. The response to GRF remains predominant throughout this developmental period: 100 times more SRIF than GRF must be added to the cultures in order to block the GRF stimulatory effect and maintain GH secretion at basal (control) levels. Finally, adding SRIF 30 min prior to the GRF does not increase the inhibitory activity of SRIF. Our data suggest that the mechanisms that permit an interaction between GRF and SRIF are developing, but slowly, in the early to midgestation human fetal somatotrope and that GRF stimulatory pathways predominate. This may help to explain the very high levels of GH in fetal serum during the first half of gestation.

Explants prepared from the neocortex and the fetal zone of the human fetal adrenal (gestational age 13 to 18weeks) were maintained under conditions of organ culture for 7 to 9 days during which time they were exposed to hACTH and various related peptides. Corticotrophic activity was monitored by the daily release of dehydroepiandrosterone sulfate (3beta-hydroxy-5-androsten-17-one, 3-sulfate; DHA-S) and cortisol as quantified by radioimmunoassay, hACTH (2.2 x 10(-9) - 2.2 x 10(-8)M) was the most active in sustaining steroidogenesis by both neocortical and fetocortical cells. alpha-MSH possessed similar properties but not at concentrations lower than 10(-6)M, whereas CLIP (4.4 x 10(-9) - 1.1 x 10(-7)M), the 18-39 C-terminal moiety of ACTH, was devoid of activity. Corticotrophic activity with respect to fetocortical explants appeared to be that of maintenance of function best illustrated by dehydroepiandrosterone sulfate biosynthesis, while enhancement of steroidogenesis was observed in the neocortex as manifested by cortisol release. Although not eliminating the possible existence of a specific fetal corticotrophin related to ACTH1-39, the data indicate that hACTH is capable of regulating steroidogenesis in the fetal zone which is primarily geared to the formation of dehydroepiandrosterone sulfate.

We have examined the regulation of GH secretion from monolayer cultures of prepubertal male lamb anterior pituitary cells. Growth hormone-releasing factor (GRF 1-44) stimulated GH release in a dose-related manner: the maximal effective dose was 10(-10) M, which caused a 500% increase in basal GH secretion, while the half-maximal effect was reached with a dose of 2.5 x 10(-11) M (ED50). Thyrotropin-releasing hormone (TRH) also elicited a dose-dependent stimulation of GH secretion, although it was approximately 1000 times less potent than GRF. GRF and TRH did not have additive or synergistic effects on GH secretion. Somatostatin (SRIF) at a concentration of 10(-7) M maximally inhibited basal GH release to 40% of that of the control; the ED50 was 2.0 x 10(-9) M. Moreover, 10(-7) M SRIF blocked the stimulation of GH secretion induced by 10(-8) M GRF. However, when the cells were incubated with these two peptides at an identical concentration (10(-8) M), GH secretion was stimulated significantly above control values. When added at the same concentration (10(-7) M, TRH ans SRIF nullified their respective effects. A dose of 100 ng/ml of synthetic IGF-I was without effect on basal GH release, but significantly decreased 10(-9) M GRF-induced stimulation of GH secretion. these data indicate that in prepubertal male lambs: the stimulatory effect of GRF is predominant over the inhibitory effect of SRIF, somatostatin inhibits TRH stimulation of GH secretion in vitro, and IGF-I may control GH secretion by modulating GRF effects at the pituitary level.

To determine the direct effects of insulin-like growth factors (IGFs) on hormone release by the human pituitary gland, human foetal, adult normal and tumour pituitary tissues were maintained in culture for 2 to 4 weeks and tested with acute (3 h) exposures to different preparations of IGF peptides. Adult normal pituitaries and adenomas were tested with a semipurified preparation of IGFs, free of immunoreactive insulin, containing IGF-I and IGF-II in a ratio of approximately 1:4. Human foetal pituitaries were tested with the semipurified IGFs as well as more purified preparations of IGF-I and IGF-II. Culture media were assayed for hGH, hPrl, hACTH and hLH using specific radioimmunoassays. Both foetal (n = 16 (No. of pituitaries), 33 (No. of observations] and normal adult (n = 3, 16) human pituitaries cultures responded to the semipurified IGFs (2-25 ngEq/ml for foetal and 2-4 ngEq/ml for adult pituitaries) with a significant decrease in hGH release compared to basal (P less than 0.01) whereas the GH-secreting pituitary tumours showed no effect when tested with from 2 to 25 ngEq/ml (n = 8, 129, NS). The effect of IGFs on human foetal somatotrope activity was dose-related for both the semipurified IGFs (2-25 ngEq/ml, n = 16, 33) and IGF-I or IGF-II (10-100 ng/ml; n = 3, 18).(ABSTRACT TRUNCATED AT 250 WORDS)