Central regulation of hypothalamic-pituitary-thyroid axis under physiological and pathophysiological conditions.

TRH is a tripeptide amide that functions as a neurotransmitter but also serves as a neurohormone that has a critical role in the central regulation of the hypothalamic-pituitary-thyroid axis. Hypophysiotropic TRH neurons involved in this neuroendocrine process are located in the hypothalamic paraventricular nucleus and secrete TRH into the pericapillary space of the external zone of the median eminence for conveyance to anterior pituitary thyrotrophs. Under basal conditions, the activity of hypophysiotropic TRH neurons is regulated by the negative feedback effects of thyroid hormone to ensure stable, circulating, thyroid hormone concentrations, a mechanism that involves complex interactions between hypophysiotropic TRH neurons and the vascular system, cerebrospinal fluid, and specialized glial cells called tanycytes. Hypophysiotropic TRH neurons also integrate other humoral and neuronal inputs that can alter the setpoint for negative feedback regulation by thyroid hormone. This mechanism facilitates adaptation of the organism to changing environmental conditions, including the shortage of food and a cold environment. The thyroid axis is also affected by other adverse conditions such as infection, but the central mechanisms mediating suppression of hypophysiotropic TRH may be pathophysiological. In this review, we discuss current knowledge about the mechanisms that contribute to the regulation of hypophysiotropic TRH neurons under physiological and pathophysiological conditions.

NR4A1 (Nur77) mediates thyrotropin-releasing hormone-induced stimulation of transcription of the thyrotropin ß gene: analysis of TRH knockout mice.

Thyrotropin-releasing hormone (TRH) is a major stimulator of thyrotropin-stimulating hormone (TSH) synthesis in the anterior pituitary, though precisely how TRH stimulates the TSHß gene remains unclear. Analysis of TRH-deficient mice differing in thyroid hormone status demonstrated that TRH was critical for the basal activity and responsiveness to thyroid hormone of the TSHß gene. cDNA microarray and K-means cluster analyses with pituitaries from wild-type mice, TRH-deficient mice and TRH-deficient mice with thyroid hormone replacement revealed that the largest and most consistent decrease in expression in the absence of TRH and on supplementation with thyroid hormone was shown by the TSHß gene, and the NR4A1 gene belonged to the same cluster as and showed a similar expression profile to the TSHß gene. Immunohistochemical analysis demonstrated that NR4A1 was expressed not only in ACTH- and FSH- producing cells but also in thyrotrophs and the expression was remarkably reduced in TRH-deficient pituitary. Furthermore, experiments in vitro demonstrated that incubation with TRH in GH4C1 cells increased the endogenous NR4A1 mRNA level by approximately 50-fold within one hour, and this stimulation was inhibited by inhibitors for PKC and ERK1/2. Western blot analysis confirmed that TRH increased NR4A1 expression within 2 h. A series of deletions of the promoter demonstrated that the region between bp -138 and +37 of the TSHß gene was responsible for the TRH-induced stimulation, and Chip analysis revealed that NR4A1 was recruited to this region. Conversely, knockdown of NR4A1 by siRNA led to a significant reduction in TRH-induced TSHß promoter activity. Furthermore, TRH stimulated NR4A1 promoter activity through the TRH receptor. These findings demonstrated that 1) TRH is a highly specific regulator of the TSHß gene, and 2) TRH mediated induction of the TSHß gene, at least in part by sequential stimulation of the NR4A1-TSHß genes through a PKC and ERK1/2 pathway.

Thyroid hormone exerts negative feedback on hypothalamic type 4 melanocortin receptor expression.

