Fasting Enhances Pyroglutamyl Peptidase II Activity in Tanycytes of the Mediobasal Hypothalamus of Male Adult Rats.

Fasting down-regulates the hypothalamus-pituitary-thyroid (HPT) axis activity through a reduction of TRH synthesis in neurons of the parvocellular paraventricular nucleus of the hypothalamus (PVN). These TRH neurons project to the median eminence (ME), where TRH terminals are close to the cytoplasmic extensions of ß2 tanycytes. Tanycytes express pyroglutamyl peptidase II (PPII), the TRH-degrading ectoenzyme that controls the amount of TRH that reaches the anterior pituitary. We tested the hypothesis that regulation of ME PPII activity is another mechanism by which fasting affects the activity of the HPT axis. Semiquantitative in situ hybridization histochemistry data indicated that PPII and deiodinase 2 mRNA levels increased in tanycytes after 48 hours of fasting. This increase was transitory, followed by an increase of PPII activity in the ME, and a partial reversion of the reduction in PVN pro-TRH mRNA levels and the number of TRH neurons detected by immunohistochemistry. In fed animals, adrenalectomy and corticosterone treatment did not change ME PPII activity 72 hours later. Methimazole-induced hypothyroidism produced a profound drop in tanycytes PPII mRNA levels, which was reverted by 3 days of treatment with T4. The activity of thyroliberinase, the serum isoform of PPII, was increased at most fasting time points studied. We conclude that delayed increases in both the ME PPII as well as the thyroliberinase activities in fasted male rats may facilitate the maintenance of the deep down-regulation of the HPT axis function, despite a partial reactivation of TRH expression in the PVN.

TRH and TRH-like peptide expression in rat following episodic or continuous corticosterone.

Sustained abnormalities of glucocorticoid levels have been associated with neuropsychiatric illnesses such as major depression, posttraumatic stress disorder (PTSD), panic disorder, and obsessive compulsive disorder. The pathophysiological effects of glucocorticoids may depend not only on the amount of glucocorticoid exposure but also on its temporal pattern, since it is well established that hormone receptors are down-regulated by continuously elevated cognate hormones. We have previously reported that TRH (pGlu-His-Pro-NH2) and TRH-like peptides (pGlu-X-Pro-NH2) have endogenous antidepressant-like properties and mediate or modulate the acute effects of a single i.p. injection of high dose corticosterone (CORT) in rats. For these reasons, two accepted methods for inducing chronic hyperglucocorticoidemia have been compared for their effects on brain and peripheral tissue levels of TRH and TRH-like peptides in male, 250 g, Sprague-Dawley rats: (1) the dosing effect of CORT hemisuccinate in drinking water, and (2) s.c. slow-release pellets. Overall, there were 93% more significant changes in TRH and TRH-like peptide levels in brain and 111% more in peripheral tissues of those rats ingesting various doses of CORT in drinking water compared to those with 1-3 s.c. pellets. We conclude that providing rats with CORT in drinking water is a convenient model for the pathophysiological effects of hyperglucocorticoidemia in rodents.

Effects of the dopamine agonist cabergoline on the pulsatile and TRH-induced secretion of prolactin, LH, and testosterone in male beagle dogs.

In the present study, the pulsatile serum profiles of prolactin, LH and testosterone were investigated in eight clinically healthy fertile male beagles of one to six years of age. Serum hormone concentrations were determined in blood samples collected at 15 min intervals over a period of 6 h before (control) and six days before the end of a four weeks treatment with the dopamine agonist cabergoline (5 microg kg(-1) bodyweight/day). In addition, the effect of cabergoline administration was investigated on thyrotropin-releasing hormone (TRH)-induced changes in the serum concentrations of these hormones. In all eight dogs, the serum prolactin concentrations (mean 3.0 +/- 0.3 ng ml(-1)) were on a relatively constant level not showing any pulsatility, while the secretion patterns of LH and testosterone were characterised by several hormone pulses. Cabergoline administration caused a minor but significant reduction of the mean prolactin concentration (2.9 +/- 0.2 ng ml(-1), p < 0.05) and did not affect the secretion of LH (mean 4.6 +/- 1.3 ng ml(-1) versus 4.4 +/- 1.7 ng ml(-1)) or testosterone (2.5 +/- 0.9 ng ml(-1) versus 2.4 +/- 1.2 ng ml(-1)). Under control conditions, a significant prolactin release was induced by intravenous TRH administration (before TRH: 3.8 +/- 0.9 ng ml(-1), 20 min after TRH: 9.1 +/- 5.9 ng ml(-1)) demonstrating the role of TRH as potent prolactin releasing factor. This prolactin increase was almost completely suppressed under cabergoline medication (before TRH: 3.0 +/- 0.2 ng ml(-1), 20 min after TRH: 3.3 +/- 0.5 ng ml(-1)). The concentrations of LH and testosterone were not affected by TRH administration. The results of these studies suggest that dopamine agonists mainly affect suprabasal secretion of prolactin in the dog.

