Receptors and neurohormones in human pituitary adenomas

In order to go further into the pathogenesis of human pituitary adenomas, we studied receptors for neurohormones (thyroliberin, TRH; dopamine, DA; somatostatin, SRIH), for estradiol and epidermal growth factor (EGF) thought to influence hormone secretion and/or cell growth. The following results were obtained: (1) the receptors listed above, with the exception of EGF receptors in the adenomas, are present in normal pituitary tissue and in prolactin (PRL)- and growth hormone (GH)-secreting adenomas; (2) they are functional and their affinities are not different in normal or tumoral tissues; (3) their density is variable and depends on the type of secreting adenoma (GH or PRL), the size of the tumor and the plasma level of the hormone which is secreted, and (4) in nonsecreting adenomas, only TRH receptors are found with characteristics identical to those observed in secreting adenomas. We also showed that TRH is contained in normal and tumoral pituitary tissues. TRH and SRIH are released in vitro from adenomatous cells in large amounts, suggesting their possible synthesis by the pituitary. In both cases a local regulation is observed. TRH release is stimulated in the presence of DA while SRIH is inhibited in the presence of TRH. This neuropeptide release may be implicated in the pituitary hormone regulation through a paracrine or an autocrine mechanism. Thus, the neurohormone receptors found in pituitary adenomas should be dependent on a more complex regulation than it has been envisaged till now.     

The species specificity of growth hormone requires the cooperative interaction of two motifs

Primate growth hormones (GH) activate both primate and non-primate somatotrophic receptors (GH receptors), but non-primate GHs do not activate primate GH receptors. Previous studies argued the interaction of Asp(171) of human GH and Arg(43) of the receptor produced an attractive ionic interaction. In non-primate GHs, His(170) replaces the homologous Asp(171), producing a repulsive interaction with Arg(43) of the primate receptor which was believed to reduce the attraction of non-primate GH for the human GH receptor, thus providing species specificity. In this report, H170D bovine GH had activity and affinity for human GH receptors approaching those of human GH. In contrast, replacing Asp(171) of human GH with His did not significantly reduce somatotrophic activity, indicating that species specificity is not wholly explained by this residue's interaction with Arg(43) of the receptor. Deletion of either Phe(44) (a residue present only in primate GHs) or residues 32-46 (20-kDa form of human GH) each only marginally reduced somatotrophic activities. But the combination of the D171H mutation with either DeltaPhe(44) or Delta32-46 in human GH reduced binding and activity in a greater than additive fashion, indicated a functional interaction between these distant structural features. In bovine GH addition of phenylalanine at position 44 increased the somatotrophic activity and receptor affinity in cells containing the human GH receptor. The combination of the H170D mutation and the addition of phenylalanine at position 44 created a bovine GH with activity indistinguishable from wild-type human GH. Based on evidence from both bovine and human GHs, the cooperative interaction of these two distant motifs determined the species specificity and indicated that structural plasticity was a critical feature necessary for the species specificity of somatotrophic activity.     

Effect of the GABAA agonist muscimol on prolactin secretion from human prolactin-secreting adenomas and GH3 rat pituitary tumour cells.

The effect of muscimol, a specific potent GABAA receptor agonist, on prolactin release from human prolactin-secreting tissue was investigated using a perifusion system. Perifusion studies on normal rat anterior pituitary tissue, which has identical GABA receptors to those found in normal human pituitary glands, show that muscimol has a specific biphasic effect on prolactin release. This is characterized by an initial transient stimulation (222.3 +/- 21.6% of basal) lasting for 5-10 min followed by a more prolonged inhibitory phase (63.9 +/- 3.1% inhibition of basal). Five human prolactin-secreting adenomas were studied, and in none of the tumours could a biphasic response be demonstrated. One of the prolactin-secreting adenomas had a blunted inhibitory response, but the other 4 showed no inhibitory effect of muscimol on prolactin release. Muscimol had no significant effect on basal or thyrotropin-releasing-hormone (TRH)-stimulated prolactin secretion from GH3 rat pituitary tumour cells. These studies suggest that the GABAergic effect on prolactin secretion is absent or altered in both rat and human prolactin-secreting tumour cells.     