The type 4 melanocortin receptor MC4R, a key relay in leptin signaling, links central energy control to peripheral reserve status. MC4R activation in different brain areas reduces food intake and increases energy expenditure. Mice lacking Mc4r are obese. Mc4r is expressed by hypothalamic paraventricular Thyrotropin-releasing hormone (TRH) neurons and increases energy usage through activation of Trh and production of the thyroid hormone tri-iodothyronine (T(3)). These facts led us to test the hypothesis that energy homeostasis should require negative feedback by T(3) on Mc4r expression. Quantitative PCR and in situ hybridization showed hyperthyroidism reduces Mc4r mRNA levels in the paraventricular nucleus. Comparative in silico analysis of Mc4r regulatory regions revealed two evolutionarily conserved potential negative thyroid hormone-response elements (nTREs). In vivo ChIP assays on mouse hypothalamus demonstrated association of thyroid hormone receptors (TRs) with a region spanning one nTRE. Further, in vivo gene reporter assays revealed dose-dependent T(3) repression of transcription from the Mc4r promoter in mouse hypothalamus, in parallel with T(3)-dependent Trh repression. Mutagenesis of the nTREs in the Mc4r promoter demonstrated direct regulation by T(3), consolidating the ChIP results. In vivo shRNA knockdown, TR over-expression approaches and use of mutant mice lacking specific TRs showed that both TRalpha and TRbeta contribute to Mc4r regulation. T(3) repression of Mc4r transcription ensures that the energy-saving effects of T(3) feedback on Trh are not overridden by MC4R activation of Trh. Thus parallel repression by T(3) on hypothalamic Mc4r and Trh contributes to energy homeostasis.

Excitatory neuromodulator reduces dopamine release, enhancing prolactin secretion.

Hypothalamic dopamine neurons inhibit pituitary prolactin secretion. In this issue of Neuron, Lyons et al. provide evidence for a novel model, whereby the excitatory neuropeptide TRH depolarizes gap-junction-coupled dopamine neurons, leading to a shift in the population pattern of action potentials from phasic burst firing to regular tonic firing, hypothetically reducing dopamine release while increasing total spike number.

Thyrotropin-releasing hormone modulates vasopressin and oxytocin synthesis and release from the hypothalamo-neurohypophysial system of different age male rats.

Thyrotropin-releasing hormone (TRH) is engaged in the modulation of the hypothalamo-neurohypophysial system activity. Effects of repeated intravenously injections of TRH in a dose of 100 ng/100 g b.w. on vasopressin (VP) and oxytocin (OT) biosynthesis and release from the hypothalamo-neurohypophysial system was investigated in rats in different age (1-, 3- or 7-months of the life). To estimate the biosynthesis rate of both neurohormones the colchicine procedure was used (the dose of 5 microg/5 microl icv 20 hours before the decapitation). It has been observed that vasopressin synthesis in the hypothalamus increased gradually with maturation of rats, while OT biosynthesis decreased in the same animals. Hypothalamic biosynthesis rate of VP and OT is most effective in youngest rats and declines during the adolescence of animals. Thyrotropin-releasing hormone directly affects VP-ergic and OT-ergic hypothalamic neurons activity and both neurohormones biosynthesis process. This effect, however, is opposed: TRH acts as a stimulator of vasopressin biosynthesis most of all in young male rats and as an inhibitor for oxytocin biosynthesis especially in mature animals.

Thyrotropin-releasing hormone (TRH) in the cerebellum.

Thyrotropin-releasing hormone (TRH) was originally isolated from the hypothalamus. Besides controlling the secretion of TSH from the anterior pituitary, this tripeptide is widely distributed in the central nervous system and regarded as a neurotransmitter or modulator of neuronal activities in extrahypothalamic regions, including the cerebellum. TRH has an important role in the regulation of energy homeostasis, feeding behavior, thermogenesis, and autonomic regulation. TRH controls energy homeostasis mainly through its hypophysiotropic actions to regulate circulating thyroid hormone levels. Recent investigations have revealed that TRH production is regulated directly at the transcriptional level by leptin, one of the adipocytokines that plays a critical role in feeding and energy expenditure. The improvement of ataxic gait is one of the important pharmacological properties of TRH. In the cerebellum, cyclic GMP has been shown to be involved in the effects of TRH. TRH knockout mice show characteristic phenotypes of tertiary hypothyroidism, but no morphological changes in their cerebellum. Further analysis of TRH-deficient mice revealed that the expression of PFTAIRE protein kinase1 (PFTK1), a cdc2-related kinase, in the cerebellum was induced by TRH through the NO-cGMP pathway. The antiataxic effect of TRH and TRH analogs has been investigated in rolling mouse Nagoya (RMN) or 3-acetylpyridine treated rats, which are regarded as a model of human cerebellar degenerative disease. TRH and TRH analogs are promising clinical therapeutic agents for inducing arousal effects, amelioration of mental depression, and improvement of cerebellar ataxia.

