Differential in vitro secretion of gonadotropin-releasing hormone (GnRH) and [hydroxyproline]GnRH from the rat hypothalamus during postnatal development.

The differential secretion of gonadotropin-releasing hormone (GnRH) and [hydroxyproline9]GnRH (HypGnRH) has been recently reported from the adult rat hypothalamus. We report here in vitro cosecretion of HypGnRH and GnRH by the hypothalamus of 2-45 day-old-rats and provide evidence that they are differentially regulated throughout development. The secretion of both forms of GnRH was increased in a dependent manner during depolarization by high K+ solutions, and was stimulated by forskolin and 12-O-tetradecanoylphorbol-13-acetate (TPA), activators of adenylate cyclase and protein kinase C pathways, respectively. The proportion of HypGnRH in the release of GnRH-like peptides remained stable and high (33-40%) under basal and K+-induced conditions until days 13 and 21, respectively. By contrast, the proportion of HypGnRH in the total GnRH-like content of the developing hypothalamus continuously decreased (from 37% to 14%). Similarly, the proportion of HypGnRH: total GnRH-like material released remained stable in TPA- (30%) and forskolin- (50%) induced secretion until postnatal day 8. Evaluation of release over tissue store ratios revealed a 1.3-to 2.8-fold higher release of HypGnRH compared to GnRH according to the different secretions and postnatal periods examined. The preferential recruitment of HypGnRH was maintained under basal and K+ conditions during postnatal development, but it disappeared under TPA stimulation from day 13 onwards. After forskolin stimulation, the preferential mobilization of HypGnRH was markedly reduced from day 2 to day 13 but recovered its high perinatal level during puberty. Taken together, our results support the hypothesis that HypGnRH may play a specific role in development. In addition, a specific function of this peptide taking place during puberty through the activation of the adenylate cyclase pathway is suggested.

Somatostatin increases mitogen-induced IL-2 secretion and proliferation of human Jurkat T cells via sst3 receptor isotype.

The neuropeptide somatostatin (SRIF) modulates normal and leukemia T cell proliferation. However, neither molecular isotypes of receptors nor mechanisms involved in these somatostatin actions have been elucidated as yet. Here we show by using RT-PCR approach that mitogen-activated leukemia T cells (Jurkat) express mRNA for a single somatostatin receptor, sst3. This mRNA is apparently translated into protein since specific somatostatin binding sites (K11 = 78 +/- 3 pM) were detected in semipurified plasma membrane preparations by using 125I-Tyr1-SRIF14 as a radioligand. Moreover, somatostatin inhibits adenylyl cyclase activity with similar efficiency (IC50 = 23 +/- 4 pM) thus strongly suggesting a functional coupling of sst3 receptor to this transduction pathway. The involvement of sst3 receptor in immuno-modulatory actions of somatostatin was assessed by analysis of neuropeptide effects on IL-2 secretion and on proliferation of mitogen-activated Jurkat cells. Our data show that in the concentrations comprised between 10 pM and 10 nM, somatostatin potentiates IL-2 secretion. This effect is correlated with somatostatin-dependent increase of Jurkat cell proliferation since the EC50 concentrations for both actions were almost identical (EC50 = 22 +/- 9 pM and EC50 = 12 +/- 1 pM for IL-2 secretion and proliferation, respectively). Altogether, these data strongly suggest that in mitogen-activated Jurkat cells, somatostatin increases cell proliferation through the increase of IL-2 secretion via a functional sst3 receptor negatively coupled to the adenylyl cyclase pathway.

Differential involvement of calcium channels and protein kinase-C activity in GnRH-induced phospholipase-C, -A2 and -D activation in a gonadotrope cell line (alpha T3-1).

