Sex-dependent and -independent regulation of thyrotropin-releasing hormone expression in the hypothalamic dorsomedial nucleus by negative energy balance, exercise, and chronic stress.

The dorsomedial nucleus of the hypothalamus (DMH) is part of the brain circuits that modulate organism responses to the circadian cycle, energy balance, and psychological stress. A large group of thyrotropin-releasing hormone (Trh) neurons is localized in the DMH; they comprise about one third of the DMH neurons that project to the lateral hypothalamus area (LH). We tested their response to various paradigms. In male Wistar rats, food restriction during adulthood, or chronic variable stress (CVS) during adolescence down-regulated adult DMH Trh mRNA levels compared to those in sedentary animals fed ad libitum; two weeks of voluntary wheel running during adulthood enhanced DMH Trh mRNA levels compared to pair-fed rats. Except for their magnitude, female responses to exercise were like those in male rats; in contrast, in female rats CVS did not change DMH Trh mRNA levels. A very strong negative correlation between DMH Trh mRNA levels and serum corticosterone concentration in rats of either sex was lost in CVS rats. CVS canceled the response to food restriction, but not that to exercise in either sex. TRH receptor 1 (Trhr) cells were numerous along the rostro-caudal extent of the medial LH. In either sex, fasting during adulthood reduced DMH Trh mRNA levels, and increased LH Trhr mRNA levels, suggesting fasting may inhibit the activity of TRHDMH->LH neurons. Thus, in Wistar rats DMH Trh mRNA levels are regulated by negative energy balance, exercise and chronic variable stress through sex-dependent and -independent pathways.

Sex Dimorphic Responses of the Hypothalamus-Pituitary-Thyroid Axis to Energy Demands and Stress.

The hypothalamus-pituitary-thyroid-axis (HPT) is one of the main neuroendocrine axes that control energy expenditure. The activity of hypophysiotropic thyrotropin releasing hormone (TRH) neurons is modulated by nutritional status, energy demands and stress, all of which are sex dependent. Sex dimorphism has been associated with sex steroids whose concentration vary along the life-span, but also to sex chromosomes that define not only sexual characteristics but the expression of relevant genes. In this review we describe sex differences in basal HPT axis activity and in its response to stress and to metabolic challenges in experimental animals at different stages of development, as well as some of the limited information available on humans. Literature review was accomplished by searching in Pubmed under the following words: "sex dimorphic" or "sex differences" or "female" or "women" and "thyrotropin" or "thyroid hormones" or "deiodinases" and "energy homeostasis" or "stress". The most representative articles were discussed, and to reduce the number of references, selected reviews were cited.

Sex Dimorphic Changes in Trh Gene Methylation and Thyroid-Axis Response to Energy Demands in Maternally Separated Rats.

The hypothalamus-pituitary-thyroid (HPT) axis regulates energy balance through the pleiotropic action of thyroid hormones. HPT basal activity and stimulation by cold or voluntary exercise are repressed by previous chronic stress in adults. Maternal separation (MS) modifies HPT basal activity; we thus studied the response of the axis to energy demands and analyzed possible epigenetic changes on Trh promoter. Nonhandled (NH) or MS male Wistar rats were cold exposed 1 h at adulthood; Trh expression in the hypothalamic paraventricular nucleus (PVN) and serum thyrotropin (TSH) concentration were increased only in NH rats. Two weeks of voluntary exercise decreased fat mass and increased Trh expression, and thyroid hormones concentration changed proportionally to running distance in NH male rats and MS male rats. Although NH females ran more than MS and much more than males, exercise decreased body weight and fat mass only in NH rats with no change on any parameter of the HPT axis but increased Pomc expression in arcuate-nucleus of NH and Npy in MS females. Overall, the methylation pattern of PVN Trh gene promoter was similar in NH males and females; MS modified methylation of specific CpG sites, a thyroid hormone receptor (THR)-binding site present after the initiation site was hypomethylated in MS males; in MS females, the THR binding site of the proximal promoter (site 4) and 2 sites in the first intron were hypermethylated. Our studies showed that, in a sex-dimorphic manner, MS blunted the responses of HPT axis to energy demands in adult animals and caused methylation changes on Trh promoter that could alter T3 feedback.

Evolution of thyrotropin-releasing factor extracellular communication units.