Thyrotropin releasing hormone (TRH) may preserve pancreatic islet cell function: potential role in the treatment of diabetes mellitus.

Thyrotropin Releasing Hormone (TRH), first identified in the hypothalamus as a regulator of the Pituitary-Thyroid axis, has also been found in the beta-cell of the pancreas co-localised with insulin. The significance of this association is emphasised by the report that the TRH knock-out (KO) mouse is hyperglycemic. These findings have led to speculation that TRH may have a physiologic role in the regulation of carbohydrate metabolism. To understand better the role of TRH in the pancreas, TRH was administered to rats rendered diabetic from streptozotocin damage to the islets of Langerhans. This resulted in almost complete normalisation of the profound hyperglycemia. TRH is capable of reversing Diabetes Mellitus (DM) in an experimental animal model, possibly by promoting neogenesis of beta cells through induction of adult stem cells in the pancreas. These studies point to a potential therapeutic role for TRH in the treatment of DM in man.

Copeptin, derived from isotocin precursor, is a probable prolactin releasing factor in carp.

Control of prolactin (PRL) release is of crucial importance for the multiple functions exerted by PRL in vertebrates. Recently identified hypothalamic PRL-releasing peptides displayed additional neuromodulatory activities and in fish only few could be detected close to lactotrophs. Here we describe the C-terminal peptide processed from the carp isotocin precursor as probable physiologically relevant regulator of PRL release in carp. The amino acid sequence derived from the complete isotocin precursor gene of Cyprinus carpio, predicted a C-terminal peptide uncleaved between the neurophysin (Np) and copeptin (Cp) domain. Accordingly, antibodies against synthetic Np- and Cp-specific oligopeptides both immunodetected a 13kDa protein (cNpCp) in total pituitary proteins and showed abundant immunoreaction in hypothalamic axons in direct contact with lactotrophs in the rostral pars distalis of carp pituitary gland sections. Finally, incubation of cultured carp pituitary explants with purified carp cNpCp resulted in a potent stimulation of PRL release.

Prolactin-releasing peptide, food intake, and hydromineral balance in goldfish.

A potential role for prolactin-releasing peptide (PrRP) in appetite regulation and hydromineral balance in goldfish was examined. PrRP was found to be expressed in discrete regions of the goldfish brain, in particular, the hypothalamus. Intraperitoneal (IP) or intracerebroventricular administration of PrRP had dose-dependent effects to suppress food intake in goldfish. Hypothalamic PrRP mRNA expression significantly increased after feeding, as well as after 7 days of food deprivation. Refeeding fish after 7 days food deprivation did not result in a postprandial increase in PrRP mRNA expression. These data suggest an anorexigenic role for PrRP in the short term around a scheduled meal time, but not over the longer term. IP injection of PrRP significantly increased pituitary prolactin (PRL) mRNA levels, suggesting involvement in the regulation of lactotroph activity. Acclimating goldfish to an ion-poor environment decreased serum osmolality and increased PrRP and PRL mRNA levels, providing evidence for PrRP involvement in hydromineral balance through its actions on lactotrophs. Acclimation to ion-poor water diminished the anorexigenic properties of PrRP in goldfish, indicating that a role for PrRP in goldfish satiation is counterbalanced by alternate systemic needs (i.e., osmoregulatory). This was further supported by an ability to reinstate the anorexigenic actions of PrRP in fish acclimated to ion-poor water by feeding a salt-rich diet. These studies provide evidence that PrRP is involved in regulating appetite and hydromineral balance in fish, and that the degree of involvement in either process varies according to overall systemic needs in response to environmental conditions.

Dominant role of thyrotropin-releasing hormone in the hypothalamic-pituitary-thyroid axis.