Release of thyrotropin releasing hormone (TRH) from human prolactin-secreting pituitary adenoma cells. Modulation by dopamine

We found, by radioimmunoassay, that thyrotropin-releasing-hormone (TRH) was present in human prolactin (PRL)-secreting adenomas (mean: 89 +/- 45 (SEM) fmol/mg proteins) and was released by perifused adenomatous cells at levels varying from 5 to 60 fmol/10(6) cell/2 min. TRH release was increased in the presence of dopamine (DA) 10(-6) M but was not modified by the presence of somatostatin (SRIH) 10(-6) M.     

In vitro secretion of somatostatin (SRIH) by human adenomatous somatotropic cells. Relation with somatotropic hormone (GH) release and modulation by thyroliberin (TRH)

SRIH and GH secretions by GH-secreting adenomatous human pituitary cells were analyzed in vitro in a perifusion system. Of the 13 adenomas studied, 7 secreted SRIH, in variable amounts (50 to 700 pg/ml/2 min., corresponding to 600 10,700 pg for the total experiment. SRIH secretion increased during the perifusion, the highest levels being observed at the end of the perifusion. GH secretion also varied from one adenoma to the other (6 to 500 ng/ml). In most cases, the secretion profiles were negatively correlated, GH secretion decreasing while SRIH secretion was increasing. In the presence of 10(-7) M TRH, GH secretion increased while that of SRIH decreased. The hypothesis of a paracrine and/or an autocrine role for SRIH as well as its possible in situ synthesis are discussed.     

Stimulation of growth hormone release in dwarf and normal chickens by thyrotrophin releasing hormone (TRH) or human pancreatic growth hormone releasing factor (hpGRF)

The effect of thyrotrophin releasing hormone (TRH) or human pancreatic growth hormone releasing factor (hpGRF) on growth hormone (GH) release was studied in both dwarf and normal Rhode Island Red chickens with a similar genotype except for a sex-linked dw gene. Both TRH (10 micrograms/kg) and hpGRF (20 micrograms/kg) injections stimulated plasma GH release within 15 min in young and adult chickens. The increase in GH release was higher in young cockerels than that in adult chickens. The age-related decline in the response to TRH stimulation was observed in both strains, while hpGRF was a still potent GH-releaser in adult chickens. The maximal and long acting response was observed in young dwarf chickens, suggesting differences in GH pools releasable by TRH and GRF in the anterior pituitary gland. The pituitary gland was stimulated directly by perifusion with hpGRF (1 microgram/ml and 10 micrograms/ml) or TRH (1 microgram/ml). Repeated perifusion of GRF at 40 min intervals blunted further increase in GH release, but successive perifusion with TRH stimulated GH release. The results suggest the possibility that desensitization to the effects of hpGRF occurs in vitro and that the extent of response depends on the number of receptors for hpGRF or TRH and/or the amount of GH stored in the pituitary gland.     

Oral thyrotropin-releasing hormone (TRH) test in acromegaly

The effects of 40 mg oral and 200 microgram intravenous TRH were studied in patients with active acromegaly. Administration of oral TRH to each of 14 acromegalics resulted in more pronounced TSH response in all patients and more pronounced response of triiodothyronine in most of them (delta max TSh after oral TRh 36.4 +/- 10.0 (SEM) mU/l vs. delta max TSH after i.v. TRH 7.7 +/- 1.5 mU/l, P less than 0.05; delta max T3 after oral TRH 0.88 +/- 0.24 nmol/vs. delta max T3 after i.v. TRH 0.23 +/- 0.06 nmol/l, P less than 0.05). Oral TRH elicited unimpaired TSH response even in those acromegalics where the TSH response to i.v. TRH was absent or blunted. In contrast to TSH stimulation, oral TRH did not elicit positive paradoxical growth hormone response in any of 8 patients with absent stimulation after i.v. TRH. In 7 growth hormone responders to TRH stimulation the oral TRH-induced growth hormone response was insignificantly lower than that after i.v. TRH (delta max GH after oral TRH 65.4 +/- 28.1 microgram/l vs. delta max GH after i.v. TRH 87.7 +/- 25.6 microgram/l, P greater than 0.05). In 7 acromegalics 200 microgram i.v. TRH represented a stronger stimulus for prolactin release than 40 mg oral TRH (delta max PRL after i.v. TRH 19.6 +/- 3.22 microgram/, delta max PRL after oral TRH 11.1 +/- 2.02 microgram/, P less than 0.05). Conclusion: In acromegalics 40 mg oral TRH stimulation is useful in the evaluation of the function of pituitary thyrotrophs because it shows more pronounced effect than 200 microgram TRH intravenously. No advantage of oral TRH stimulation was seen in the assessment of prolactin stimulation and paradoxical growth hormone responses.     