Effects of PRI-2191--a low-calcemic analog of 1,25-dihydroxyvitamin D3 on the seizure-induced changes in brain gene expression and immune system activity in the rat.

Apart from the essential role of 1alpha,25-dihydroxyvitamin D3 in calcium and phosphorus metabolism, this compound and its analogs are involved in regulating the functions of the central nervous and immune systems. Active forms of vitamin D3 have been reported to stimulate neurotrophin gene expression and to prevent neuronal damage against a variety of insults. In the present study, we evaluated the effects of PRI-2191-a low-calcemic analog of 1alpha,25-dihydroxyvitamin D3 (50 ng/kg i.p., once daily for 8 days) on seizure-related neuronal degeneration, changes in some brain gene expression and immune system activity. Seizures were induced by pilocarpine (400 mg/kg; i.p.) administration. An in situ hybridization study showed that the pilocarpine-induced seizures led to time-dependent changes in the brain-derived neurotrophic factor (BDNF), heat shock protein 70 (HSP-70) and prepro-thyreoliberin (prepro-TRH) gene expression in several cortical and hippocampal regions. The maximal induction of gene expression was 3 h for BDNF and 24 h for HSP-70 and prepro-TRH; however, only in the case of prepro-TRH that effect was long-lasting. PRI-2191 alone had no effect on gene expression, but it enhanced the seizure-evoked expression of HSP-70, had an opposite effect on BDNF mRNA level and did not affect prepro-TRH mRNA level. Moreover, PRI-2191 had a moderate inhibitory effect on the seizure-related hippocampal damage in the CA1 field only. An immunological study revealed that PRI-2191 reversed the seizure-induced decrease in the proliferative activity of splenocytes and their ability to produce interferon-gamma. Summing up, the present study demonstrated that subchronic administration of PRI-2191 significantly modulated the seizure-related changes in both the brain and the peripheral immune system of rats.

Thyroid hypofunction in patients with rapid-cycling bipolar disorder after lithium challenge.

BACKGROUND: There is debate whether patients with rapid-cycling bipolar disorder (BD) are predisposed to thyroid axis abnormalities and whether this may contribute to development of rapid mood shifts. Using lithium carbonate as a challenge to the hypothalamic-pituitary-thyroid (HPT) system, we determined whether patients with rapid-cycling BD are sensitive to the "antithyroid" properties of lithium. METHODS: We studied the response to thyrotropin-releasing hormone (TRH) of HPT system hormones in 20 medication-free patients with rapid-cycling BD and compared these measurements with those of 20 healthy age- and gender-matched control subjects. The same measurements were repeated after both groups had received lithium carbonate for 4 weeks in sufficient doses to maintain blood levels between.7-1.2 mEq/L. RESULTS: At baseline, the results of thyroid function tests, including the TRH challenge test, did not differ between patients and control subjects. After treatment with lithium, serum concentrations of thyroxine significantly decreased, whereas basal thyrotropin (TSH) and DeltaTSH(max) significantly increased in both patients and control subjects; however, patients had significantly higher DeltaTSH(max) after TRH stimulation. More patients than control subjects developed laboratory evidence consistent with grade III hypothyroidism after lithium treatment. CONCLUSIONS: Rapid-cycling BD is associated with a latent hypofunction of the HPT system. This dysfunction becomes manifest with short-term lithium challenge.