The mode of action of GnRH on pituitary gonadotropes involves metabolism of phospholipids, protein kinase-C (PKC) and voltage sensitive Ca2+ channels (VSCC) activation. We have studied the differential role of PKC and VSCC on the coupling of the GnRH receptor with phospholipases-C (PLC), -A2 (PLA2) and -D (PLD) activities in a gonadotrope cell line (alpha T3-1), by measuring the production of inositol phosphates (IPs), arachidonic acid (AA) and phosphatidylethanol (PEt) respectively. We demonstrated that in these cells GnRH stimulated through a specific receptor, IPs formation, a rapid and sustained diacylglycerol generation, consequently AA release and a delayed PEt production in a dose-dependent manner. In contrast to GnRH-induced PLC activity, the PLA2 and PLD stimulation by the neuropeptide involved Ca2+ mobilization via VSCC activation. BAY-K8644 a VSCC agonist significantly potentiated, while the VSCC antagonist nitrendipine markedly inhibited GnRH-induced AA release and PEt production. TPA, a phorbol ester which induced a rapid and important redistribution of PKC, although unable to elicit PLC or PLA2 stimulation, specifically provoked PLD activation in a PKC-dependent but Ca(2+)-independent manner. The PKC stimulation by TPA significantly inhibited the GnRH-stimulated IPs and AA formation, while it potentiated the GnRH-evoked PEt production. This negative feed-back of PKC on GnRH-Induced PLC and PLA2 activities was reversed when PKC was either down regulated after long TPA treatments or inhibited by the PKC inhibitors, staurosporine or GF109203X. The GnRH-induced PEt formation was markedly diminished in PKC depleted cells or after PKC inhibition. Under such conditions, both agonist and antagonist of VSCC became less effective in modulating the remaining GnRH-evoked PEt formation. These results suggest that PKC, in coordination with Ca2+, plays a key role in regulating the cross-talk between the multiple phospholipases implicated in the GnRH signal transduction.

Nature and bioactivity of gonadotropin-releasing hormone (GnRH) secreted during the GnRH surge.

Previous studies have demonstrated a neural action of estradiol in inducing a surge of GnRH in the ewe. However, although the GnRH and LH surges began concurrently, the GnRH surge consistently continued well beyond the surge of LH. Three experiments were conducted to test the hypothesis that the termination of the LH surge results from the secretion of a relatively inactive variant of GnRH during the later phases of the GnRH surge. In the first experiment, hypophyseal portal blood collected during an estrogen-induced LH surge was analyzed for GnRH immunoreactivity using two antibodies having specificity for the N- or C-terminal portion of the GnRH molecule. The duration, amplitude, and time course of the GnRH surge were found to be similar irrespective of the antisera used. In a second experiment, a competitive GnRH antagonist was administered at the beginning of the estrogen-induced GnRH/LH surge at a dose capable of blocking pituitary responsiveness for approximately half the duration of the GnRH surge. Antagonist treatment did not result in any change in the time of onset of the GnRH surge, but there was no increase in LH that naturally occurs coincident with onset of the GnRH surge. Rather, a persistent increase in LH secretion was observed during the latter stages of the GnRH surge, indicating that the GnRH molecules secreted at this time were biologically active. Finally, a sensitive and specific ovine pituitary cell bioassay was used to test bioactivity of GnRH in hypophyseal portal blood during different phases of the GnRH surge. GnRH bioactivity in samples collected early in the GnRH surge was greater than that before the onset of the GnRH surge but no greater than that collected during the descending limb of the surge. The results of all three experiments fail to support the hypothesis that the LH surge ends because of a change in the nature of the GnRH secreted. Rather they show that GnRH secreted throughout the surge is biologically active. Thus, the termination of the LH surge before that of the GnRH surge occurs for reasons other than lack of a bioactive GnRH signal.

Characterization of [hydroxyproline9]luteinizing hormone-releasing hormone and its smallest precursor forms in immortalized luteinizing hormone-releasing hormone-secreting neurons (GT1-7), and evaluation of their mode of action on pituitary cells.

