The interrelationship between the effects of insulin-like growth factor I and somatostatin on growth hormone secretion by normal rat pituitary cells: the role of glucocorticoids

Both insulin-like growth factor I (IGF-I) and somatostatin (SRIH) have been shown to directly inhibit GH release and the total GH content of cultured pituitary cells. In the present study we evaluated the interrelationship between the effects of a recombinant human IGF-I analog ([Thr59]IGF-I) and SRIH on GH release by cultured normal rat pituitary cells together with the effects of glucocorticoids. In all experiments anterior pituitary cells were preincubated for 24 h without or with IGF-I, SRIH, and/or dexamethasone. Thereafter, 24-h incubations without or with IGF-I, dexamethasone, SRIH, and GHRH were performed. Both IGF-I and SRIH inhibited basal and GHRH-stimulated GH release in a dose-dependent manner; the maximal inhibitory concentrations were 5 nM IGF-I and 10 nM SRIH. These concentrations inhibited basal and GHRH-stimulated GH release by 23% and 40% (IGF-I) and 80% and 85% (SRIH), respectively. The combination of IGF-I and low concentrations of SRIH exerted an additive inhibitory effect on GHRH-stimulated GH release; IGF-I (1 nM) and SRIH (10 pM) together inhibited GH release by 50%, while the maximal inhibitory concentrations of 5 nM IGF-I and 10 nM SRIH virtually completely inhibited GH release (by 93%). Preincubation with 5 and 100 nM dexamethasone attenuated the sensitivity of somatotrophs to SRIH and completely abolished the inhibitory effects of IGF-I. This effect of dexamethasone could be reversed by coincubation with the glucocorticoid receptor antagonist RU 38486. High concentrations of 5-10 nM of the recombinant human IGF-I analog stimulated PRL cell content (5 and 10 nM) and release (10 nM), while a purified IGF-I preparation extracted from human blood exerted a parallel inhibitory effect on GH and PRL release. We conclude that 1) IGF-I and SRIH exert an additive direct inhibitory effect on basal and GHRH-stimulated GH secretion by normal cultured pituitary cells; 2) glucocorticoids directly attenuate the sensitivity of somatotrophs to SRIH, but completely prevent the inhibitory effects of IGF-I on GH secretion; and 3) in contrast to a purified IGF-I preparation extracted from human blood (which inhibits GH and PRL release) high concentrations of the recombinant IGF-I preparation (which inhibit GH release) stimulate PRL production.     

The effect of age on somatostatin suppression of basal, growth hormone (GH)-releasing factor-stimulated, and dibutyryl adenosine 3',5'-monophosphate-stimulated GH release from rat pituitary cells in monolayer culture

To test the hypothesis that relative resistance of the somatotroph to somatostatin (SRIF) contributes to elevated circulating levels of GH in the newborn rat, we examined the effects of SRIF (0.1, 0.33, and 1 nM) on basal, human pancreatic GH-releasing factor-40 (hpGRF-40; 1 nM)-stimulated, and (Bu)2cAMP (0.5 mM)-stimulated GH release from pituitary cells of 2-day-old, 15-day-old, and adult Sprague-Dawley rats in monolayer culture. The effect of SRIF on basal GH release varied markedly with age. SRIF, in the doses studied, did not inhibit basal GH release (nanograms of GH per 10(5) cells/3 h) from pituitary cultures of 2-day-old rats. In those of 15-day-old rats, only the two higher doses of SRIF (0.33 and 1 nM) suppressed GH release. By contrast, in pituitary cell cultures of adult male and female rats, all doses of SRIF significantly inhibited basal GH release (P less than 0.001). Similarly, the degree of SRIF suppression of both hpGRF-40- and (Bu)2cAMP-stimulated GH release differed among the age groups. In pituitary cultures of 2-day-old rats, SRIF did not significantly inhibit stimulated GH release. In 15-day-old rat pituitary cells, SRIF inhibited GH release, but did not eradicate the stimulatory effect of hpGRF-40 or (Bu)2cAMP. By contrast, in pituitary cell cultures of adult male and female rats, SRIF completely abolished the stimulatory effect of both hpGRF-40 and (Bu)2cAMP. When expressed as a percentage of the control (or stimulated) value, GH release at each SRIF dose varied markedly with age (P less than 0.001). Furthermore, a similar age-associated trend was evident when, in a separate series of experiments (n = 37), we examined the suppressive effect of a single concentration of SRIF (0.33 nM) on (Bu)2cAMP (0.5 mM)-stimulated GH release in cultured pituitary cells of rats ranging in age from -1 (day 20 of gestation) to 78 days. The degree of suppression increased progressively with advancing age; GH release decreased from 82 +/- 2% (+/- SE) of stimulated values in cultured cells of perinatal rats to 20 +/- 1% of stimulated values in cultured cells of 78-day-old rats. There was a significant negative correlation between age and SRIF-inhibited GH release (r = -0.89; P less than 0.001).(ABSTRACT TRUNCATED AT 400 WORDS)     