Disturbances in hypothalamo pituitary adrenal and thyroid axis identify different sleep EEG patterns in major depressed patients.

This study was aimed at investigating the relationships between sleep EEG abnormalities and hypothalamo pituitary adrenal (HPA) and hypothalamo pituitary thyroid (HPT) disturbances in major depressive disorder. Post dexamethasone (DXM) cortisol levels and the dual TSH response to 08:00 h and 23:00 h TRH administration were determined after a 2 weeks wash-out period in a group of 113 DSM-IV major depressed patients (72 females aged 44.3+/-13.0 and 41 males aged 45.7+/-11) who were consecutively admitted to undergo sleep EEG recordings. Post-DXM cortisolemia, 08:00 and 23:00 post-TRH TSH values, time spent in rapid eye movement sleep (REMS), in slow wave sleep (SWS), and in stage 2 as well as time awake after sleep onset were introduced in a principal component (PC) analysis. The four 3 PC scores explaining up to 74% of the data set were further calculated for each patients and used in a cluster analysis. A three-cluster solution was retained. Controlling for the effects of age and gender, patients belonging to these three clusters could clearly be differentiated on the basis of their neuroendocrine responses and on their sleep EEG profiles. Compared to the two other clusters, cluster I (n=26) patients showed the most severe sleep continuity disturbances. Post-DXM cortisol escape and sleep architecture disturbances (consisting of a shortening of REMS latency and a decreased SWS) identified patients belonging to cluster II (n=39). Patients in cluster III (n=48) had the lowest TSH response to TRH and the less marked sleep EEG alteration. Clinical or demographic variables were unable to differentiate the three clusters. Our results suggest that different biological dysfunctions could each underlie particular neuroendocrine and sleep EEG disturbances in major depression.

[Aminoglycoside antibiotics neomycin and kanamycin inhibit the increase of of pyroglutamyl aminopeptidase activity by depolarizing synaptosomes of the frontal cortex of the rate]. / Los antibióticos aminoglucósidos neomicina y kanamicina inhiben el incremento de actividad piroglutamato aminopeptidasa obtenido tras despolarización de sinaptosomas de corteza frontal de rata.

INTRODUCTION: Thyrotrophin releasing hormone (TRH) has emerging in the last few years as a neuropeptide with important functions, not only as neurohormone into the hypothalamus-pituitary axis, but as neurotransmitter in several areas of the nervous system. Although little is known about its extra-endocrine functions, TRH has been related with several types of psychiatric disorders. Pyroglutamyl aminopeptidase (pGluAP) is the enzyme involved in the degradation of TRH. OBJECTIVES: The present research studies the levels of pGluAP activity under basal (resting) and KCl-stimulated (depolarized) conditions. The role of intracellular free calcium homeostasis, by means of the aminoglycoside antibiotics neomycin and kanamycin as voltage-dependent calcium channels blockers, is also studied. MATERIAL AND METHODS: Both pGluAP activity and intracellular free calcium concentration were analyzed in synaptosomes obtained from the frontal cortex of rats. Synaptosomes were incubated in artificial cerebrospinal fluid, under basal (resting) or KCl-stimulated (depolarized) conditions, with of without neomycin or kanamycin at different concentrations. RESULTS: Depolarization increases significantly pGluAP activity, which is completely abolished by neomycin and kanamycin at the lower concentrations used. On the contrary, aminoglycoside antibiotics do not block completely the increase on intracellular free calcium concentration induced by depolarization. Under basal conditions, no changes were found on pGluAP activity nor intracellular free calcium. CONCLUSIONS: pGluAP activity could regulate the neurotransmitter/neuromodulatory functions of TRH trough intracellular free calcium movements through aminoglycoside-sensitive voltage-dependent calcium channels. A role for inositol 4,5-bisphosphate breakdown products is also suggested.

Evaluation of thyroid function in lithium-naive bipolar patients.