Hypothalamic thyrotropin-releasing hormone (TRH) stimulates thyroid-stimulating hormone (TSH) secretion from the anterior pituitary. TSH then initiates thyroid hormone (TH) synthesis and release from the thyroid gland. Although opposing TRH and TH inputs regulate the hypothalamic-pituitary-thyroid axis, TH negative feedback is thought to be the primary regulator. This hypothesis, however, has yet to be proven in vivo. To elucidate the relative importance of TRH and TH in regulating the hypothalamic-pituitary-thyroid axis, we have generated mice that lack either TRH, the beta isoforms of TH receptors (TRbeta KO), or both (double KO). TRbeta knock-out (KO) mice have significantly higher TH and TSH levels compared with wild-type mice, in contrast to double KO mice, which have reduced TH and TSH levels. Unexpectedly, hypothyroid double KO mice also failed to mount a significant rise in serum TSH levels, and pituitary TSH immunostaining was markedly reduced compared with all other hypothyroid mouse genotypes. This impaired TSH response, however, was not due to a reduced number of pituitary thyrotrophs because thyrotroph cell number, as assessed by counting TSH immunopositive cells, was restored after chronic TRH treatment. Thus, TRH is absolutely required for both TSH and TH synthesis but is not necessary for thyrotroph cell development.

Hypothalamic control of the thyroidal axis in the chicken: over the boundaries of the classical hormonal axes.

The pituitary gland, occupying a central position in the hypothalamo-pituitary thyroidal axis, produces thyrotropin (TSH), which is known to stimulate the thyroid gland to synthetize and release its products, thyroid hormones. TSH is produced by a specific cell population in the pituitary, the so-called thyrotropes. Their secretory activity is controlled by the hypothalamus, releasing both stimulatory and inhibitory factors that reach the pituitary through a portal system of blood vessels. Based on early experiments in mammals, thyrotropin-releasing hormone (TRH) is generally mentioned as the main stimulator of the thyrotropes. During the past few decades, it has become clear that the hypophysiotropic function of the hypothalamus is more complex, with different hormonal axes interacting with each other. In the chicken, it was found that not only TRH, but also corticotropin-releasing hormone (CRH), the main stimulator of corticotropin release, is a potent stimulator of TSH secretion. Somatostatin (SRIH), a hypothalamic factor known for its inhibitory effect on growth hormone secretion, was demonstrated to blunt the TSH response to TRH and CRH. In this review we summarize the latest studies concerning the "interaxial" hypothalamic control of TSH release in the chicken, with a special emphasis on the molecular components of these control mechanisms. It remains to be demonstrated if these findings could also be extrapolated to other species or classes of vertebrates.

Thyrotropin-releasing hormone induced thermogenesis in Syrian hamsters: site of action and receptor subtype.

Early work in our laboratory has revealed the important role played by thyrotropin-releasing hormone (TRH) in the arousal from hibernation in Syrian hamsters. In the present study, we investigated the thermogenic mechanism of TRH in Syrian hamsters. Six to 10 female Syrian hamsters were used in the respective experiments. Intracerebroventricular (icv) injection of TRH elevated the intrascapular brown adipose tissue (IBAT) temperature (T(IBAT)) and rectal temperature (T rec) in Syrian hamsters. Thermogenic response of icv TRH was suppressed by bilateral denervation of the sympathetic nerve. Icv injection of TRH increased the norepinephrin (NE) turnover rate in IBAT without affecting the total serum triiodothyronine (T3) level. Moreover, TRH microinjections into the dorsomedial hypothalamus (DMH), preoptic area (PO), anterior hypothalamus (AH) and ventromedial hypothalamus (VMH) induced T(IBAT) and T(rec) increases. However, neither T(IBAT) nor T rec was affected by similar TRH administrations into the lateral hypothalamus and posterior hypothalamus. Interestingly, although TRH-induced hyperthermia was suppressed by pretreatment of anti-TRH-R1 antibodies, no changes were induced by anti-TRH-R2 antibodies. These results suggest that the sites of action of TRH associated with thermogenesis are probably localized in the DMH, PO, AH and VMH. In addition, TRH-induced thermogenesis is probably elicited by facilitation of the sympathetic nerve system via the central TRH-R1 irrelevant of T3.

Excitatory effects of thyrotropin-releasing hormone in the thalamus.