The interrelationship between the effects of somatostatin and human pancreatic growth hormone-releasing factor on growth hormone release by cultured pituitary tumor cells from patients with acromegaly

The effects of somatostatin (SRIF) and human pancreatic tumor GRF on GH release by cultured pituitary tumor cells obtained during transsphenoidal operation from 15 acromegalic patients were investigated. In a study of the sensitivity of pathological GH release to SRIF, 1-10 nM SRIF induced maximal inhibition of hormone release in 3 consecutive tumors. In 12 of 15 tumor cell cultures, 10 nM SRIF produced statistically significant inhibition of basal GH release by 39 +/- 3% (mean +/- SEM). In 2 of the 3 other tumors, SRIF inhibited GRF-stimulated GH release, while this was not investigated in the third tumor. A dose-response study of the effect of GRF on GH release by cultured pituitary tumor cells showed that doses of 0.1, 1, 10, and 100 nM induced similar maximal (35%) stimulation of hormone secretion. In four of five consecutive tumor cell suspensions, 1 and 10 nM GRF induced statistically significant GH stimulation by 18-300%. Preincubation of the tumor cells with 5 nM dexamethasone greatly increased the sensitivity and the maximal stimulation in response to GRF and made one tumor cell suspension, which did not react to GRF initially, sensitive to GRF. In the tumors of four patients, the interrelationship between the effects of SRIF and GRF on GH release were also studied. SRIF (10 nM) inhibited the stimulatory effects of GRF on GH release virtually completely. In conclusion, GH release by in vitro cell cultures of GH-secreting pituitary adenomas was inhibited by SRIF and stimulated by GRF. The interaction of GRF and SRIF on GH release by these pituitary tumor cells was similar to that in normal rat GH cells, as SRIF virtually completely overcame the GRF-induced GH release.     

The growth hormone (GH)-releasing hormone (GHRH)-GH-somatomedin axis: evidence for rapid inhibition of GHRH-elicited GH release by insulin-like growth factors I and II

Hypothalamic-pituitary-end-organ axes are frequently controlled by long loop negative feedback homeostatic mechanisms. Insulin-like growth factor I (IGF-I), IGF-II, and insulin receptors have recently been described in normal and neoplastic rat and acromegalic human pituitary cells, a finding which suggests the possibility that somatomedins might exert feedback at the level of the anterior pituitary. To study the kinetics of this feedback response, we used perifused dispersed rat anterior pituitary cells to learn if somatomedins or insulin could inhibit GH-releasing hormone (GHRH)-stimulated GH secretion. Cells were exposed to hourly boluses of 1 nM GHRH with or without varying doses of IGF or insulin. IGF-I inhibited GHRH-elicited GH release with an IC50 of 6.5 nM; maximal inhibition (approximately 67%) was achieved with 10 nM IGF-I. IGF-II was a less potent hormone, with 10 nM inhibiting about 30% of GHRH-stimulated GH release. Slight inhibition of stimulated GH release (less than 15%) was seen when cells were treated with insulin, but only when doses of insulin of 10 nM or more were used. In conclusion, nanomolar concentrations of IGF-I and IGF-II inhibited GHRH-elicited GH release from perifused rat pituitary cells in a dose-dependent manner; and insulin was not an effective inhibitor of stimulated GH release at physiological peptide concentrations. In conjunction with our previous findings that the concentrations of IGF-I and IGF-II receptors greatly exceed that of insulin receptors on normal rat pituitary cells, we hypothesize that the GH-inhibiting action of high dose insulin is mediated through an IGF receptor.     

