TRH/TRH-R1 receptor signaling in the brain medulla as a pathway of vagally mediated gut responses during the cephalic phase.

Pavlov's seminal findings in the early twentieth century showed that the sight, smell or taste of food in dogs with chronic esophagostomy induces a vagal-dependent gastric acid secretion. These observations established the concept of the cephalic phase of digestion. Compelling experimental evidence in rats indicates that the three amino acid peptide thyrotropin-releasing hormone (TRH) expressed in the brainstem plays a key role in the vagal stimulation of gastric function. Neurons in the dorsal motor nucleus of the vagus (DMN) expressed TRH receptor subtype (TRH-R1) and received efferent input from TRH containing fibers arising from TRH synthesizing neurons in the raphe pallidus, raphe obscurus, and the parapyramidal regions. TRH microinjected into the DMN or intracisternally excites the firing of DMN neurons and stimulates efferent activity in the gastric branch of the vagus nerve and gastric myenteric cholinergic neurons. At the functional level, this results in a vagally-mediated and atropine-sensitive stimulation of gastric epithelial and endocrine cells secreting acid, pepsin, serotonin, histamine and ghrelin, and enteric neurons leading to increased gastric motility and emptying. Importantly, the blockade of TRH or TRH-R1 in the brainstem by pretreatment into the cisterna magna or the DMN with TRH antibody or TRH-R1 oligodeoxynucleotide antisense respectively abolishes the stimulation of gastric acid induced by sham-feeding. The gastric response to TRH injected into the DMN is potentiated by serotonin and the proTRH flanking peptide, Ps4 and suppressed by a number of brainstem peptides and cytokines activated during stress or immune response and inhibiting food intake and gastric acid secretion. These convergent data strongly support a physiological involvement of TRH signaling pathway in the brainstem to stimulate vagal activity and identified TRH-TRH-R1 system as a major effector in the dorsal vagal complex to drive the vagally mediated gut response triggered by the cephalic phase.

Thyrotropin-releasing hormone receptor type 1 (TRH-R1), not TRH-R2, primarily mediates taltirelin actions in the CNS of mice.

Thyrotropin-releasing hormone receptor type 2 (TRH-R2), not TRH-R1, has been proposed to mediate the CNS effects of TRH and its more effective analog taltirelin (TAL). Consistent with this idea, TAL exhibited higher binding affinity and signaling potency at mouse TRH-R2 than TRH-R1 in a model cell system. We used TRH-R1 knockout (R1ko), R2ko and R1/R2ko mice to determine which receptor mediates the CNS effects of TAL. There was no TRH-R1 mRNA in R1ko and R1/R2ko mice and no TRH-R2 mRNA in R2ko and R1/R2ko mice. Specific [(3)H]MeTRH binding to whole brain membranes was 5% of wild type (WT) for R1ko mice, 100% for R2ko mice and 0% for R1/R2ko mice, indicating TRH-R1 is the predominant receptor expressed in the brain. In arousal assays, TAL shortened sleep time with pentobarbital sedation in WT and R2ko mice by 44 and 49% and with ketamine/xylazine sedation by 66 and 55%, but had no effect in R1ko and R1/R2ko mice. In a tail flick assay of nociception, TAL increased response latency by 65 and 70% in WT and R2ko mice, but had no effect in R1ko and R1/R2ko mice. In a tail suspension test of depression-like behavior, TAL increased mobility time by 49 and 37% in WT and R2ko mice, but had no effect in R1ko and R1/R2ko mice. Thus, in contrast to the generally accepted view that the CNS effects of TAL are mediated by TRH-R2, these effects are mediated primarily if not exclusively by TRH-R1 in mice.

Persistent signaling by thyrotropin-releasing hormone receptors correlates with G-protein and receptor levels.