The activity of the thalamus is state dependent. During slow-wave sleep, rhythmic burst firing is prominent, whereas during waking or rapid eye movement sleep, tonic, single-spike activity dominates. These state-dependent changes result from the actions of modulatory neurotransmitters. In the present study, we investigated the functional and cellular effects of the neuropeptide thyrotropin-releasing hormone (TRH) on the spontaneously active ferret geniculate slice. This peptide and its receptors are prominently expressed in the thalamic network, yet the role of thalamic TRH remains obscure. Bath application of TRH resulted in a transient cessation of both spindle waves and the epileptiform slow oscillation induced by application of bicuculline. With intracellular recordings, TRH application to the GABAergic neurons of the perigeniculate (PGN) or thalamocortical cells in the lateral geniculate nucleus resulted in depolarization and increased membrane resistance. In perigeniculate neurons, this effect reversed near the reversal potential for K+, suggesting that it is mediated by a decrease in K+ conductance. In thalamocortical cells, the TRH-induced depolarization was of sufficient amplitude to block the generation of rebound Ca2+ spikes, whereas the even larger direct depolarization of PGN neurons transformed these cells from the burst to tonic, single-spike mode of action potential generation. Furthermore, application of TRH prominently enhanced the afterdepolarization that follows rebound Ca2+ spikes, suggesting that this transmitter may also enhance Ca2+-activated nonspecific currents. These data suggest a novel role for TRH in the brain as an intrinsic regulator of thalamocortical network activity and provide a potential mechanism for the wake-promoting and anti-epileptic effects of this peptide.

Hypothalamic hormones a.k.a. hypothalamic releasing factors.

Originally searched for and eventually isolated as factors of hypothalamic origin controlling anterior pituitary secretions, these hypophysiotropic peptides are now a chapter of physiology and medical endocrinology of their own. Defying the concept of 'neuropeptides' they and their receptors are now known to be ubiquitous and to have subtle as well as profound effects on a large number of functions of both soma and psyche. This review will be composed of brief essays on current knowledge of each of the original 'hypothalamic hormones', TRH, GnRH, somatostatin, GHRH and corticotropin releasing hormone and will close on possible and probable futures.

Low temperature stimulates alpha-melanophore-stimulating hormone secretion and inhibits background adaptation in Xenopus laevis.

It is well-known that alpha-melanophore-stimulating hormone (alpha-MSH) release from the amphibian pars intermedia (PI) depends on the light condition of the animal's background, permitting the animal to adapt the colour of its skin to background light intensity. In the present study, we carried out nine experiments on the effect of low temperature on this skin adaptation process in the toad Xenopus laevis, using the skin melanophore index (MI) bioassay and a radioimmunoassay to measure skin colour adaptation and alpha-MSH secretion, respectively. We show that temperatures below 8 degrees C stimulate alpha-MSH secretion and skin darkening, with a maximum at 5 degrees C, independent of the illumination state of the background. No significant stimulatory effect of low temperature on the MI and alpha-MSH plasma contents was noted when the experiment was repeated with toads from which the neurointermediate lobe (NIL) had been surgically extirpated. This indicates that low temperature stimulates alpha-MSH release from melanotrope cells located in the PI. An in vitro superfusion study with the NIL demonstrated that low temperature does not act directly on the PI. A possible role of the central nervous system in cold-induced alpha-MSH release from the PI was tested by studying the hypothalamic expression of c-Fos (as an indicator for neuronal activity) and the coexistence of c-Fos with the regulators of melanotrope cell activity, neuropeptide Y (NPY) and thyrotrophin-releasing hormone (TRH), using double fluorescence immunocytochemistry. Upon lowering temperature from 22 degrees C to 5 degrees C, in white-adapted animals c-Fos expression decreased in NPY-producing suprachiasmatic-melanotrope-inhibiting neurones (SMIN) in the ventrolateral area of the suprachiasmatic nucleus (SC) but increased in TRH-containing neurones of the magnocellular nucleus. TRH is known to stimulate melanotrope alpha-MSH release. We conclude that temperatures around 5 degrees C inactivate the SMIN in the SC and activate TRH-neurones in the magnocellular nucleus, resulting in enhanced alpha-MSH secretion from the PI, darkening the skin of white-adapted X. laevis.

Feedback regulation of thyrotropin-releasing hormone (TRH): mechanisms for the non-thyroidal illness syndrome.