Bombesin stimulates prolactin and growth hormone release by pituitary cells in culture

Bombesin (BBS) has been previously shown to stimulate the secretion of PRL and GH in steroid-primed rats. To determine whether these effects were mediated by the central nervous system or were due to direct action on the pituitary gland, we studied the interaction of BBS with GH4C1 cells, a clonal strain of rat pituitary cells which synthesizes and secretes PRL and GH. The addition of 100 nM BBS to GH4C1 cells for 60 min increased PRL release to 140 +/- 3% of the control value (mean +/- SE) and GH release to 133 +/- 5% of the control value. Stimulation of hormone secretion was observed within 15 min of treatment with 100 nM BBS and continued for at least 2 h. Half-maximal stimulation of PRL release occurred with 0.5 nM BBS, and a maximal effect was observed with 10 nM peptide. The BBS analogs ranatensin, litorin, and [Tyr4]BBS, each at a concentration of 100 nM, caused the same stimulation of PRL release as maximal concentrations of BBS itself. BBS stimulated hormone release selectively in two of five different clonal pituitary cell strains examined. Pretreatment of GH4C1 cells with 1 nM estradiol and/or 100 nM insulin resulted in more powerful stimulation of PRL release by both TRH and BBS. When epidermal growth factor and vasoactive intestinal peptide were added simultaneously with BBS, PRL release was greater than in the presence of either peptide alone. In contrast, the stimulatory effects of TRH and BBS were not additive. Somatostatin inhibited both basal and stimulated PRL release. Thus, low concentrations of BBS can directly stimulate PRL and GH release by a clonal pituitary cell strain in culture. These results suggest that BBS may stimulate PRL and GH secretion in vivo by direct action on the pituitary gland.     

In vitro regulation of growth hormone (GH) release from ovine pituitary cells during fetal and neonatal development: effects of GH-releasing factor, somatostatin, and insulin-like growth factor I

Using a monolayer approach, we have examined the acute (3 h) effects of GRF, somatostatin (SRIF), and insulin-like growth factor I (IGF-I) on GH release from pituitary cells of male and female 70-, 100-, and 130-day-old fetuses and newborn lambs and of prepubertal male lambs. GRF stimulated basal GH release in a dose-dependent (10(-12)-10(-8) M) manner at each stage in development. There was no linear relationship between maximal response and increasing age of the donor animals. The ED50 values for GRF were similar in all groups, except in the pituitaries from male and female 130-day-old fetuses, where the ED50 values were significantly higher. SRIF elicited a dose-related (10(-10)-10(-6) M) inhibition of basal GH secretion at each stage of fetal life and in the prepubertal period; although the response was lower in the youngest fetal pituitaries, there was no significant change in maximal response during the fetal or prepubertal period. No effect of SRIF on basal GH secretion was observed in newborn lambs. However, SRIF (10(-7) M) was able to block GRF (10(-8) M)-stimulated GH release in 100- and 130-day-old fetal and prepubertal as well as newborn lamb pituitary cells. Plasma IGF-I concentrations increased from 15.0 +/- 0.7 (mean +/- SE) and 13.8 +/- 0.9 ng/ml for male and female animals, respectively, at 70 days gestation to 55.8 +/- 3.2 and 51.8 +/- 11.1 ng/ml at the time of birth. The increase was much more pronounced in prepubertal lambs, especially in male animals, where IGF-I levels reached 300.8 +/- 37.7 ng/ml. IGF-I (100 ng/ml) had no effect on basal GH release in 70- and 100-day-old fetal, newborn, and prepubertal lamb pituitary cultures, but significantly inhibited basal GH secretion from 130-day-old fetal cells. This dose of IGF-I had no effect on GRF (10(-9) M)-stimulated GH release at 70 days gestation. It significantly inhibited this effect at 100 days and in prepubertal lamb cells. In 130-day-old fetal and newborn lamb pituitary cultures, IGF-I completely blocked the GH response to GRF.(ABSTRACT TRUNCATED AT 400 WORDS)     

Stimulation of growth hormone release by vasoactive intestinal peptide and peptide PHI in rat anterior pituitary reaggregates. Permissive action of a glucocorticoid and inhibition by thyrotropin-releasing hormone

In superfused rat anterior pituitary cell reaggregates, cultured for 5 days in serum-free defined medium, vasoactive intestinal peptide (VIP) concentration-dependently stimulated prolactin (Prl) release but had only a marginal effect on growth hormone (GH) release. When reaggregates were cultured in the presence of 80 nM dexamethasone (Dex) VIP strongly stimulated GH release from a concentration as low as 0.1 nM. VIP did not stimulate LH release. Peptide PHI also stimulated GH release but thyrotropin-releasing hormone (TRH) or angiotensin II did not. In fact, TRH slightly but transiently inhibited basal GH release and strongly inhibited VIP-stimulated GH release. GH-releasing factor (GRF) stimulated GH more potently and with higher intrinsic activity than VIP but GRF did not increase Prl release. The present data indicate that under defined hormonal conditions VIP and PHI are capable of stimulating GH release and that TRH can antagonize this effect by a direct action on the pituitary.     