G-protein-coupled receptors with dissociable agonists for thyrotropin, parathyroid hormone, and sphingosine-1-phosphate were found to signal persistently hours after agonist withdrawal. Here we show that mouse thyrotropin-releasing hormone (TRH) receptors, subtypes 2 and 1(TRH-R2 and TRH-R1), can signal persistently in HEK-EM293 cells under appropriate conditions, but TRH-R2 exhibits higher persistent signaling activity. Both receptors couple primarily to Gα(q/11). To gain insight into the mechanism of persistent signaling, we compared proximal steps of inositolmonophosphate (IP1) signaling by TRH-Rs. Persistent signaling was not caused by slower dissociation of TRH from TRH-R2 (t(1/2)=77 ± 8.1 min) compared with TRH-R1 (t(1/2)=82 ± 12 min) and was independent of internalization, as inhibition of internalization did not affect persistent signaling (115% of control), but required continuously activated receptors, as an inverse agonist decreased persistent signaling by 60%. Gα(q/11) knockdown decreased persistent signaling by TRH-R2 by 82%, and overexpression of Gα(q/11) induced persistent signaling in cells expressing TRH-R1. Lastly, persistent signaling was induced in cells expressing high levels of TRH-R1. We suggest that persistent signaling by TRHRs is exhibited when sufficient levels of agonist/receptor/G-protein complexes are established and maintained and that TRH-R2 forms and maintains these complexes more efficiently than TRH-R1.

TRH-like peptides.

TRH-like peptides are characterized by substitution of basic amino acid histidine (related to authentic TRH) with neutral or acidic amino acid, like glutamic acid, phenylalanine, glutamine, tyrosine, leucin, valin, aspartic acid and asparagine. The presence of extrahypothalamic TRH-like peptides was reported in peripheral tissues including gastrointestinal tract, placenta, neural tissues, male reproductive system and certain endocrine tissues. Work deals with the biological function of TRH-like peptides in different parts of organisms where various mechanisms may serve for realisation of biological function of TRH-like peptides as negative feedback to the pituitary exerted by the TRH-like peptides, the role of pEEPam such as fertilization-promoting peptide, the mechanism influencing the proliferative ability of prostatic tissues, the neuroprotective and antidepressant function of TRH-like peptides in brain and the regulation of thyroid status by TRH-like peptides.

Role of helix 8 of the thyrotropin-releasing hormone receptor in phosphorylation by G protein-coupled receptor kinase.

The thyrotropin-releasing hormone (TRH) receptor undergoes rapid and extensive agonist-dependent phosphorylation attributable to G protein-coupled receptor (GPCR) kinases (GRKs), particularly GRK2. Like many GPCRs, the TRH receptor is predicted to form an amphipathic helix, helix 8, between the NPXXY motif at the cytoplasmic end of the seventh transmembrane domain and palmitoylation sites at Cys335 and Cys337. Mutation of all six lysine and arginine residues between the NPXXY and residue 340 to glutamine (6Q receptor) did not prevent the receptor from stimulating inositol phosphate turnover but almost completely prevented receptor phosphorylation in response to TRH. Phosphorylation at all sites in the cytoplasmic tail was inhibited. The phosphorylation defect was not reversed by long incubation times or high TRH concentrations. As expected for a phosphorylation-defective receptor, the 6Q-TRH receptor did not recruit arrestin, undergo the typical arrestin-dependent increase in agonist affinity, or internalize well. Lys326, directly before phenylalanine in the common GPCR motif NPXXY(X)(5-6)F(R/K), was critical for phosphorylation. The 6Q-TRH receptor was not phosphorylated effectively in cells overexpressing GRK2 or in in vitro kinase assays containing purified GRK2. Phosphorylation of the 6Q receptor was partially restored by coexpression of a receptor with an intact helix 8 but without phosphorylation sites. Phosphorylation was inhibited but not completely prevented by alanine substitution for cysteine palmitoylation sites. Positively charged amino acids in the proximal tail of the beta2-adrenergic receptor were also important for GRK-dependent phosphorylation. The results indicate that positive residues in helix 8 of GPCRs are important for GRK-dependent phosphorylation.