A high prevalence of thyroid hypofunction has been found in bipolar patients. However, the samples used in previous studies included a high percentage of patients in treatment with lithium and carbamazepine. Since the use of these drugs may explain the high prevalence of thyroid disturbances found in bipolar patients, we designed the present study to assess thyroid function in a sample of bipolar patients who had not been treated previously with lithium or carbamazepine. Patients included in the sample met Research Diagnostic Criteria for bipolar affective disorder. Assessment included determination of serum levels for total tyroxine (T4), total triiodothyronine (T3), and thyrotropin both basally and in response to infusion of 500 mg of Protilerin. The rate of thyroid hypofunction in the total sample (9.2%) was considerably lower than that reported in other studies with bipolar patients undergoing lithium therapy. Five patients (9.2%) showed some thyroid hyperfunction parameter. Our results do not show significant differences in thyroid function indices between long-term and short-term duration of illness, between outpatients and inpatients, between high and low number of episodes, and between rapid- and non-rapid-cycling cases. Comparison between bipolar I and bipolar II patients shows a statistically significant difference in the values of TSH levels, with the bipolar II group having a higher mean value. Our data suggest that thyroid dysfunction is not related to gender, duration of illness, number of episodes, or rapid-cycling course of illness. The higher TRH-stimulated TSH levels in the bipolar II group could be considered a differential biological feature.

Endocrine mechanism of action of toremifene at the level of the central nervous system in advanced breast cancer patients.

PURPOSE: To differentiate the antagonistic and agonistic effect of toremifene at the level of the hypothalamus-hypophysis axis a leutinizing hormone-releasing hormone (LHRH) test was performed during a phase II clinical trial. METHODS: In 15 postmenopausal patients with advanced breast cancer, follicle-stimulating hormone (FSH) and LH release--induced by an LHRH agonist (Suprefact injection, 0.5 mg s.c.)--was monitored during a 16-week period of toremifene treatment (60 mg/day p.o.). Prolactin, estradiol, and sex hormone-binding globulin (SHBG) levels were also measured. The functional test was carried out prior to toremifene therapy and then 4, 8, 12, and 16 weeks afterward. RESULTS: The drug sensitized the pituitary to the action of the gonadotrophins; the LHRH-induced FSH and LH release showed a considerably increasing tendency during the toremifene therapy. Estradiol levels decreased statistically significantly and SHBG levels showed a statistically significant increase. A decreased level of prolactin is the sign of an antiestrogenic effect of toremifene on the hypophysis and, as a result, provides evidence for a direct influence of toremifene upon the pituitary. An increase in LH and prolactin release in response to the LHRH test was characteristic in the responders. CONCLUSION: According to the LHRH test, the antagonistic effect of toremifene seems to be more dominant than the concomitantly existing agonistic property. Neither clinical nor endocrinological side effects could be observed at the level of the CNS during a prolonged period of toremifene administration.

Regulation of the human TRH (hTRH) gene by human thyroid hormone receptor beta 1 (hTR beta 1) mutants.

TRH is negatively regulated by T3 both in the hypothalamic paraventricular nucleus and transient transfection models. Mutations in hTR beta 1 genes are associated with the syndrome of generalized resistance to thyroid hormone. To investigate potential effects of mutant TRs on T3 regulation of the hTRH gene, transient gene expression assays were performed in human neuroblastoma (HTB-11) cells with an hTRH promoter-luciferase construct, wild type (WT) hTR beta 1, and three qualitatively distinct hTR beta 1 mutant forms (ED, OK and PV). In the presence of T3 (10(-9) M), liganded WT-hTR beta 1 inhibited hTRH promoter activity significantly (40%). Cotransfection of each of the two mutants (ED and OK) achieved similar levels of inhibition only at 10 to 100 fold increased T3 concentrations. Of interest, a 10x excess of mutant ED or OK could also exert dominant negative effects upon WT hTR beta 1-T3 mediated inhibitory actions on the hTRH promoter. In contrast, mutant TR-PV exerted neither inhibitory nor dominant negative effects at even higher concentrations of T3. Moreover, all three unliganded mutant forms stimulated TRH promoter activity significantly in the absence of T3, despite their different mutations in the ligand-binding domain (LBD). These data demonstrate that thyroid hormone resistance at the level of TRH gene regulation, due to reduced inhibitory actions of mutant TR-T3 complexes, as well as dominant negative effects upon WT hTR beta 1 mediated inhibition, likely contribute to elevated TSH values observed in the syndrome of thyroid hormone resistance.