Synthetic human pancreatic growth hormone releasing factor (GRF) stimulates growth hormone secretion in the domestic fowl (Gallus domesticus)

Synthetic human pancreatic Growth Hormone-Releasing Factor (hpGRF) elevated the plasma concentration of growth hormone (GH) in young and adult domestic fowl. This in vivo effect of hpGRF appeared to be largely similar for both the 32 amino-acid (hpGRF 1-32) or 40 amino-acid (hpGRF 1-40) polypeptide, although the effect of hpGRF 1-32 was more prolonged than that of hpGRF 1-40 in adult domestic fowl. The increase in plasma GH concentrations following hpGRF administration (10 micrograms/kg) was somewhat greater in young than adult chickens (the increase in plasma concentration of GH being 230 ng/ml at 1 week old, 282 ng/ml at 6 week old, 241 ng/ml at 10 weeks and 150 ng/ml in adults). In the adult domestic fowl hpGRF stimulated a greater increase in the plasma concentration of GH than did thyrotropin-releasing hormone (TRH). However in the young chicks TRH was more active. The in vitro release of GH from dispersed chicken pituitary cells was elevated by hpGRF (1-32) and hpGRF (1-40).     

Paradoxical growth hormone response following thyroid-stimulating hormone administration in the polycystic ovary syndrome

Growth hormone (GH) and prolactin (PRL) responses after TRH administration were studied in 31 women presenting with the clinical, biochemical and ultrasonographic characteristics of the polycystic ovarian (PCO) syndrome; their results were compared with those of 20 normally menstruating women investigated during the early follicular phase of the cycle. Based on the GH responses two PCO subgroups were observed: (a) nonresponders (n = 16) who showed delta max GH responses (0.7 +/- 0.27 ng/ml, x +/- SE) similar to those of the normals (0.97 +/- 0.20 ng/ml), and (b) responders (n = 15), 48.4% of the PCO patients who showed a paradoxical increase in GH levels (delta max GH, 18.0 +/- 1.96 ng/ml) following thyrotropin-releasing hormone (TRH) administration significantly higher than those observed either in nonresponder PCO patients or in normals. Furthermore, basal GH levels were found to be significantly higher in the responder PCO subgroup (5.65 +/- 0.75 ng/ml) compared to either nonresponders (1.58 +/- 0.21 ng/ml) or normals (1.8 +/- 0.18 ng/ml). However, no correlation was found between basal GH levels and delta max GH responses observed. Additionally, basal PRL and delta max PRL levels following TRH administration did not differ either between the two PCO subgroups or those observed in normal controls. delta 4A, T and E2 levels were similar between the two PCO subgroups. No correlation was found between the delta max GH responses to delta max PRL or the post-luteinizing hormone-releasing hormone stimulation test delta max luteinizing hormone:follicle-stimulating hormone ratio observed or to steroid levels.(ABSTRACT TRUNCATED AT 250 WORDS)     

Receptor number determines latency and amplitude of the thyrotropin-releasing hormone response in Xenopus oocytes injected with pituitary RNA

TRH evokes depolarizing membrane electrical responses in Xenopus laevis oocytes injected with RNA from pituitary cells. We have shown previously that the amplitude of this response is directly proportional to the dose of TRH and the amount of RNA injected. Herein we show that the number of TRH receptors expressed on oocytes after injection of rat pituitary (GH3) cell RNA or mouse thyrotropic (TtT) tumor RNA determines the latency as well as the amplitude of the response. In oocytes injected with a maximally effective amount of GH3 cell RNA, the latency of the response decreased from a maximal duration of 103 +/- 16 to 10 +/- 1 sec when the TRH concentration was increased from 5 to 3000 nM. When oocytes injected with different amounts of GH3 cell RNA were stimulated with 3000 nM TRH, the latency decreased from 31 +/- 4 to 11 +/- 0.5 sec when the amount of RNA injected was increased from 30 to 400 ng. Specific binding of [3H]methylhistidine-TRH increased when increasing amounts of TtT poly(A)+ RNA was injected, and binding correlated with increased response amplitude. To show that these effects were caused by mRNA for the TRH receptor and did not depend on other mRNAs, TtT poly(A)+ RNA was fractionated on a sucrose gradient. Using RNA from each fraction, there was an inverse correlation between response amplitude and latency. For size-fractionated RNA, as for unfractionated RNA, there was a direct correlation between specific [3H]methylhistidine-TRH binding and response amplitude.(ABSTRACT TRUNCATED AT 250 WORDS)     