Excitation of histaminergic tuberomamillary neurons by thyrotropin-releasing hormone.

The histaminergic tuberomamillary nucleus (TMN) controls arousal and attention, and the firing of TMN neurons is state-dependent, active during waking, silent during sleep. Thyrotropin-releasing hormone (TRH) promotes arousal and combats sleepiness associated with narcolepsy. Single-cell reverse-transcription-PCR demonstrated variable expression of the two known TRH receptors in the majority of TMN neurons. TRH increased the firing rate of most (ca 70%) TMN neurons. This excitation was abolished by the TRH receptor antagonist chlordiazepoxide (CDZ; 50 mum). In the presence of tetrodotoxin (TTX), TRH depolarized TMN neurons without obvious change of their input resistance. This effect reversed at the potential typical for nonselective cation channels. The potassium channel blockers barium and cesium did not influence the TRH-induced depolarization. TRH effects were antagonized by inhibitors of the Na(+)/Ca(2+) exchanger, KB-R7943 and benzamil. The frequency of GABAergic spontaneous IPSCs was either increased (TTX-insensitive) or decreased [TTX-sensitive spontaneous IPSCs (sIPSCs)] by TRH, indicating a heterogeneous modulation of GABAergic inputs by TRH. Facilitation but not depression of sIPSC frequency by TRH was missing in the presence of the kappa-opioid receptor antagonist nor-binaltorphimine. Montirelin (TRH analog, 1 mg/kg, i.p.) induced waking in wild-type mice but not in histidine decarboxylase knock-out mice lacking histamine. Inhibition of histamine synthesis by (S)-alpha-fluoromethylhistidine blocked the arousal effect of montirelin in wild-type mice. We conclude that direct receptor-mediated excitation of rodent TMN neurons by TRH demands activation of nonselective cation channels as well as electrogenic Na(+)/Ca(2+) exchange. Our findings indicate a key role of the brain histamine system in TRH-induced arousal.

Participation of HERG channel cytoplasmic structures on regulation by the G protein-coupled TRH receptor.

Human ether-a-go-go-related gene (HERG) channels heterologously expressed in Xenopus oocytes are regulated by the activation of G protein-coupled hormone receptors that, like the thyrotropin-releasing hormone (TRH) receptor, activate phospholipase C. Previous work with serially deleted HERG mutants suggested that residues 326-345 located in the proximal domain of the channels amino terminus might be required for the hormonal modulation of HERG activation. Generation of new channel mutants deleted in this region further point to the amino acid sequence between residues 326 and 332 as a possible determinant of the TRH effects, but individual or combined single-point mutations in this sequence demonstrate that maintenance of its consensus sites for phosphorylation and/or interaction with regulatory components is not important for the modulatory response(s). The TRH-induced effects also remained unaltered when a basic amino acid cluster located between residues 362 and 366 is eliminated. Additionally, no effect of TRH was observed in channels carrying single-point mutations at the beginning of the intracellular loop linking transmembrane domains S4 and S5. Our results indicate that a correct structural arrangement of the amino terminal domains is essential for the hormone-induced modifications of HERG activation. They also suggest that the hormonal regulatory action is transmitted to the transmembrane channel core through interactions between the cytoplasmic domains and the initial portion of the S4-S5 linker.

High levels of thyrotropin-releasing hormone receptors activate programmed cell death in human pancreatic precursors.