Thyroid hormones (THs) are ancient signaling molecules that contribute to the regulation of metabolism, energy homeostasis and growth. In vertebrates, the hypothalamus-pituitary-thyroid (HPT) axis links the corresponding organs through hormonal signals, including thyrotropin releasing factor (TRF), and thyroid stimulating hormone (TSH) that ultimately activates the synthesis and secretion of THs from the thyroid gland. Although this axis is conserved among most vertebrates, the identity of the hypothalamic TRF that positively regulates TSH synthesis and secretion varies. We review the evolution of the hypothalamic factors that induce TSH secretion, including thyrotropin-releasing hormone (TRH), corticotrophin-releasing hormone (CRH), urotensin-1-3, and sauvagine, and non-mammalian glucagon-like peptide in metazoans. Each of these peptides is part of an extracellular communication unit likely composed of at least 3 elements: the peptide, G-protein coupled receptor and bioavailability regulator, set up on the central neuroendocrine articulation. The bioavailability regulators include a TRH-specific ecto-peptidase, pyroglutamyl peptidase II, and a CRH-binding protein, that together with peptide secretion/transport rate and transduction coupling and efficiency at receptor level shape TRF signal intensity and duration. These vertebrate TRF communication units were coopted from bilaterian ancestors. The bona fide elements appeared early in chordates, and are either used alternatively, in parallel, or sequentially, in different vertebrate classes to control centrally the activity of the HPT axis. Available data also suggest coincidence between apparition of ligand and bioavailability regulator.

The Thyrotropin-Releasing Hormone-Degrading Ectoenzyme, a Therapeutic Target?

Thyrotropin releasing hormone (TRH: Glp-His-Pro-NH2) is a peptide mainly produced by brain neurons. In mammals, hypophysiotropic TRH neurons of the paraventricular nucleus of the hypothalamus integrate metabolic information and drive the secretion of thyrotropin from the anterior pituitary, and thus the activity of the thyroid axis. Other hypothalamic or extrahypothalamic TRH neurons have less understood functions although pharmacological studies have shown that TRH has multiple central effects, such as promoting arousal, anorexia and anxiolysis, as well as controlling gastric, cardiac and respiratory autonomic functions. Two G-protein-coupled TRH receptors (TRH-R1 and TRH-R2) transduce TRH effects in some mammals although humans lack TRH-R2. TRH effects are of short duration, in part because the peptide is hydrolyzed in blood and extracellular space by a M1 family metallopeptidase, the TRH-degrading ectoenzyme (TRH-DE), also called pyroglutamyl peptidase II. TRH-DE is enriched in various brain regions but is also expressed in peripheral tissues including the anterior pituitary and the liver, which secretes a soluble form into blood. Among the M1 metallopeptidases, TRH-DE is the only member with a very narrow specificity; its best characterized biological substrate is TRH, making it a target for the specific manipulation of TRH activity. Two other substrates of TRH-DE, Glp-Phe-Pro-NH2 and Glp-Tyr-Pro-NH2, are also present in many tissues. Analogs of TRH resistant to hydrolysis by TRH-DE have prolonged central efficiency. Structure-activity studies allowed the identification of residues critical for activity and specificity. Research with specific inhibitors has confirmed that TRH-DE controls TRH actions. TRH-DE expression by ß2-tanycytes of the median eminence of the hypothalamus allows the control of TRH flux into the hypothalamus-pituitary portal vessels and may regulate serum thyrotropin secretion. In this review we describe the critical evidences that suggest that modification of TRH-DE activity in tanycytes, and/or in other brain regions, may generate beneficial consequences in some central and metabolic disorders and identify potential drawbacks and missing information needed to test these hypotheses.

A Glial-Neuronal Circuit in the Median Eminence Regulates Thyrotropin-Releasing Hormone-Release via the Endocannabinoid System.

Based on the type-I cannabinoid receptor (CB1) content of hypophysiotropic axons and the involvement of tanycytes in the regulation of the hypothalamic-pituitary-thyroid (HPT) axis, we hypothesized that endocannabinoids are involved in the tanycyte-induced regulation of TRH release in the median eminence (ME). We demonstrated that CB1-immunoreactive TRH axons were associated to DAGLα-immunoreactive tanycyte processes in the external zone of ME and showed that endocannabinoids tonically inhibit the TRH release in this tissue. We showed that glutamate depolarizes the tanycytes, increases their intracellular Ca2+ level and the 2-AG level of the ME via AMPA and kainite receptors and glutamate transport. Using optogenetics, we demonstrated that glutamate released from TRH neurons influences the tanycytes in the ME. In summary, tanycytes regulate TRH secretion in the ME via endocannabinoid release, whereas TRH axons regulate tanycytes by glutamate, suggesting the existence of a reciprocal microcircuit between tanycytes and TRH terminals that controls TRH release.