PRF activity of VIP in vitro (author's transl)

The effect of VIP on prolactin secretion from incubated rat hemipituitaries was characterized. Under these conditions, the secretion of GH, LH, FSH, ACTH was not affected, indicating that the effect of VIP is hormone specific. The stimulation of prolactin was dose-dependent, with an apparent affinity of VIP of 10.9 +/- 3.1 nM and a maximal stimulation of 57.7 +/- 4.2%. Secretin, a structurally related peptide, was also active at higher concentrations, whereas another partial analogue, glucagon, was ineffective. Furthermore, VIP does not act through pituitary DA receptors since alpha-flupentixol, a potent dopaminergic antagonist, does not block the stimulation of prolactin secretion by VIP. In addition, stimulation by VIP and TRH was additive. Naloxone and met-enkephalin were ineffective on the VIP effect on prolactin release. In contrast, SRIF seems to inhibit the VIP stimulation of prolactin release. Our data suggest that VIP, which was found in the hypothalamo-hypophyseal blood at concentrations of the same order of magnitude as that found to stimulate PRL in vitro, could be a physiological PRF.     

A decreased capacity of hepatic growth hormone (GH) receptors and failure of thyrotrophin-releasing hormone to stimulate the peripheral conversion of thyroxine into triiodothyronine in sex-linked dwarf broiler hens

The effect of two different doses of thyrotrophic releasing hormone (TRH) upon the plasma levels of growth (GH) and thyroid hormones in both sex-linked dwarf (dw) and normal (Dw) broiler hens was determined. In normal hens, 1.5 and 24 microg TRH/kg increased the GH plasma concentrations after 15 min. Plasma concentrations of T3 increased significantly 1 h after TRH injection, whereas T4 concentration decreased after 2 following injection of 24 microg/kg TRH. In dwarf hens both doses of TRH increased the plasma concentrations of GH and the GH response lasted longer. However, TRH was ineffective in raising T3 and T4 levels. Saline-injected dwarf birds showed no differences in plasma T4 and T3 levels in comparison with normal hens. A smaller number of hepatic cGH receptors was found in dwarf hens, whereas the affinity of the hepatic GH receptor was not influenced by the genotype. It is concluded that the sex-linked dwarf broiler hen is unable to respond to a TRH-induced GH stimulus probably because of a deficiency in hepatic GH receptors resulting in a failure to stimulate the T4 to T3 converting activity.     

Increase of IGF I binding to erythrocyte receptors during the TRH-test: correlation between IGF I binding and thyroid hormone values

The effects of TRH on insulin-like growth factor I receptors were investigated on erythrocytes from 7 GH-deficient children having plasma GH levels less than 10 ng/ml during two provocation tests. Intravenous injection of synthetic TRH (0.2 mg/m2) was followed by a marked increase of IGF I binding on erythrocytes, from 3.9% +/- 0.3% to 5.9% +/- 0.3% (P less than 0.005) after 1 hour and 7.3% +/- 0.4% (P less than 0.005) after 2 hours. The IGF I binding variations were due to an increase in both the receptor affinity and the number of sites. The levels of plasma GH, IGF I, T3, T4, free T4, TSH and prolactin having been determined during the TRH test at 0, 1 hour, and 2 hours after the injection, the increase in the IGF I binding to erythrocytes at the same time correlated with the rise of thyroid hormones: triiodothyronine T3 (P less than 0.001) and thyroxine T4 (P less than 0.005) and not with the level of the other hormones. These findings suggest that thyroid hormones play a role in the regulation of insulin-like growth factor I receptors.     

Thyrotropin-releasing hormone regulates the number of its own receptors in the GH3 strain of pituitary cells in culture.