[Hydroxyproline9]luteinizing hormone-releasing hormone ([Hyp9]LHRH), an endogenous hydroxylated post-translational product of the LHRH sequence, has been isolated from mammalian hypothalamus. Using the LHRH-hypothalamic cell line (GT1-7) of fetal origin, we attempted to define the substrates available for the hydroxylation process during LHRH synthesis and to characterize immunologically the [Hyp9]LHRH and pro-[Hyp9]LHRH forms with anti-LHRH antibodies of different specificities after separation by HPLC. Their biological activity and mode of action were evaluated and compared to that of LHRH and LHRH intermediate precursors in normal pituitary cells and in a gonanodotrope cell line alpha T3-1. immunoreactivity was progressively increased in cells and media during cell culture. [Hyp9]LHRH and its two smallest precursor forms ([Hyp9]LHRH-(Gly11) and -(11-13)) were detected in cells and in media. They were simultaneously detected with the homologous LHRH molecular forms indicating that the hydroxylation occurs early in the processing of pro-LHRH. [Hyp9]LHRH-like molecules were more abundant than LHRH forms in media. This predominant release may thus represent a physiological process occurring during fetal life. Free acid forms of both decapeptides were detected only in cells. Furthermore, the results obtained suggest that conversion of Gln1 in pyroGlu1 occurs before or during processing into the hydroxylated or non-hydroxylated LHRH intermediate (11-13)-precursors. The biosynthetic pathway is thus common for both decapeptides and it is not altered by the hydroxylation process. LHRH and [Hyp9]LHRH shared the same receptor for their biological activity, as assessed by measuring luteinizing hormone release and activation of phospholipase C and A2. [Hyp9]LHRH was, however, less potent than LHRH.

Choline acetyltransferase and somatostatin levels in aged Microcebus murinus brain.

beta-Amyloid protein (beta-AP) deposits, analoguous to those found in Alzheimer's disease (AD) are observed in the brain of aging Microcebus murinus. Because choline acetyltransferase (ChAT) activity and somatostatin (SRIH) content are consistently decreased in AD, we tested whether such changes could be observed in middle aged to aged Microcebus cerebral cortex and whether they were accompanied by beta-AP deposits. A positive correlation was observed between age and ChAT activity. By HPLC, SRIH immunoreactivity eluted as four peaks, two of which being identical with SRIH-28 and SRIH-14 while the other two likely represented precursor forms. Cortical SRIH content was not significantly affected by age. ChAT activity and SRIH content were not significantly correlated. Amyloid angiopathy was observed in every brain examined and the presence of cortical lesions analoguous to senile plaques observed in the oldest case only which did not demonstrate important alterations in ChAT and somatostatin levels.

Preferential distribution of C-terminal fragments of [hydroxyproline9]LHRH in the rat hippocampus and olfactory bulb.

Several molecular forms related to the decapeptide LHRH were characterized and quantified in various brain structures of intact and castrated male and female rats. Distinct moieties were separated by high performance liquid chromatography (HPLC) and radioimmunoassayed against anti-LHRH antibodies of different specificities. The hypothalamus contained the highest concentration of LHRH-like material detected by the antisera. The predominant (89%) molecular form recovered from that structure was LHRH itself; 9% of the material corresponded to [hydroxyproline9]LHRH ([Hyp9]LHRH), an endogenous posttranslational product of the LHRH precursor, and the residual immunoreactivity was accounted for by C-terminal fragments of both decapeptides, as assessed after labelling HPLC columns with appropriate synthetic or endogenous hypothalamic peptides. The proportions were the same in both sexes and were not affected by castration, in spite of a lesser overall LHRH activity in females and in castrates. LHRH and [Hyp9]LHRH were also detected in the olfactory bulb and the hippocampus. In these structures however, most (97%) LHRH-related molecules corresponded to C-fragments derived from [Hyp9]LHRH, whereas only very few fragments derived from the nonhydroxylated decapeptide were found. Sex or castration affected neither total nor relative concentrations of LHRH-derived molecules in the olfactory bulb and the hippocampus. Taken altogether, these observations are suggestive of a different LHRH metabolic regulation in neurons projecting to either the median eminence or extrahypothalamic areas. In the latter case, larger amounts of the LHRH precursor appear processed to [Hyp9]LHRH. Recovery of relatively high concentrations of [Hyp9]LHRH C-fragments in the olfactory bulb and the hippocampus reflects the higher resistance of the Hyp9-Gly10-NH2 than the Pro9-Gly10-NH2 peptide bond to hydrolysis by the postproline cleaving enzyme. In view of reports that intracerebral administration of C-terminal fragments of LHRH are able to trigger sex behavior, our finding that extrahypothalamic structures contain relatively high concentrations of the [Hyp9]LHRH-derived, more stable C-fragments suggests that these catabolites may have a role in the regulation of sex behavior.