Regulation of growth hormone release from cultured human pituitary adenomas by somatomedins and insulin

GH secretion is stimulated by hypothalamic GH-releasing factor (GHRH) and inhibited by somatostatin. Since GH induces the production of insulin-like growth factors (IGF) in liver and other tissues, it is of interest to learn whether IGF alters GH release through long loop feedback inhibition. Pituitary adenomas which had been removed from six acromegalic patients were processed for dispersed cell cultures and/or cell membrane preparations. Binding studies using 125I-labeled IGF-I, IGF-II, and insulin revealed specific hormone binding for each ligand to cell membranes derived from four somatotropinomas. A partially purified somatomedin preparation inhibited basal and/or GHRH-stimulated GH release from cultured pituitary cells derived from three of four adenomas; there was no effect of somatomedin in one tumor. In a single tumor, insulin also partially inhibited GHRH-stimulated GH release. Additionally, in one nonadenomatous pituitary removed from a patient with diabetes mellitus, insulin and somatomedin inhibited GHRH-stimulated GH release, and insulin inhibited basal GH secretion. These results indicate that specific cell membrane receptors for somatomedin peptides and insulin may be found on cell membranes from GH-secreting tumors, and that somatomedins and insulin can inhibit GH release in cultured human somatotropinoma cells. Thus, these data suggest that somatomedins may exert feedback inhibition of GH secretion in some patients with acromegaly.     

The effects of thyroid hormone deprivation in vivo and in vitro on growth hormone (GH) responses to human pancreatic (tumor) GH-releasing factor (1-40) by dispersed rat anterior pituitary cells

Anterior pituitary cells from euthyroid and hypothyroid male rats have been cultured as monolayers for 3 days with or without 5 nM T3 and stimulated with either human pancreatic GH-releasing factor 1-40 (hpGRF), TRH, or the Ca2+ channel ionophore A23187. Basal GH secretion was reduced in the hypothyroid cultures (P less than 0.001) and basal TSH secretion increased (P less than 0.001). Culture with T3 increased GH secretion and intracellular GH content in euthyroid and hypothyroid cultures but suppressed TSH secretion with no effect on intracellular TSH content in either euthyroid or hypothyroid cultures. hpGRF released more GH from euthyroid [3.52 +/- 0.2 (SE) micrograms/6 h X 10(5) cells] than hypothyroid cultures of (0.17 +/- 0.01 micrograms/6 h X 10(5) cells, P less than 0.001) without a change in ED50 (approximately 0.02 nM). The reduction in hpGRF-induced GH release remained significant when corrected for the reduced intracellular GH content in the hypothyroid cultures. hpGRF-induced GH release also declined relative to A23187-induced GH release in hypothyroid cultures. Culture with 5 nM T3 doubled maximum hpGRF-induced GH release in euthyroid cultures and increased maximum release 10-fold in hypothyroid cultures without altering the ED50 of hpGRF action. In contrast, T3 suppressed TRH-induced TSH release in euthyroid cultures but was without effect on TRH-induced TSH release in the hypothyroid cultures. T3 did not effect the ED50 of TRH action (2-5 nM). In summary, hypothyroid rat anterior pituitary cells in culture have a reduced maximal GH response to hpGRF, but the same ED50. hpGRF activity can be partially restored by physiological concentrations of T3 in vitro.     

Growth hormone secretagogue (GHS) analogue, hexarelin stimulates GH from peripheral lymphocytes.