Thyrotropin-releasing hormone (TRH), a hypothalamic tripeptide, binds rapidly and reversibly to specific membrane receptors on GH3 cells, a clonal strain of rat pituitary cells grown in culture. GH3 cells were incubated for 1-72 hr with unlabeled TRH, washed, and then incubated for 1 hr with [3H]TRH. Under these conditions 80% of any bound, unlabeled TRH exchanges with [3H]TRH in the medium, and the amount of radioactivity bound to the cells gives a measure of the number of TRH receptors. In GH3 cells, the number of available TRH receptors decreased from 92% of control after 1 hr to 35% after 48 or 72 hr of incubation with unlabeled TRH. Binding of [3H]TRH to both intact control and TRH-treated cells was half-maximal at 8 nM [3H]TRH, but the maximum amount of [3H]TRH bound was decreased by 75% in cells previously incubated for 48 hr with unlabeled TRH. Equilibrium binding studies were performed using membrane fractions prepared from control cells and cells previously exposed to TRH for various periods. The dissociation constant of the TRH-receptor complex was the same in all cases, but the maximum amount of TRH bound decreased progressively in membrane fractions from cells incubated with TRH for 1-51 hr. TRH receptors were not found in cytoplasmic fractions of control or TRH-treated cells. The loss of TRH receptors was reversible within 4 days. In the continued presence of the tripeptide the number of receptors remained low for 12 days. After incubation for 2 days with different concentrations of TRH, the number of receptors was decreased to 33% of control at 100-300 nM TRH, and half of this decrease occurred at about 1 nM TRH; half-maximal biological responses occur at 2 nM TRH. The biologically active Ntau-methylhistidyl derivative of TRH also effected a loss of receptors, while three inactive analogs of TRH did not cause reductions in the number of TRH receptors. In cultures incubated for 40 hr with cycloheximide, protein synthesis was inhibited by 85%, but the number of TRH receptors was 76% of control suggesting that the receptor has a long half-life. When GH3 cells were incubated with cycloheximide plus TRH, the number of TRH receptors decreased by only 23% as compared to a decrease of 73% in cells incubated with TRH alone, suggesting that receptor loss is partially dependent on active protein synthesis. We conclude that in GH3 cells TRH regulates the number of its own receptors.     

Differential regulation of insulin-like growth factor-(IGF) I and IGF-binding protein (IGFBP) secretion by human peripheral blood mononuclear cells

BACKGROUND: Recent studies have shown that immunocompetent cells synthesize and express growth hormone (GH), growth hormone receptors (GH-R), insulin-like growth factor I (IGF-I), IGF-I receptors (IGF-I-R) and different insulin-like growth factor binding proteins (IGFBPs). The aim of the current study was to evaluate the regulation of IGFBP and IGF-I secretion from immunocompetent cells by different mitogens. METHODS/RESULTS: We studied the in vitro secretion pattern of IGFBPs and IGF-I from human peripheral blood mononuclear cells (PBMC), derived from 10 normal adults and 8 GH-deficient patients with adult onset. In serum-free conditioned medium of unstimulated PBMC, derived from normal adults, Western ligand blotting (1D-WLB) revealed a 24-kD, a 34-kD and a 39/43-kD doublet band to be most prominent. According to their molecular weight and two-dimensional Western ligand blot analysis (2D-WLB), these bands are deglycosylated IGFBP-4, IGFBP-2 and IGFBP-3, respectively. When the cells were treated with the T-cell mitogen phytohemagglutinin (PHA) (10 microg/ml), a differential stimulation of IGFBPs was found with a 2.57 +/- 0.48-fold increase of IGFBP-4 (p < 0.01), a 1.55 +/- 0.13-fold increase of IGFBP-2 (p < 0.01), and a 1.35 +/- 0.19-fold increase of IGFBP-3 (n.s.). In contrast, treatment with the B-cell mitogen pokeweed mitogen (PWM) (10 microg/ml) caused only a modest 1.40 +/- 0.07-fold increase of IGFBP-4 (p < 0.01). Treatment with rhGH (100 ng/ml) or rhIGF-I (200 ng/ml) caused no significant induction of any specific band, respectively. In contrast to the secretion pattern of IGFBPs, IGF-I secretion of the PBMC was not stimulated by either PHA or PWM, but showed a significant increase after GH incubation (p < 0.01). A similar differentiated secretion pattern of IGFBPs and IGF-I was also observed in the conditioned medium of PBMC, derived from GH-deficient patients. CONCLUSION: In summary, at least three different IGFBPs are secreted by human PBMC. Secretion of IGFBPs by PBMC is differentially regulated by different lymphocyte mitogens. Secretion of IGFBPs by PBMC is independent of GH or IGF-I, whereas the secretion of IGF-I is stimulated by GH. PBMC derived from normal adults and GH-deficient patients show similar patterns of IGF-I and IGFBPs secretion, thus indicating that the paracrine/autocrine IGF-I-IGFBPs interactions of the PBMC are not altered by pituitary GH deficiency.     