OBJECTIVES: Thyrotropin-releasing hormone (TRH) is expressed in rodent and human adult pancreata and in mouse pancreas during embryonic development. However, expression of TRH receptors (TRHRs) in the pancreas is controversial. We sought to provide evidence that the TRH/TRHR system might play a role in fetal development. METHODS: We used quantitative reverse transcription-polymerase chain reaction to measure TRH and TRHR messenger RNA (mRNA). To study the effects of TRHR expression in a pancreatic progenitor population, we expressed TRHRs in human islet-derived precursor cells (hIPCs) by infection with adenoviral vector AdCMVmTRHR. Thyrotropin-releasing hormone receptor signaling was measured as inositol phosphate production and intracellular calcium transients. Thyrotropin-releasing hormone receptor expression was measured by [H]methyl-TRH binding. Apoptosis was monitored by release of cytochrome c from mitochondria. RESULTS: We show that TRH mRNA is expressed in human fetal and adult pancreata, and that TRHR mRNA is expressed in fetal human pancreas but not in adult human pancreas. Thyrotropin-releasing hormone receptors expressed in hIPCs were shown to signal normally. Most importantly, TRH treatment for several days stimulated apoptosis in hIPCs expressing approximately 400,000 TRHRs per cell. CONCLUSIONS: These findings suggest a possible role for TRH/TRHR signaling in pancreatic precursors to promote programmed cell death, a normal constituent of morphogenesis during embryonic development in humans.

TRH-receptor-type-2-deficient mice are euthyroid and exhibit increased depression and reduced anxiety phenotypes.

Thyrotropin-releasing hormone (TRH) is a neuropeptide that initiates its effects in mice by interacting with two G-protein-coupled receptors, TRH receptor type 1 (TRH-R1) and TRH receptor type 2 (TRH-R2). Two previous reports described the effects of deleting TRH-R1 in mice. TRH-R1 knockout mice exhibit hypothyroidism, hyperglycemia, and increased depression and anxiety-like behavior. Here we report the generation of TRH-R2 knockout mice. The phenotype of these mice was characterized using gross and histological analyses along with blood hematological assays and chemistries. Standard metabolic tests to assess glucose and insulin tolerance were performed. Behavioral testing included elevated plus maze, open field, tail suspension, forced swim, and novelty-induced hypophagia tests. TRH-R2 knockout mice are euthyroid with normal basal and TRH-stimulated serum levels of thyroid-stimulating hormone (thyrotropin), are normoglycemic, and exhibit normal development and growth. Female, but not male, TRH-R2 knockout mice exhibit moderately increased depression-like and reduced anxiety-like phenotypes. Because the behavioral changes in TRH-R1 knockout mice may have been caused secondarily by their hypothyroidism whereas TRH-R2 knockout mice are euthyroid, these data provide the first evidence for the involvement of the TRH/TRH-R system, specifically extrahypothalamic TRH/TRH-R2, in regulating mood and affect.

Understanding the structural and functional differences between mouse thyrotropin-releasing hormone receptors 1 and 2.

Multiple computational methods have been employed in a comparative study of thyrotropin-releasing hormone receptors 1 and 2 (TRH-R1 and TRH-R2) to explore the structural bases for the different functional properties of these G protein-coupled receptors. Three-dimensional models of both murine TRH receptors have been built and optimized by means of homology modeling based on the crystal structure of bovine rhodopsin, molecular dynamics simulations, and energy minimizations in a membrane-aqueous environment. The comparison between the two models showed a correlation between the higher flexibility and higher basal activity of TRH-R2 versus the lesser flexibility and lower basal activity of TRH-R1 and supported the involvement of the highly conserved W6.48 in the signaling process. A correlation between the level of basal activity and conformational changes of TM5 was detected also. Comparison between models of the wild type receptors and their W6.48A mutants, which have reversed basal activities compared with their respective wild types, further supported these correlations. A flexible molecular docking procedure revealed that TRH establishes a direct interaction with W6.48 in TRH-R2 but not in TRH-R1. We designed and performed new mutagenesis experiments that strongly supported these observations.

Thyrotropin-releasing hormone receptor 1-deficient mice display increased depression and anxiety-like behavior.