Precursor and deaminated forms of both luteinizing hormone-releasing hormone (LHRH) and (hydroxyproline9)LHRH are present in the rat hypothalamus.

A naturally occurring analog of the decapeptide luteinizing hormone-releasing hormone ([Hyp9]LHRH) has been described previously in the hypothalamus of several mammals. It derives from post-translational hydroxylation of the LHRH proline9 residue. In the present work, intermediate LHRH precursors exhibiting both Pro9 or Hyp9 residues in the LHRH sequence were characterized in the rat hypothalamus. Hydroxylation of the Pro9 residue can thus be assumed to occur at an early stage of post-translational maturation. Deaminated, free acid forms of both native decapeptides were also detected. They correspond most likely to catabolites from incompletely processed precursors.

A second endogenous molecular form of mammalian hypothalamic luteinizing hormone-releasing hormone (LHRH), (hydroxyproline9)LHRH, releases luteinizing hormone and follicle-stimulating hormone in vitro and in vivo.

In vitro and in vivo release of pituitary hormones were studied in the presence of (hydroxyproline9)LHRH ((Hyp)LHRH), a newly characterized endogenous molecular form of LHRH. Results were compared to those obtained with LHRH itself. (Hyp)LHRH, as LHRH, stimulated both luteinizing hormone (LH) and follicle-stimulating hormone (FSH) release in a homothetic manner. The hydroxylated compound was, however, 24 times (in vitro) and 5 times (in vivo) less potent than LHRH. The lower activity of (Hyp)LHRH than of LHRH in the in vitro assay correlated well with a 28-fold lesser potency in a binding test using pituitary membrane preparations. The higher relative potency and the prolonged effect of (Hyp)LHRH in the in vivo test were related to a lesser susceptibility of the hydroxylated form to proteolytic degradation. Effects of LHRH and of (Hyp)LHRH were not additive, both peptides were equally able to desensitize gonadotrophs to a subsequent challenge by the other. Taken together, these observations suggest that both forms of LHRH act at the same receptor site. The lesser affinity of the hydroxylated compound is compensated to a certain extent by its higher resistance to enzymatic degradation. It is concluded that in spite of its lesser potency, (Hyp)LHRH may participate in the regulation of gonadotropins.

Luteinizing hormone-releasing hormone (LHRH)- and (hydroxyproline9)LHRH-stimulated human chorionic gonadotropin secretion from perifused first trimester placental cells.

(Hydroxyproline9)LHRH [(Hyp9)LHRH] has been isolated from human, sheep, rodent, and frog hypothalamus. (Hyp9)LHRH is the major LHRH moiety in fetal rat hypothalamus. This study compared 1) synthetic LHRH-stimulated hCG secretion from term trophoblast vs. first trimester placental cells and 2) the ability and specificity with which synthetic LHRH and (Hyp9)LHRH could stimulate hCG secretion from 8- to 12-week gestation placenta. Physically dissociated cells from multiple placentae were pooled, plated on microcarrier beads, and perifused in 1.5-ml chambers (1.5 x 10(6) cells/chamber). Effluent fractions were analyzed for hCG. Each chamber was its own control. Basal hCG secretion did not depend upon exogenous LHRH stimulation. The amplitude of LHRH-stimulated hCG pulses was greater from first trimester placental than term trophoblast cells (mean +/- SEM, 6.99 +/- 1.47 vs. 0.50 +/- 0.05 mIU/ml perifusate; peak minus basal; n = 4 chambers; P less than 0.01). LHRH and (Hyp9)LHRH (10(-9) M) increased hCG secretion from first trimester placental cells (5.02 +/- 1.29 vs. 8.64 +/- 1.61 and 4.36 +/- 0.58 vs. 7.44 +/- 1.01 mIU/ml; n = 15 and 9, respectively; P less than 0.01). At the concentrations used, LHRH and (Hyp9)LHRH seemed to stimulate hCG secretion equipotently (P greater than 0.05). Simultaneous perifusion with an LHRH antagonist, (Nal-Glu)LHRH blocked the hCG secretory response to LHRH or (Hyp9)LHRH (equimolar 10(-9) M concentrations; n = 5; P less than 0.05). (Nal-Glu)LHRH alone (10(-9) M) did not affect hCG secretion (n = 5; P greater than 0.05). The results suggested that first trimester placental cells are more responsive to LHRH than are term trophoblast cells. (Hyp9)LHRH is a potential physiological secretagogue of hCG.