Sexually dimorphic dynamics of thyroid axis activity during fasting in rats.

Starvation induces tertiary hypothyroidism in adult rodents. Response of the hypothalamus-pituitary-thyroid (HPT) axis to starvation is stronger in adult males than in females. To improve the description of this sexual dimorphism, we analyzed the dynamics of HPT axis response to fasting at multiple levels. In adult rats of the same cohort, 24 and 48 h of starvation inhibited paraventricular nucleus Trh expression and serum concentrations of TSH and T4 earlier in males than in females, with lower intensity in females than in males. In adult females fasted for 36-72 h, serum TSH concentration decreased after 36 h, when the activity of thyrotropin-releasing hormone (TRH)-degrading ectoenzyme was increased in the median eminence. The kinetics of these events were distinct from those previously observed in male rats. We suggest that the sex difference in TSH secretion kinetics is driven not only at the level of paraventricular nucleus TRH neurons, but also by differences in post-secretory catabolism of TRH, with enhancement of TRH-degrading activity more sustained in male than female animals.

Tanycytes and the Control of Thyrotropin-Releasing Hormone Flux Into Portal Capillaries.

Central and peripheral mechanisms that modulate energy intake, partition and expenditure determine energy homeostasis. Thyroid hormones (TH) regulate energy expenditure through the control of basal metabolic rate and thermogenesis; they also modulate food intake. TH concentrations are regulated by the hypothalamus-pituitary-thyroid (HPT) axis, and by transport and metabolism in blood and target tissues. In mammals, hypophysiotropic thyrotropin-releasing hormone (TRH) neurons of the paraventricular nucleus of the hypothalamus integrate energy-related information. They project to the external zone of the median eminence (ME), a brain circumventricular organ rich in neuron terminal varicosities and buttons, tanycytes, other glial cells and capillaries. These capillary vessels form a portal system that links the base of the hypothalamus with the anterior pituitary. Tanycytes of the medio-basal hypothalamus express a repertoire of proteins involved in transport, sensing, and metabolism of TH; among them is type 2 deiodinase, a source of 3,3',5-triiodo-L-thyronine necessary for negative feedback on TRH neurons. Tanycytes subtypes are distinguished by position and phenotype. The end-feet of ß2-tanycytes intermingle with TRH varicosities and terminals in the external layer of the ME and terminate close to the ME capillaries. Besides type 2 deiodinase, ß2-tanycytes express the TRH-degrading ectoenzyme (TRH-DE); this enzyme likely controls the amount of TRH entering portal vessels. TRH-DE is rapidly upregulated by TH, contributing to TH negative feedback on HPT axis. Alterations in energy balance also regulate the expression and activity of TRH-DE in the ME, making ß2-tanycytes a hub for energy-related regulation of HPT axis activity. ß2-tanycytes also express TRH-R1, which mediates positive effects of TRH on TRH-DE activity and the size of ß2-tanycyte end-feet contacts with the basal lamina adjacent to ME capillaries. These end-feet associations with ME capillaries, and TRH-DE activity, appear to coordinately control HPT axis activity. Thus, down-stream of neuronal control of TRH release by action potentials arrival in the external layer of the median eminence, imbricated intercellular processes may coordinate the flux of TRH into the portal capillaries. In conclusion, ß2-tanycytes appear as a critical cellular element for the somatic and post-secretory control of TRH flux into portal vessels, and HPT axis regulation in mammals.

Voluntary Exercise-Induced Activation of Thyroid Axis and Reduction of White Fat Depots Is Attenuated by Chronic Stress in a Sex Dimorphic Pattern in Adult Rats.