Preprothyrotropin-releasing hormone mRNA in the rat central gray is strongly and persistently induced during morphine withdrawal.

In vivo evidence strongly implicates the central gray in expression of the physical symptoms of opiate withdrawal. Preprothyrotropin-releasing hormone (ppTRH) mRNA is highly expressed in the central gray. Furthermore, systemic administration of thyrotropin-releasing hormone (TRH) inhibits the development of opiate dependence in rats. To elucidate the link between TRH and opiate withdrawal, we examined the regulation of ppTRH mRNA in the central gray of rats made dependent on morphine, and during opiate withdrawal, using quantitative in situ hybridization. In the ventrolateral central gray, a significant increase in ppTRH mRNA was observed 3 h after precipitation of withdrawal, and this increase persisted for 36 h. Upregulation of ppTRH mRNA was not seen with chronic morphine or acute naltrexone treatment alone and was specific for the ventrolateral central gray. These findings support a role for TRH or other ppTRH-derived peptides in the central gray during morphine withdrawal.

Facilitatory role of serotonin (5-HT) in the control of thyrotropin releasing hormone/thyrotropin (TRH/TSH) secretion in rats.

In order to investigate the role of serotonin in the regulation of thyrotropin (TSH) secretion, control and propylthiouracil (PTU)-treated male Wistar rats weighing approximately 250 g were subjected to ip injections of methysergide (MET, 10 micrograms/100 g body weight), a serotonergic receptor blocker, and killed 60 min later by decapitation. Serum and pituitary concentrations of TSH were measured by radioimmunoassay. An addition, the pituitary release of TSH was estimated in an in vitro system in which pituitary glands were incubated with hypothalamic extracts. MET treatment led to a decrease in pituitary (94.12 +/- 18.55 vs 199.30 +/- 31.47 micrograms/mg, N = 20), and serum (1.95 +/- 0.92 vs 4.26 +/- 1.40 ng/ml, N = 20) TSH concentration (P < 0.001) and also to a decreased in vitro pituitary response to control hypothalamic extracts (55 +/- 8 vs 78 +/- 7%, N = 5, P < 0.005). In addition, hypothalamic extracts of MET-treated rats significantly facilitated in vitro pituitary TSH secretion, suggesting an enhanced hypothalamic thyrotropin releasing hormone (TRH) activity (347 +/- 62 vs 78 +/- 7%, N = 5, P < 0.001). These results suggest that serotonin participates in the physiological control of TRH/TSH secretion, probably by increasing TRH production/secretion, and/or by facilitating the pituitary TSH response to TRH.

Facilitatory role of serotonin (5-HT) in the control of thyrotropin releasing hormone/thyrotropin (TRH/TSH) secretion in rats

In order to investigate the role of serotonin in the regulation of thyrotropin (TSH) secretion, control and propylthiouracil (PTU)treated male Wistar rats weighing approximately 250 g were subjected to ip injections of methysergide (MET, 10 mug/l00 g body weight), a serotonergic receptor blocker, and killed 60 min later by decapitation. Serum and pituitary concentrations of TSH were measured by radioimmunoassay. In addition, the pituitary release of TSH was estimated in an in vitro system in which pituitary glands were incubated with hypothalamic extracts. MET treatment led to a decrease in pituitary (94.12 ñ 18.55 vs 199.30 ñ 31.47 mug/mg, N = 20), and serum (l.95 ñ 0.92 vs 4.26 ñ 1.40 ng/ml, N = 20) TSH concentration (P

Phorbol ester or cAMP enhance thyrotropin-releasing hormone mRNA in primary cultures of hypothalamic cells.