Thyrotropin releasing hormone (TRH) selectively binds and activates the melanocortin 1 receptor.

We tested the endogenous tripeptide TRH (thyrotropin releasing hormone) ability to bind to MC (melanocortin) receptor subtypes. We discovered that TRH binds to the human MCI receptor expressed in COS cells and to murine B16 melanoma cells with 5790+/-1010 nM and 6370+/-1260 nM Ki's, respectively. TRH did not bind to the human MC3, MC4 or MC5 receptor subtypes. Moreover, TRH also stimulated cAMP production in murine B16 melanoma cells reaching the same maximum level of cAMP as found for alpha-MSH. However, several analogues of TRH, including TRH-OH, TRH-Gly-NH2 and other analogues, where each of the three amino acid residues in TRH had been exchanged by a related residue, did not bind to any of the MC receptors tested in this study. C(alpha) atoms of molecular models of TRH and the core of a MSH peptide were aligned with r.m.s. of 0.01 A. Moreover, TRH could be docked into a binding pocket of a molecular model of the MC1 receptor at only a little higher energy than a short cyclic MSH peptide. The data indicate a similarity in the mode of TRH and MSH activation of the MCI receptor.     

Growth hormone-binding proteins in high-speed cytosols of multiple tissues of the rabbit

Soluble, specific binding protein(s) for growth hormone (GH) have been identified and partially characterized in high-speed cytosolic preparations from a number of rabbit tissues. The binding of 125I-labelled human GH to proteins in liver, heart, adipose tissue, skeletal muscle and kidney cytosols was dependent on time and cytosolic protein concentration. By Scatchard analysis, the binding affinities (KA: (2-7) X 10(9) M-1) were somewhat higher than those generally reported for membrane GH receptors. The binding proteins had a greater specificity for somatotrophic hormones than lactogenic hormones, although the kidney appeared to have, in addition, a lactogen-binding protein. By gel filtration, the Mr of the cytosolic GH-binding protein was approximately 100 000 in all tissues. None of the binding proteins was detectable by the poly(ethylene glycol) precipitation method used widely for soluble hormone receptors. The cytosolic GH-binding proteins also cross-reacted with a monoclonal antibody to the rabbit liver membrane GH receptor. These results indicate the ubiquitous presence of apparently naturally soluble GH-binding proteins in the cytosolic fractions of several tissues in the rabbit. Of great interest is their presence in muscle, where GH receptors or binding proteins have not previously been detected, despite muscle being recognized as a classical GH target tissue.     

Effect of 9-cis retinoic acid on dopamine D2 receptor expression in pituitary adenoma cells

The dopamine receptor subtype 2 (D2R) promoter contains a functional retinoic acid response element involved in the control of D2R expression. The aim of the study was to evaluate the effect of 9-cis retinoic acid (9-cis RA) on D2R protein expression in human pituitary adenomas and GH3 cell line. Treatment with 9-cis RA (100 nM for 48 hrs) caused a 109 +/- 32% increase of basal D2R levels in five of eight growth hormone (GH)-secreting adenomas (GH-omas), a 129 +/- 28% increase in 7 of 11 nonfunctioning adenomas, and no effect in two resistant prolactinomas by Western blotting. The lack of D2R induction in some tumors was not associated with a different pattern of retinoid x receptor (RXR) and retinoic acid receptor (RAR) isoform expression that was similar in all tumors by immunohistochemistry. While the induction of D2R did not affect the slight but significant inhibitory effect exerted by dopamine (10 nM) on in vitro GH release by GH-oma cultured cells, in pituitary GH3 cell lines cis-9 RA enhanced the dopamine-induced inhibition of in vitro GH release (% inhibition: 16 +/- 2 versus 26 +/- 5, P < 0.05), cell proliferation (25 +/- 2% versus 44 +/- 5%, P < 0.05) and cell viability (16 +/- 0.8% versus 29 +/- 1%, P < 0.05), likely by activating caspase-3 (28 +/- 3% versus basal, P < 0.05). In conclusion, this study provides novel evidence for a permissive role of retinoids on the expression of D2R in a good proportion of pituitary tumors and on the generation of pro-apoptotic signals in GH3 cell line.     