The role of growth hormone releasing hormone (GHRH) and growth hormone releasing peptide-6 (GHRP-6) analogue hexarelin was investigated in the regulation of GH production from lymphocytes. Porcine and bovine blood mononuclear cells were separated using density gradient centrifugation method by layering the whole blood or buffy coat cells on lymphodex. Cells were incubated for 3 or 5 days with or without phytohemagglutinin (PHA-M), GHRH, GHRP-6 analogue hexarelin, somatostatin or GHRH + hexarelin. Growth hormone was fractionated from supernatants by gel chromatography and further concentrated by lyophilization at - 20 degrees C. A nearly two fold increase in basal secretion of GH (porcine: 3.5 +/- 0.1 ng/ml, bovine: 3.2 +/- 0.2 ng/ml) was achieved by GHRH and hexarelin at concentrations of 0.1, 1.0, 10 and 100 nM in both porcine and bovine cells. Lymphocytic GH release was also stimulated in response to PHA-M (10 micro g/well). Neither a dose dependent nor a synergistic nor an additive effect was apparent on GH secretion from lymphocytes. GHRH stimulated lymphocytic GH secretion, whereas, somatostatin had no effect. This study reports for the first time that hexarelin stimulates the secretion of GH from peripheral lymphocytes.     

Insulin-like growth factor I regulates growth hormone secretion and messenger ribonucleic acid levels in human pituitary tumor cells

GH secretion and mRNA levels were measured in cultured human GH adenoma cells incubated in serum-free medium for up to 48 h. A human recombinant insulin-like growth factor I (IGF-I) analog, Thr-59-IGF-I (6.5 nM), inhibited basal GH secretion by up to 60% in tumor cell cultures. The 30-50% stimulation of GH secretion by GH-releasing hormone (GHRH) was prevented by simultaneous exposure of the cells to IGF-I (6.5 nM). Gel electrophoresis of total RNA derived from GH cell adenoma tissue, followed by transfer and hybridization with 32P-labeled human GH cDNA, revealed a distinct mRNA species of about 1.0 kilobases. Using cytoplasmic dot blot hybridization, IGF-I inhibited the levels of human GH mRNA sequences in these cells and also prevented the GHRH-induced stimulation of GH mRNA. A monoclonal antibody to the type I IGF-I receptor (alpha IR3) prevented the inhibitory effects of IGF-I on basal and GHRH-stimulated GH secretion. This antibody also prevented the IGF-I-induced suppression of GH mRNA sequences. PRL secretion in these cells was not altered by IGF-I. Furthermore, relative levels of beta-actin mRNA were unaltered by IGF-I. Thus, IGF-I suppresses basal and GHRH-stimulated GH secretion and GH mRNA levels in pituitary adenoma cells, indicating that IGF-I acts selectively on the somatotroph to directly regulate GH gene expression.     

Effects of insulin-like growth factors on adult male rat pituitary function in tissue culture

To determine the direct effects of insulin-like growth factors (IGFs) on pituitary secretion of GH, PRL, and ACTH, adult male rat pituitary explant cultures were tested with acute (3-4 h) or chronic (24 h) exposure to a semipurified preparation of IGF peptides, free of immunoreactive insulin, containing IGF-I and IGF-II in a ratio of approximately 1:4. To examine the effect of serum binding proteins on IGF bioactivity, certain experiments were run in parallel using culture medium supplemented with 10% fetal bovine serum or 1% purified BSA. To compare IGF effects with those of known regulators of pituitary function, cultures were also tested with SRIF, TRH, human pancreatic GH-releasing factor, insulin, and human GH (hGH). IGFs, at 10-100 ngeq/ml, were able to inhibit significantly both basal and (1 mM) theophylline-stimulated rat GH (rGH) and rat PRL (rPRL) release during acute (3-4 h) exposure. Only the higher concentration (100 ngeq/ml) was consistently effective in inhibiting rGH and rPRL output after 24 h in culture, due to gradual metabolism of IGF peptides by the cells. Parallel experiments carried out in medium containing 10% fetal bovine serum or 1% BSA gave similar results, demonstrating that IGF serum binding proteins did not interfere with IGF bioactivity in this test system. Chronic 5-day exposure to IGFs, at 100 ngeq/ml, resulted in a significant inhibition of rGH release for the entire 5-day period and rPRL release for the first 3 days. IGFs (10-100 ngeq/ml) had no acute or chronic effect on basal or theophylline-stimulated ACTH release. Purified IGF-I (50 ng/ml) and IGF-II (50 ng/ml) gave approximately equivalent effects on basal rGH and rPRL release during an acute (3 h) exposure suggesting that both IGFs can exert inhibitory influence on pituitary function. Ten thousand nanograms per ml insulin and 10(-9) M SRIF had acute inhibitory effects on rGH and rPRL release similar to what were observed for 100 ngeq/ml semipurified IGFs. hGH (200 and 1000 ng/ml) had no effect on rGH, rPRL, or ACTH release when administered either acutely (3-4 h) or chronically (24 h). Conclusions: These studies demonstrate that IGFs, administered acutely or chronically, directly inhibit basal as well as theophylline-stimulated rGH and rPRL output by the rat pituitary; ACTH release remains unaltered. Insulin, at high concentrations, can mimic these effects, whereas hGH has no effect either acutely or chronically.     