Thyrotropin releasing hormone (TRH) biosynthesis is subject to a multifactorial control. TRH mRNA levels are negatively regulated by thyroid hormones in the paraventricular hypothalamic nucleus, and positively in cold exposure or suckling. Effect of second messenger pathways stimulation, a known response to membrane receptors, was studied in vitro; cultures of rat embryonic hypothalami (18 day gestation) were treated with 12-O-tetradecanoylphorbol-13-acetate (TPA, 100 nM) or dibutiryl cAMP (dBcAMP, 1 mM) for various times. Levels of TRH mRNA were raised after the first hour of dBcAMP or 2 h of TPA treatment and were still increased at 24 h. These results suggest a neural regulation of TRH biosynthesis.

Effect of a novel prolyl endopeptidase inhibitor, JTP-4819, on thyrotropin-releasing hormone-like immunoreactivity in the cerebral cortex and hippocampus of aged rats.

We investigated the effects of a novel prolyl endopeptidase inhibitor, JTP-4819 ((S)-2-[[(S)-2-(hydroxyacetyl)-1-pyrrolidinyl]carbonyl]-N- (phenylmethyl)-1-pyrrolidinecarboxamide), on thyrotropin-releasing hormone (TRH)-like immunoreactivity (TRH-LI) in the cerebral cortex and hippocampus of aged rats. The TRH-LI content of both brain regions in aged rats was significantly lower than that in young rats. A single oral dose of JTP-4819 (3 mg/kg) restored the cortical TRH-LI content in aged rats, while doses of 0.3-3 mg/kg restored it in the hippocampus. Repeated oral administration of JTP-4819 at a dose of 1 mg/kg for 21 days produced a significant increase of TRH-LI in the cerebral cortex, while it did so in the hippocampus at doses of 0.3 and 1 mg/kg. Our findings suggest that JTP-4819 may improve the functioning of TRHergic neurons, which deteriorate with senescence.

Different regulation of thyrotropin releasing hormone content and release in paraventricular nucleus (PVN) and median eminence (ME) of rat hypothalamus during in vitro incubation.

TRH is present throughout the central nervous system possessing many different functions. Only TRH synthesized in the hypothalamic PVN and transported to the ME regulates anterior pituitary secretion of thyrotropin. To investigate this hypophysiotropic system, we have developed a method using dissected rat PVN and ME, representing mostly cell bodies and nerve terminals respectively, of the same TRH system. Tissues were incubated for four 30 min periods each in Locke's medium with alternatively normal and high KCl concentration. Repeated KCl-induced membrane depolarization resulted in significantly increased TRH release from both tissues (15% of TRH content) which was dependent on Ca2+ influx. Some important differences were found in the regulation of TRH in the PVN and ME: Comparison of TRH content in the tissue before and after incubation with the amount of secreted TRH showed new production of TRH during incubation in the ME, but not in the PVN. Frequent medium replacement during depolarization revealed that TRH secretion at the level of the ME, but not the PVN, is probably inhibited by some substance released during incubation. These data suggest that there is a different regulation of TRH in the PVN and ME and that TRH secreted by the isolated PVN may come mostly from the perikarya and represent paracrine neurohormone secretion.

The influence of muscarinic and prostaglandic mechanisms on regional cerebral and peripheral blood flows and on the vascular effects of thyrotropin releasing hormone (TRH).

TRH has pronounced vascular effects. The final transmitter mechanisms of these effects are not fully understood. The present study was conducted in order to elucidate whether these effects are mediated by prostaglandic or muscarinic mechanisms. Muscarinic blockade augmented the vasoconstricting- and pressor effect of TRH; vasodilation in the brain was attenuated only in the caudate nucleus. Indomethacin provoked a decrease in regional cerebral blood flow and in the gastric mucosal blood flow. No effect of indomethacin was observed on the vascular effects of TRH. It is concluded that the cerebral vasodilating and peripheral vasoconstricting effects of TRH are not mediated by prostaglandins. Muscarinic mechanisms are involved in the vasodilating effect of TRH only in the caudate nucleus.

TRH-evoked entry of extracellular calcium in GH4C1 cells: possible importance of arachidonic acid metabolites.