TRH is a neuropeptide with a variety of hormonal and neurotransmitter/neuromodulator functions. In particular, TRH has pronounced acute antidepressant effects in both humans and animals and has been implicated in the mediation of the effects of other antidepressive therapies. Two G protein-coupled receptors, TRH receptor 1 (TRH-R1) and TRH-R2, have been identified. Here we report the generation and phenotypic characterization of mice deficient in TRH-R1. The TRH-R1 knockout mice have central hypothyroidism and mild hyperglycemia but exhibit normal growth and development and normal body weight and food intake. Behaviorally, the TRH-R1 knockout mice display increased anxiety and depression levels while behaving normally in a number of other aspects examined. These results provide the first direct evidence that the endogenous TRH system is involved in mood regulation, and this function is carried out through TRH-R1-mediated neural pathways.

Discovery of a dual action first-in-class peptide that mimics and enhances CNS-mediated actions of thyrotropin-releasing hormone.

Thyrotropin-releasing hormone (TRH) displays multiple CNS-mediated actions that have long been recognized to have therapeutic potential in treating a wide range of neurological disorders. Investigations of CNS functions and clinical use of TRH are hindered, however, due to its rapid degradation by TRH-degrading ectoenzyme (TRH-DE). We now report the discovery of a set of first-in-class compounds that display unique ability to both potently inhibit TRH-DE and bind to central TRH receptors with unparalleled affinity. This dual pharmacological activity within one molecular entity was found through selective manipulation of peptide stereochemistry. Notably, the lead compound of this set, L-pyroglutamyl-L-asparaginyl-L-prolyl-D-tyrosyl-D-tryptophan amide (Glp-Asn-Pro-D-Tyr-D-TrpNH(2)), is effective in vivo at producing and potentiating central actions of TRH without evoking release of thyroid-stimulating hormone (TSH). Specifically, this peptide displayed high plasma stability and combined potent inhibition of TRH-DE (K(i) 151 nM) with high affinity binding to central TRH receptors (K(i) 6.8 nM). Moreover, intraperitoneal injection of this peptide mimicked and augmented the effects of TRH on behavioural activity in rat. Analogous to TRH, it also antagonized pentobarbital-induced narcosis when administered intravenously. This discovery provides new opportunities for probing the role of TRH actions in the CNS and a basis for development of novel TRH-based neurotherapeutics.

Effects of TRH on heteromeric rat erg1a/1b K+ channels are dominated by the rerg1b subunit.

The erg1a (HERG) K+ channel subunit and its N-terminal splice variant erg1b are coexpressed in several tissues and both isoforms have been shown to form heteromultimeric erg channels in heart and brain. The reduction of erg1a current by thyrotropin-releasing hormone (TRH) is well studied, but no comparable data exist for erg1b. Since TRH and TRH receptors are widely expressed in the brain, we have now studied the different TRH effects on the biophysical properties of homomeric rat erg1b as well as heteromeric rat erg1a/1b channels. The erg channels were overexpressed in the clonal somatomammotroph pituitary cell line GH3/B6, which contains TRH receptors and endogenous erg channels. Compared to rerg1a, homomeric rerg1b channels exhibited not only faster deactivation kinetics, but also considerably less steady-state inactivation, and half-maximal activation occurred at about 10 mV more positive potentials. Coexpression of both isoforms resulted in erg currents with intermediate properties concerning the deactivation kinetics, whereas rerg1a dominated the voltage dependence of activation and rerg1b strongly influenced steady-state inactivation. Application of TRH induced a reduction of maximal erg conductance for all tested erg1 currents without effects on the voltage dependence of steady-state inactivation. Nevertheless, homomeric rerg1b channels significantly differed in their response to TRH from rerg1a channels. The TRH-induced shift in the activation curve to more positive potentials, the dramatic slowing of activation and the acceleration of deactivation typical for rerg1a modulation were absent in rerg1b channels. Surprisingly, most effects of TRH on heteromeric rerg1 channels were dominated by the rerg1b subunit.