Calcitonin inhibits basal and thyrotropin-releasing hormone-induced release of prolactin from anterior pituitary cells: evidence for a selective action exerted proximal to secretagogue-induced increases in cytosolic Ca2+

Salmon calcitonin (sCT)-like peptide is present in the central nervous system and pituitary gland of the rat, and this peptide inhibits basal and TRH-stimulated PRL release from cultured rat anterior pituitary (AP) cells. The present studies were designed to examine further the inhibitory actions of sCT on basal and TRH-stimulated PRL release and investigated 1) the temporal dynamics of the responses, 2) the effects of sCT on PRL release induced by other secretogogues, and particularly those acting via elevations of cytosolic Ca2+, and 3) the selectivity of sCT action on basal and stimulated AP hormone release. The inhibition of basal PRL release by sCT (0.1-10 nM) was dose-dependent and was characterized by a rapid onset with a gradual recovery to normal rates of release after the period of sCT inhibition. The inhibitory effect of sCT on basal PRL release was reversed by treatment with either the Ca2+ ionophore A23187 or with the phorbol ester, phorbol myristate acetate (PMA). sCT infusion did not affect the basal release of GH, TSH, FSH, or LH by perifused AP cells. When administered in short pulses, TRH, at concentrations from 1-100 nM, elicited a dose-dependent increase in PRL release. When coadministered with short 10 nM TRH, sCT (1-100 nM) inhibited TRH-induced PRL release in a dose-dependent manner, with a maximal inhibition of 78% at a concentration of 10 nM, and an ED50 concentration of approximately 3 nM. During longer (30 min) pulses of TRH (100 nM), PRL release increased sharply over 4-fold within 2 min, followed within 12 min by a rapid decline to a level 1.5-2-fold higher than basal, and this level was maintained for the remainder of the stimulation period. sCT pretreatment inhibited the overall PRL response to TRH. In contrast to its inhibition of TRH-induced PRL release, sCT failed to prevent the stimulation of PRL release by either ionophore A23187, PMA, vasoactive intestinal peptide, or forskolin. In addition, sCT failed to block TRH-induced TSH release or GnRH-induced LH release.(ABSTRACT TRUNCATED AT 400 WORDS)     

Effects of interleukin-1 on hormone release from normal rat pituitary cells in primary culture

The present study was performed mainly to determine whether interleukin-1 (IL-1), a polypeptide produced by immunologically activated monocytes, plays a physiological role in the regulation of adrenocorticotropic hormone (ACTH) using primary monolayer cultures of rat anterior pituitary cells. Neither human IL-1 alpha nor IL-1 beta stimulated the ACTH release from normal pituitary cells in concentrations ranging from 0.01 to 10 nM. IL-1 beta caused a slight, but significant, increase in ACTH release at a concentration of 100 nM, while IL-1 alpha did not, even at the highest dose tested. IL-1 beta exhibited a synergistic action with corticotropin-releasing factor (CRF) in ACTH secretion at 10 and 100 nM of CRF, but the interaction was not striking. Both of the monokines failed to cause any change in the secretions of growth hormone, prolactin, follicle-stimulating hormone and luteinizing hormone throughout concentrations ranging from 0.01 to 100 nM. The effects of possible sex-related differences and prolonged preincubation of cultured pituitary cells in serum-free medium prior to assay incubation were also tested, providing no significantly different findings. These results suggest that the physiological significance of IL-1 as a tissue CRF is indeed questionable and should be further clarified.     