Previous studies have shown that stimulating pituitary GH4C1 cells with thyrotropin-releasing hormone (TRH) evoked a biphasic change in cytosolic free Ca2+ concentration ([Ca2+]i): a rapid release of sequestered Ca2+ due to the production of inositol-1,4,5-trisphosphate, and Ca2+ entry via both voltage-operated Ca2+ channels and a presently unknown voltage-independent influx pathway. The aim of the present study was to further evaluate to which extent the TRH-evoked changes in [Ca2+]i were dependent on entry of extracellular Ca2+, and which mechanisms participated in regulating this Ca2+ entry. Pretreatment of the cells with 4-bromophenylacylbromide (an inhibitor of phospholipase A2), nordihydroguaiaretic acid (an inhibitor of lipoxygenase), and econazole (an inhibitor of both lipoxygenase and cytochrome P-450 enzymes), attenuated the TRH-evoked increase in [Ca2+]i, suggesting that noncyclooxygenase metabolites of arachidonic acid or cytochrome P-450 metabolites may participate in regulating the TRH-evoked entry of extracellular Ca2+. Both nordihydroguaiaretic acid and econazole showed a similar inhibition of the Ca2+ entry, as did SKF 96365, a compound previously shown to inhibit receptor-activated Ca2+ entry. We also showed that arachidonic acid per se increased [Ca2+]i, and acidified the cytosol in GH4C1 cells in a dose-dependent manner. The effects of arachidonic acid was reversed by addition of BSA to the cell suspension. The calcium entry and the activation of the metabolism of arachidonic acid may thus be important components of the TRH-evoked signal-transduction pathway in GH4C1 cells.

TRH-immunoreactivity in chronic pancreatitis.

Thyrotropin-releasing hormone (TRH) is abundantly present in the pancreas. We studied the circulating TRH-immunoreactivity (IR) in 27 patients with chronic pancreatitis (CP) and different degrees of exocrine pancreatic insufficiency (EPI), as well as in 23 normal subjects. Furthermore we examined the effect of oral administration of 100 g glucose on peripheral TRH-IR in normal subjects (n = 5) and in patients with severe exocrine insufficiency (SEI, n = 5). Basal TRH-IR plasma levels in the CP group (20.8 +/- 7 fmol/ml, mean +/- SD) were significantly lower (p < 0.005) as compared with the normal subjects (38 +/- 14). TRH-IR plasma levels in patients with CP and SEI (15.8 +/- 3) were significantly lower (p < 0.05) than in patients with normal pancreatic function (28.1 +/- 8), but were no different from those in patients with CP and moderate exocrine insufficiency (18.7 +/- 5). In normal controls TRH-IR rose 120-180 min after glucose ingestion from 33 +/- 5 to 64 +/- 20 fmol/ml, while no increase in TRH-IR levels was observed in patients with SEI. We conclude that circulating TRH-IR levels are mainly of pancreatic origin. Patients with SEI have very low peripheral TRH-IR, indicating that CP does indeed influence TRH-release.

Effect of serotonin on the immunoreactive thyrotropin-releasing hormone concentrations of the rat stomach.

The effects of serotonin and its related compounds on immunoreactive thyrotropin-releasing hormone (ir-TRH) concentrations of the rat stomach wall and gastric juice were studied. Either serotonin, cyproheptadine or GR38032F was injected intraperitoneally, and the rats were decapitated at various times after the injection. ir-TRH concentrations of the stomach wall and gastric juice were measured by radioimmunoassay, and gastric serotonin concentrations were measured by HPLC. ir-TRH concentrations of the stomach wall decreased, and ir-TRH concentrations of gastric juice increased significantly after serotonin injection. On the other hand, both cyproheptadine and GR38032F did not affect ir-TRH concentrations of the stomach. The effects of serotonin on ir-TRH concentrations of the stomach were significantly blocked by the pretreatment of cyproheptadine and GR38032F. The reciprocal changes of gastric ir-TRH and serotonin concentrations were observed without changes in gastric juice pH. These findings suggest that serotonin stimulates ir-TRH release from the stomach wall into gastric juice, and the effects of serotonin on ir-TRH release may be partly mediated via 5-HT2- and 5-HT3-receptors.