Beta-arrestin mediates desensitization and internalization but does not affect dephosphorylation of the thyrotropin-releasing hormone receptor.

The G protein-coupled thyrotropin-releasing hormone (TRH) receptor is phosphorylated and binds to beta-arrestin after agonist exposure. To define the importance of receptor phosphorylation and beta-arrestin binding in desensitization, and to determine whether beta-arrestin binding and receptor endocytosis are required for receptor dephosphorylation, we expressed TRH receptors in fibroblasts from mice lacking beta-arrestin-1 and/or beta-arrestin-2. Apparent affinity for [(3)H]MeTRH was increased 8-fold in cells expressing beta-arrestins, including a beta-arrestin mutant that did not permit receptor internalization. TRH caused extensive receptor endocytosis in the presence of beta-arrestins, but receptors remained primarily on the plasma membrane without beta-arrestin. beta-Arrestins strongly inhibited inositol 1,4,5-trisphosphate production within 10 s. At 30 min, endogenous beta-arrestins reduced TRH-stimulated inositol phosphate production by 48% (beta-arrestin-1), 71% (beta-arrestin-2), and 84% (beta-arrestins-1 and -2). In contrast, receptor phosphorylation, detected by the mobility shift of deglycosylated receptor, was unaffected by beta-arrestins. Receptors were fully phosphorylated within 15 s of TRH addition. Receptor dephosphorylation was identical with or without beta-arrestins and almost complete 20 min after TRH withdrawal. Blocking endocytosis with hypertonic sucrose did not alter the rate of receptor phosphorylation or dephosphorylation. Expressing receptors in cells lacking Galpha(q) and Galpha(11) or inhibiting protein kinase C pharmacologically did not prevent receptor phosphorylation or dephosphorylation. Overexpression of dominant negative G protein-coupled receptor kinase-2 (GRK2), however, retarded receptor phosphorylation. Receptor activation caused translocation of endogenous GRK2 to the plasma membrane. The results show conclusively that receptor dephosphorylation can take place on the plasma membrane and that beta-arrestin binding is critical for desensitization and internalization.

Carboxyl tail cysteine mutants of the thyrotropin-releasing hormone receptor type 1 exhibit constitutive signaling: role of palmitoylation.

We studied the role of carboxyl tail cysteine residues and their palmitoylation in constitutive signaling by the thyrotropin-releasing hormone (TRH) receptor type 1 (TRH-R1) in transfected mammalian cells and in Xenopus laevis oocytes. To study palmitoylation, we inserted a factor Xa cleavage site within the third extracellular loop of TRH-R1, added a carboxyl-terminal C9 immunotag and expressed the mutant receptor in Chinese hamster ovary cells. We identified TRH-R1-specific palmitoylation in the transmembrane helix-7/carboxyl-tail receptor fragment mainly at Cys-335 and Cys-337. In contrast to a mutant truncated at Cys-335 that was reported previously to be constitutively active, a receptor truncated at Lys-338 (K338Stop), which preserves Cys-335 and Cys-337, and C337Stop and N336Stop, which preserve Cys-335, did not exhibit increased constitutive signaling. TRH-R1 mutants substituted singly by Gly or Ser at Cys-335 or Cys-337 did not exhibit constitutive signaling. By contrast, substitution of both cysteines (C335G/C337G or C335S/C337S) yielded TRH-R1 mutants that exhibited marked constitutive signaling in mammalian cells. In the oocyte, constitutive signaling by C335G/C337G resulted in homologous (of C335G/C337G) and heterologous (of M1 muscarinic receptor) desensitization. Because both Cys-335 and Cys-337 have to be substituted or deleted for constitutive signaling, we propose that a single palmitoylation site in the proximal carboxyl tail is sufficient to constrain TRH-R1 in an inactive conformation.