Thyrotropin-releasing hormone (TRH) and phorbol myristate acetate decrease TRH receptor messenger RNA in rat pituitary GH3 cells: evidence that protein kinase-C mediates the TRH effect.

In a previous report we showed that TRH-induced down-regulation of the density of its receptors (TRH-Rs) on rat pituitary tumor (GH3) cells was preceded by a decrease in the activity of the mRNA for the TRH-R, as assayed in Xenopus oocytes. Here we report the effects of TRH, elevation of cytoplasmic free Ca2+ concentration, phorbol myristate acetate (PMA), and H-7 [1-(5-isoquinolinesulfonyl)2-methylpiperazine dihydrochloride], an inhibitor of protein kinases, on the levels of TRH-R mRNA, which were measured by Northern analysis and in nuclease protection assays using probes made from mouse pituitary TRH-R cDNA, in GH3 cells. These agents were studied to gain insight into the mechanism of the TRH effect, because signal transduction by TRH involves generation of inositol 1,4,5-trisphosphate and elevation of cytoplasmic free Ca2+ concentration, which leads to activation of Ca2+/calmodulin-dependent protein kinase, and of 1,2-diacylglycerol, which leads to activation of protein kinase-C. TRH (1 microM TRH, a maximally effective dose) caused a marked transient decrease in TRH-R mRNA that attained a nadir of 20-45% of control by 3-6 h, increased after 9 h, but was still below control levels after 24 h. Elevation of the cytoplasmic free Ca2+ concentration had no effect on TRH-R mRNA. A maximally effective dose of PMA (1 microM) caused decreases in TRH-R mRNA that were similar in magnitude and time course to those induced by 1 microM TRH. H-7 (20 microM) blocked the effects of TRH and PMA to lower TRH-R mRNA to similar extents.(ABSTRACT TRUNCATED AT 250 WORDS)     

Ectopic growth hormone-releasing hormone (GHRH) syndrome in a case with multiple endocrine neoplasia type I

A 36-yr-old man with multiple endocrine neoplasia (MEN) type I had an ectopic growth hormone-releasing hormone (GHRH) syndrome due to a GHRH-secreting pancreatic tumor. The immunoreactive (IR)-GHRH concentration in his plasma ranged from 161 to 400 pg/ml (299 +/- 61 pg/ml, mean +/- SD; normal, 10.4 +/- 4.1 pg/ml), and a significant correlation was found between his plasma IR-GHRH and GH (r = 0.622, p less than 0.02). After removal of the pancreatic tumor, the high plasma GH concentration returned to nearly the normal range (42.2 +/- 31.3 to 9.6 +/- 3.8 ng/ml). These changes paralleled the normalization of his plasma IR-GHRH (16.1 +/- 3.8 pg/ml) and some of his symptoms related to acromegaly improved. However, plasma GH (7.7 +/- 1.3 ng/ml) and IGF-I (591 +/- 22 ng/ml) concentrations were high at 12 months after surgery, suggesting adenomatous changes in the pituitary somatotrophs. Before surgery, exogenous GHRH induced a marked increase in plasma GH, and somatostatin and its agonist (SMS201-995) completely suppressed GH secretion, but not IR-GHRH release. No pulsatile secretion of either IR-GHRH or GH was observed during sleep. An apparent increase in the plasma GH concentration was observed in response to administration of TRH, glucose, arginine or insulin, while plasma IR-GHRH did not show any fluctuation. However, these responses of plasma GH were reduced or no longer observed one month and one year after surgery. These results indicate that 1) a moderate increase in circulating GHRH due to ectopic secretion from a pancreatic tumor stimulated GH secretion resulting in acromegaly, and evoked GH responses to various provocative tests indistinguishable from those in patients with classical acromegaly, and 2) the ectopic secretion of GHRH may play an etiological role in the pituitary lesion of this patient with MEN type I.