[New data on a second endogenous molecular form of mammalian hypothalamic LHRH, (hydroxyproline 9) LHRH]. / Nouvelles données sur une seconde forme moléculaire endogène de la LHRH hypothalamique de mammifère, la (hydroxyproline9) LHRH.

An endogenous hydroxylated form of LHRH, (Hyp) LHRH, is able to displace LHRH bound to pituitary membrane preparations. In parallel, it stimulates release of both LH and FSH from pituitary cells in primary culture. The potency ratio of (Hyp)LHRH is approximately 1:20 and 1:5 with respect to the native decapeptide when peptidasic degradation is or is not inhibited. This correlates with a greater resistance of (Hyp) LHRH towards enzymatic degradation; in contrast to LHRH, the C-terminal (residues 6 to 10) end of (Hyp) LHRH is not degraded and generates C-terminal fragments which account for 64% of the LHRH immunoreactivity in extrahypothalamic areas as the hippocampus. Besides its weak gonadotropin releasing activity and its action or its localization in peripheral organs (placenta, gonads), a major role of the hydroxylated decapeptide may thus be to serve as a precursor of smaller active fragments on targets other than pituitary receptors.

Seasonal changes in thyroid-gonadal interactions in the edible dormouse, Glis glis.

This study was conducted to determine changes in thyroid-gonadal interaction in the edible dormouse during the phase of the annual cycle that corresponds to the end of the breeding season (from June to September). We evaluated intra-hypothalamic luteinizing hormone-releasing hormone (LHRH) content, and plasma concentrations of luteinizing hormone (LH), testosterone, thyroid-stimulating hormone (TSH) and thyroxine (T4) in three groups of dormice: (1) controls; (2) dormice receiving sufficient T4 supplementation to maintain June levels in control animals until September, thus counteracting the seasonal reduction of T4 that normally begins in July; and (3) thyroidectomized dormice. The effect of thyroidectomy was only detectable in June, when plasma T4 concentration in the control group was maximal, and consisted of a significant decrease in plasma testosterone levels. This provides strong support for the hypothesis that thyroid function positively influences gondal function during the breeding season. The T4 supplementation resulted in a decrease in hypothalamic LHRH concentration, suggesting that an increased LHRH release led to the observed stimulated hypophyseal secretion of LH in June and September and the increased circulating testosterone levels in September. There was no detectable effect in July and August. These results show that thyroid axis activation of the hypothalamic-pituitary-gondal system is only possible during certain phases of the annual cycle, particularly evidenced here during the breeding season. They also reinforce our conclusions drawn from the thyroidectomy results. Conversely, the summer testicular regression which normally occurs after the breeding season is no longer controlled by plasma T4 levels.(ABSTRACT TRUNCATED AT 250 WORDS)

(Hydroxyproline(9)) luteinizing hormone-releasing hormone: A novel peptide in mammalian and frog hypothalamus.