Thyrotropin-releasing hormone in cardiovascular pathophysiology.

Thyrotropin (TSH)-releasing hormone (TRH) also known as thyroliberin was the first of a number of peptides exerting several roles as a hormone and as a neuropeptide. Its ubiquitous distribution in the hypothalamus and in the extrahypothalamic regions and its diverse pharmacological and physiological effects are all features of its dual functions. For this reason, TRH has been the subject of much research throughout the past 20 years, work that has examined the structure, function, distribution, and regulation of the tripeptide and it has been extensively reviewed elsewhere [O'Leary R., O'Connor B. Thyrotropin-releasing hormone. J Neurochem. 1995;65:953-963.; Nillni E., Sevarino K. The biology of pro-thyrotropin-releasing hormone-derived peptides. Endocrine Reviews, 1999;20:599-664.]. After a brief overview of its distribution, hypothalamic and extrahypothalamic functions, and receptors involved, this review discusses efforts devoted to support TRH role in cardiovascular regulation with a main focus on hypertension pathophysiology in experimental models and humans.

Generation of thyrotropin-releasing hormone receptor 1-deficient mice as an animal model of central hypothyroidism.

To provide an animal model of central hypothyroidism, mice deficient in the TRH-receptor 1 (TRH-R1) gene were generated by homologous recombination. The pituitaries of TRH-R1-/- mice are devoid of any TRH-binding capacity, demonstrating that TRH-R1 is the only receptor localized on TRH target cells of the pituitary. With the exception of some retardation in growth rate, TRH-R1-/- mice appear normal, but compared with control animals they exhibit a considerable decrease in serum T(3), T(4), and prolactin (PRL) levels but not in serum TSH levels. In situ hybridization histochemistry and real-time RT-PCR analysis revealed that in adult TRH-R1-/- animals TSHbeta-mRNA expression is not impaired whereas PRL mRNA and GH mRNA levels are considerably reduced compared with control mice. The numbers of thyrotropes, somatotropes, and lactotropes, however, are not affected by the deletion of the TRH-R1 gene. The mutant mice are fertile, and the dams nourish their pups well, indicating that TRH is not a decisive factor for suckling-induced PRL release. In situ hybridization and quantitative RT-PCR analysis, furthermore, revealed that, as in control animals, pituitary PRL-mRNA expression in TRH-R1-/- is considerably increased during lactation, albeit strongly reduced as compared with lactating control animals.

Involvement of thyrotropin-releasing hormone receptor, somatostatin receptor subtype 2 and corticotropin-releasing hormone receptor type 1 in the control of chicken thyrotropin secretion.

Thyrotropin or thyroid-stimulating hormone (TSH) secretion in the chicken is controlled by several hypothalamic hormones. It is stimulated by thyrotropin-releasing hormone (TRH) and corticotropin-releasing hormone (CRH), whereas somatostatin (SRIH) exerts an inhibitory effect. In order to determine the mechanism by which these hypothalamic hormones modulate chicken TSH release, we examined the cellular localization of TRH receptors (TRH-R), CRH receptors type 1 (CRH-R1) and somatostatin subtype 2 receptors (SSTR2) in the chicken pars distalis by in situ hybridization (ISH), combined with immunological staining of thyrotropes. We show that thyrotropes express TRH-Rs and SSTR2s, allowing a direct action of TRH and SRIH at the level of the thyrotropes. CRH-R1 expression is virtually confined to corticotropes, suggesting that CRH-induced adrenocorticotropin release is the result of a direct stimulation of corticotropes, whereas CRH-stimulated TSH release is not directly mediated by the known chicken CRH-R1. Possibly CRH-induced TSH secretion is mediated by a yet unknown type of CRH-R in the chicken. Alternatively, a pro-opiomelanocortin (POMC)-derived peptide, secreted by the corticotropes following CRH stimulation, could act as an activator of TSH secretion in a paracrine way.