Thyrotropin-releasing hormone stimulates growth hormone release from the anterior pituitary of hypothyroid rats in vitro

We have examined the interaction of thyroid hormone and TRH on GH release from rat pituitary monolayer cultures and perifused rat pituitary fragments. TRH (10(-9) and 10(-8)M) consistently stimulated the release of TSH and PRL, but not GH, in pituitary cell cultures of euthyroid male rats. Basal and TRH-stimulated TSH secretion were significantly increased in cells from thyroidectomized rats cultured in medium supplemented with hypothyroid serum, and a dose-related stimulation of GH release by 10(-9)-10(-8) M TRH was observed. The minimum duration of hypothyroidism required to demonstrate the onset of this GH stimulatory effect of TRH was 4 weeks, a period significantly longer than that required to cause intracellular GH depletion, decreased basal secretion of GH, elevated serum TSH, or increased basal secretion of TSH by cultured cells. In vivo T4 replacement of hypothyroid rats (20 micrograms/kg, ip, daily for 4 days) restored serum TSH, intracellular GH, and basal secretion of GH and TSH to normal levels, but suppressed only slightly the stimulatory effect of TRH on GH release. The GH response to TRH was maintained for up to 10 days of T4 replacement. In vitro addition of T3 (10(-6) M) during the 4-day primary culture period significantly stimulated basal GH release, but did not affect the GH response to TRH. A GH stimulatory effect of TRH was also demonstrated in cultured adenohypophyseal cells from rats rendered hypothyroid by oral administration of methimazole for 6 weeks. TRH stimulated GH secretion in perifused [3H]leucine-prelabeled anterior pituitary fragments from euthyroid rats. A 15-min pulse of 10(-8) M TRH stimulated the release of both immunoprecipitable [3H]rat GH and [3H]rat PRL. The GH release response was markedly enhanced in pituitary fragments from hypothyroid rats, and this enhanced response was significantly suppressed by T4 replacement for 4 days. The PRL response to TRH was enhanced to a lesser extent by thyroidectomy and was not affected by T4 replacement. These data suggest the existence of TRH receptors on somatotrophs which are suppressed by normal amounts of thyroid hormones and may provide an explanation for the TRH-stimulated GH secretion observed clinically in primary hypothyroidism.     

Platelet-activating factor stimulates prolactin release from dispersed rat anterior pituitary cells in vitro

The biologically active phospholipid (platelet-activating factor (PAF; 1-O-alkyl-2-acetyl-sn-glycero-3-phosphocholine) stimulated PRL release from dispersed rat anterior pituitary cells in culture. PAF-induced PRL release was dose dependent, with threshold stimulation at 1 nM and maximal stimulation at 100 nM. Stimulation occurred as early as 1 min of incubation and persisted for 2 h. The action of PAF on PRL release is consistent with a receptor-mediated mechanism based on the observations that the action of PAF is blocked by dopamine agonists and the PAF receptor antagonists L 652731 and SRI 63072. The structural analogs 1-O-alkyl-2-oleoyl-sn-glycero-3-phosphocholine and 1-O-alkyl-2-acetyl-sn-glycero-3-phosphoethanolamine, which lack the biological activity of PAF, are not able to stimulate PRL release over the dose range 0.2-2 microM. In addition, the PAF precursor lyso PAF and diacyl-sn-glycero-3-phosphocholine (phosphatidylcholine) were ineffective in stimulating PRL release. PAF induced the secretion of PRL and GH but not that of LH or TSH from hemipituitaries in short term incubations. PAF did not effect PRL release from GH3 cells. In conclusion, these data indicate that PAF stimulates PRL release from primary cultures of rat anterior pituitary cells in a dose-related, rapid, and specific manner.     

Inhibition of hypothalamic somatostatin release by beta-adrenergic antagonists

A number of in vivo studies suggest that hypothalamic somatostatin (SRIF) tone is stimulated by the beta-adrenergic system. Employing dispersed adult male rat hypothalamic cells, we studied the effects of beta-adrenergic antagonists on the release of hypothalamic SRIF. Propranolol, at concentrations of 1-100 microM, had no detectable effect on basal SRIF release, but caused dose-dependent inhibition of SRIF release stimulated by ouabain. Two other beta-adrenergic antagonists, labetolol and metoprolol, also caused inhibition of ouabain-stimulated SRIF release. The alpha 2-agonist clonidine was without effect on SRIF release under basal or stimulated conditions. GH secretion from monolayers of dispersed rat anterior pituitary cells was also examined. Propranolol (1-100 microM) had no significant effect on basal GH secretion or GH secretion stimulated by rat GRF. In conclusion, 1) beta-adrenergic antagonists caused inhibition of stimulated SRIF release; 2) clonidine had no detectable effect on SRIF release; and 3) propranolol did not affect GH secretion in vitro. These findings support the hypothesis that beta-adrenergic antagonists augment GH responsivity by inhibiting hypothalamic SRIF release.