Luteinizing hormone-releasing hormone (LHRH)-like immunoreactive material was detected in mammalian and frog hypothalamic extracts after high performance liquid chromatography. Radioimmunological detection was performed with three different antibodies directed respectively against the N-(N-antibody) or the C-terminus (C-antibody) as well as the conformational structure (W-antibody) of synthetic mammalian LHRH (mLHRH). Under the chromatographic conditions used, two immunoreactive peptides could be separated. One of those was detected in a homothetic manner by the three antibodies. It coeluted with synthetic mLHRH, had the same amino acid composition as mLHRH, and was shown to correspond to authentic endogenous LHRH. The other peptide was more efficiently detected by the N- than by the two other antibodies. It was eluted prior to mLHRH on reverse phase chromatography but did not correspond to LHRH fragments or to any other known LHRH sequences. Its molecular weight was 1198 Da. Amino acid analysis and enzymatic digestion indicated a very close LHRH sequence homology, with a substitution of proline (Pro) by a hydroxyproline (Hyp) residue in position 9. The new peptide was found in extracts of human, rodent, ovine and amphibian hypothalamus, with a markedly higher concentration in fetal than in adult rats. Various extraction and chromatographic control procedures led to the conclusion that the new peptide was genuinely endogenous and did not result from artifactual modification(s) of LHRH during the purification process. It is concluded that, in the mammalian as in the frog hypothalamus, LHRH-like material corresponds to at least two native molecular forms: (Pro(9)) and (Hyp(9))LHRH.

The helminth Schistosoma mansoni expresses a peptide similar to human beta-endorphin and possesses a proopiomelanocortin-related gene.

Opioid peptides, a group of transmitter substances with a high degree of phylogenic conservation, have many different functions, including a role in modulation of cells of the immune system. We have postulated the existence of such peptides in the parasite Schistosoma mansoni in view of their possible role in host-parasite interactions. In this report we show that beta-endorphin, which is a member of the opiate family and is derived from the proopiomelanocortin (POMC) precursor, is present in S. mansoni. Southern blots of cercarial genomic DNA, hybridized with two oligonucleotide probes complementary to highly conserved POMC sequences, showed a POMC-related gene in this trematode. Northern blot analysis of adult worm RNA indicated that this gene was actively transcribed. Significant amounts of beta-endorphin, adrenocorticotropin (ACTH), and alpha-melanocyte stimulating hormone (alpha-MSH) were detected in all developmental stages of the parasite by radioimmunoassays with the use of antisera to human peptides. By means of reverse-phase high-performance liquid chromatography (HPLC), we found that the parasite beta-endorphin-like material and the human opiate have a high degree of homology. These results appear to constitute the first demonstration of a POMC-related gene transcribed in an invertebrate.

[Neurohormones coming from the normal and tumoral human anterior pituitary. Secretion and regulation in vitro]. / Neurohormones provenant de l'antéhypophyse humaine normale et tumorale. Sécrétion et régulation in vitro.

Several neuropeptides, classically associated with the hypothalamus have been found in the anterior pituitary and their local synthesis has been hypothesized. Using normal and tumoral human pituitaries we found in the tissue itself different neuropeptides (TRH, SRIH, GHRH) and dopamine in variable quantities according to the nature of the tissue. They were all present in normal pituitaries while only stimulatory neurohormones like TRH and GHRH were found in tumoral tissue implying an imbalance between the stimulatory and inhibitory control of hypophyseal hormones (PRL and GH) in pituitary adenomas. Fragments from normal pituitaries and dispersed cells from GH, PRL and nonsecreting adenomas, were perifused for 4 hours in a Krebs-Ringer medium collected every 2 min and GH, PRL, TRH, GHRH and SRIH were measured by RIA under basal conditions and in the presence of 10(-6) mol/L DA, TRH or SRIH. Neuropeptides and DA were characterized by HPLC. Both normal and tumoral pituitaries released TRH, SRIH and GHRH in large amounts suggesting their local synthesis. There was an in situ regulation between SRIH and GH as their secretion profile was negatively correlated, GH secretion decreasing while SRIH secretion was increasing. Moreover the release of TRH was stimulated 5 to 20 folds by DA, while PRL decreased at the same time. Pulses of TRH and SRIH had differential effects on GHRH and SRIH release according to the nature of the tissue as TRH stimulated SRIH release from normal pituitary while it inhibited SRIH release from adenoma. These results indicate that anterior pituitary cells can release neuropeptides which are probably endogenously synthesized and have a local regulation.(ABSTRACT TRUNCATED AT 250 WORDS)

Normal and growth hormone (GH)-secreting adenomatous human pituitaries release somatostatin and GH-releasing hormone.