Regulation of the hypothalamic-pituitary-thyroid (HPT) axis is dependent upon the secretion of thyrotropin-releasing hormone (TRH), a tripeptide originating in the hypothalamic paraventricular nucleus (PVN). These so-called hypophysiotropic neurons are under feedback inhibition by circulating levels of thyroid hormone, mediated through interactions with the beta2 thyroid hormone receptor (TRbeta2) and competition with the phosphorylated form of cyclic adenosine 5'-monophosphate response element binding protein (CREB) for a multifunctional binding site in the TRH gene. The non-thyroidal illness syndrome, characterized by low circulating thyroid hormone levels yet suppression of TRH gene expression in hypophysiotropic neurons, is due to alteration in the regulatory factors that modulate TRH gene expression to result in central hypothyroidism. These factors include alpha melanocyte-stimulating hormone (alphaMSH) and cocaine- and amphetamine-regulated transcript (CART), and agouti-related protein (AGRP) and neuropeptide Y (NPY), substances co-produced by distinct populations of leptin-responsive neurons in the hypothalamic arcuate nucleus. Through monosynaptic projections from arcuate nucleus neurons to hypophysiotropic TRH neurons, these factors contribute to suppression of HPT axis during fasting and starvation by exerting opposing actions on the TRH gene, altering the sensitivity for feedback inhibition by thyroid hormone. In contrast, central hypothyroidism associated with infection may be due to upregulation of type 2 deiodinase activity in tanycytes, specialized glial cells that line the infralateral walls and floor of the third ventricle. Through tanycyte-cerebrospinal fluid, -vascular or -neuronal associations, these cells may lead to inhibition of TRH gene expression in hypophysiotropic neurons by increasing local triiodothyronine production.

A novel TRH-PFTAIRE protein kinase 1 pathway in the cerebellum: subtractive hybridization analysis of TRH-deficient mice.

TRH has been reported to possess several neurophysiological actions in the brain. To gain insights into the molecular mechanisms underlying these effects, particularly in the cerebellum, we attempted to clone a cDNA that was regulated by TRH using TRH knockout mice and subtractive cDNA analysis. Over 100 clones obtained by subtractive hybridization analysis between the wild-type and TRH-1-cerebellum were analyzed. Four clones among them were identical and cdc2-related kinase (PFTAIRE protein kinase 1 (PFTK1)) cDNA, which was previously reported to be expressed only in the brain and testis. PFTK1 mRNA levels in the euthyroid TRH-1- cerebellum supplemented with thyroid hormone were significantly decreased compared with those in the wild-type. Induction of PFTK1 mRNA by TRH was also observed in a time- and dose-dependent manner in human medulloblastoma-derived HTB-185 cells that expressed TRH receptor subtype I mRNA. In addition, treatment of 8-Br-cGMP significantly increased PFTK1 mRNA levels, and a specific inhibitor of cGMP production, ODQ, completely blocked TRH-induced expression of PFTK1 mRNA. Furthermore, induction of PFrK1 mRNA by TRH was significantly inhibited by a NOS specific inhibitor, L-NAME, but not by a MEK inhibitor, PD98059 or a calcium channel inhibitor, nimodipine. These findings demonstrated, for the first time, a novel pathway between a neuropeptide and a cell cycle related peptide in the brain, and PFTK1 may be a key regulator for TRH action in t he cerebellum through t he NO-cGMP pathway.

A novel function of prolactin-releasing peptide in the control of growth hormone via secretion of somatostatin from the hypothalamus.

The present study examined a novel function of PRL-releasing peptide (PrRP) on the neuroendocrine. PrRP-immunoreactive nerve fibers and nerve terminals were located in the vicinity of the somatostatin (SOM)-neurons in the hypothalamic periventricular nucleus (PerVN). Immuno-electron microscopy revealed that PrRP-immunoreactive nerve terminals made synaptic contacts with nonimmunoreactive neuronal elements in the PerVN. Intracerebroventricular (icv) administration of PrRP induced immediate early gene, NGFI-A, in SOM-neurons in the PerVN. Double-labeling in situ hybridization showed that some parts of SOM-neurons in the PerVN expressed PrRP receptor messenger RNA. Therefore, some parts of SOM-neurons in the PerVN are considered to be directly innervated by PrRP via PrRP receptor. In addition to the above morphological characteristics, icv administration of PrRP decreased plasma GH levels. Such inhibitory effects of PrRP on the secretion of GH from the anterior pituitary were diminished by depletion or neutralization of SOM. From these findings it was strongly suggested that SOM-neurons respond to PrRP and secrete SOM into the portal vessels and thus inhibit GH secretion from the anterior pituitary.