Neuropeptides such as vasoactive intestinal peptide, LHRH, or TRH have been found in rat pituitary tissue and could act via paracrine or autocrine actions in this tissue. In this study we investigated whether normal human pituitary tissue and GH-secreting human pituitary adenomas could release somatostatin (SRIH) and GHRH. Fragments from three human pituitaries and dispersed cells from six GH-secreting adenomas (four adenomas were studied for GHRH release and five for SRIH release) were perifused using a Krebs-Ringer culture medium, and the perifusion medium was collected every 2 min (1 mL/fraction for 5 h). GH, GHRH, and SRIH were measured by RIA under basal conditions and in the presence of 10(-6) mol/L TRH or SRIH. Both normal pituitaries and GH-secreting pituitary adenomas released SRIH and GHRH. SRIH release commenced 90-180 min after initiation of the perifusion, at which time GH secretion had decreased significantly. TRH stimulated SRIH release from normal pituitary tissue and inhibited SRIH release from adenoma tissue. GHRH was present at the start of the perifusion, but rapidly disappeared. However, SRIH stimulated GHRH release from normal pituitary tissue, but not from adenoma tissue. Significant amounts of GHRH and SRIH were released during the experiments, suggesting their local synthesis. These results indicate that pituitary cells can release hypothalamic peptides. The liberation of these neuropeptides is regulated, and moreover, their regulation differs between normal and adenomatous pituitaries.

LHRH-like immunoreactivity in the human placenta is not identical to LHRH.

LHRH-like immunoreactive material was separated from human placental homogenates by chromatography and tested against two antibodies directed respectively against the C- or the N-terminus (N- and C-antibodies) of the synthetic peptide. Extraction yield was 94 per cent as assessed by the recovery of the radiolabelled LHRH added to the homogenates. Placental LHRH-like immunoreactivity in crude extracts appeared largely overestimated, due to interference of endogenous substances non-related to LHRH. After elimination of this contamination, a higher residual activity was detected with the C- than with the N-antibodies. Eluates tested under these conditions contained only 550 pg of LHRH-like material per kg of placenta. This concentration was comparable whether placental tissue had been sampled at 12 weeks of pregnancy or at the end of gestation. The corresponding peptides however neither co-eluted with synthetic LHRH nor with catabolite fragments of the peptide. It is concluded that the human placenta contains a lesser amount of LHRH-like material than anticipated on the basis of earlier results and that this material does not correspond to the native decapeptide.

[In vitro secretion of somatostatin (SRIH) by human adenomatous somatotropic cells. Relation with somatotropic hormone (GH) release and modulation by thyroliberin (TRH)]. / Sécrétion in vitro de somatostatine (SRIH) par les cellules somatotropes adénomateuses humaines. Relation avec la libération d'hormone somatotrope (GH) et modulation par la thyrolibérine (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.

Effects of estradiol and progesterone on immunoreactive forms of hypothalamic luteinizing hormone-releasing hormone.

In order to investigate mechanisms underlying the ovarian steroid action on hypothalamic luteinizing hormone-releasing hormone (LHRH) neurons, LHRH and a higher immunoreactive molecular form (MW 1,800 daltons) of the decapeptide were immunoassayed with antibodies of different specificities in hypothalamic subcellular fractions, after molecular sieve filtration on Biogel P4 columns equilibrated with 0.2 N acetic acid containing 0.02% sodium azide. The study was performed in ovariectomized (OVX), ovariectomized estradiol-implanted (OVX + E2) or OVX + E2 progesterone one-treated rats (OVX + E2 + P). The animals were killed before or during the circadian luteinizing hormone (LH) surge. The amount of LHRH-like immunoreactivity recovered from the synaptosomal fraction was slightly increased in OVX + E2-implanted animals but very markedly augmented in OVX + E2 + P-treated rats. In contrast, the higher molecular form recovered from a high-speed supernatant was markedly decreased in OVX + E2 + P-treated rats when compared to the other groups. At the time of maximal LH release induced by E2 + P administration, hypothalamic LHRH was markedly depleted, whereas the larger molecular form was notably augmented. The data suggest that ovarian steroids not only influence release of hypothalamic LHRH but also the processing of LHRH precursor forms.