The activity of the hypothalamus-pituitary-thyroid (HPT) axis is inhibited by energy deficit, by acute or chronic stress, but activated by cold exposure or exercise. Because stress curtails acute cold induced activation of HPT, we evaluated the effect of chronic stress on HPT axis response to voluntary exercise, a persistent energy-demanding situation. Adult male and female Wistar rats were exposed to restraint stress, 30 min/day for 2 weeks, or to isolation (Iso) [post-natal day [PND] 30-63]. Exercise was performed (7 p.m.-7 a.m.) in a running wheel, sedentary controls stayed in individual cages (Sed); at 7 a.m. they were housed with their cage mate or individually (Iso); food intake by the exercised group was measured day and night to pair-fed Sed. At sacrifice, hormones, mRNA levels and tissue weights were quantified. Control or restrained adult rats had access to running wheel daily for 2 weeks. Compared to C, exercise decreased white adipose tissue (WAT) mass in females and males, increased hypothalamic paraventricular nucleus (PVN)-Trh expression in males proportionally to exercise performed, and increased TSH and T4 serum concentration in females. These changes were not detected in restrained groups. Starting at PND 63 control (2/cage) and isolated (1/cage) rats either exercised on 10 alternated nights or were sedentary. In control male animals, compared to Sed rats, exercise did not decrease WAT mass, nor changed HPT axis activity, but increased Pomc and deiodinase 2 (Dio2) expression in mediobasal hypothalamus (MBH), adrenergic receptor ß3 and uncoupling protein-1 in brown adipose tissue. In control female animals, exercise decreased WAT mass, increased Pomc, Dio2, and Trhde expression in MBH, and TSH serum concentration. Iso females had lower TSH and T4 serum concentration, Dio2 and Trhde expression in MBH than controls. The stress response was higher in isolated males than females, but in males it did not alter the effects of exercise, in contrast to isolated females that had a blunted response to exercise compared to controls. In conclusion, chronic stress interferes with metabolic effects produced by exercise, such as loss of WAT mass, coincident with dampening of HPT activity.

Sex Dimorphic Responses of the Hypothalamus-Pituitary-Thyroid Axis to Maternal Separation and Palatable Diet.

Neonatal stress contributes to the development of obesity and has long-lasting effects on elements of the hypothalamus-pituitary-thyroid (HPT) axis. Given the importance of thyroid hormones in metabolic regulation, we studied the effects of maternal separation and a high-fat/high-carbohydrate diet (HFC), offered from puberty or adulthood, on HPT axis activity of adult male and female Wistar rats. Pups were non-handled (NH) or maternally separated (MS) 3 h/day at postnatal days (Pd) 2-21. In a first experiment, at Pd60, rats had access to chow or an HFC diet (cookies, peanuts, chow) for 1 month. Male and female NH and MS rats that consumed the HFC diet increased their caloric intake, body weight, and serum insulin levels; fat weight increased in all groups except in MS males, and serum leptin concentration increased only in females. Mediobasal hypothalamus (MBH) Pomc expression increased in NH-HFC females and Npy decreased in NH-HFC males. MS males showed insulinemia and hypercortisolemia that was attenuated by the HFC diet. The HPT axis activity response to an HFC diet was sex-specific; expression of MBH thyrotropin-releasing hormone-degrading ectoenzyme (Trhde) increased in NH and MS males; serum TSH concentration decreased in NH males, and T4 increased in NH females. In a second experiment, rats were fed chow or an HFC diet from Pd30 or 60 until Pd160 and exposed to 1 h restraint before sacrifice. Regardless of neonatal stress, age of diet exposition, or sex, the HFC diet increased body and fat weight and serum leptin concentration; it induced insulinemia in males, but in females only in Pd30 rats. The HFC diet's capacity to curtail the hypothalamus-pituitary-adrenal axis response to restraint was impaired in MS males. In restrained rats, expression of Trh in the paraventricular nucleus of the hypothalamus, Dio2 and Trhde in MBH, and serum thyroid hormone concentration were altered differently depending on sex, age of diet exposition, and neonatal stress. In conclusion, metabolic alterations associated to an HFC-diet-induced obesity are affected by sex or time of exposition, while various parameters of the HPT axis activity are additionally altered by MS, pointing to the complex interplay that these developmental influences exert on HPT axis activity in adult rats.

The Kv1.3 channel blocker Vm24 enhances muscle glucose transporter 4 mobilization but does not reduce body-weight gain in diet-induced obese male rats.

AIMS: Voltage-gated potassium channels 1.3 (Kv1.3) can be targeted to reduce diet-induced obesity and insulin resistance in mice. Since species-specific differences in Kv1.3 expression and pharmacology have been observed, we tested the effect of Vm24, a high-affinity specific blocker of Kv1.3 channels from Vaejovis mexicanus smithi, on body weight (BW), glucose tolerance and insulin resistance in diet-induced obese rats. MATERIALS AND METHODS: Young adult male Wistar rats were switched to a high-fat/high-fructose (HFF) diet. Eighteen days later animals were divided in two groups: vehicle and Vm24 group. Subcutaneous injections were applied every other day until sacrifice 2months later. An additional cohort was maintained on standard chow. KEY FINDINGS: The HFF diet promoted obesity. Treatment with Vm24 did not alter various metabolic parameters such as food intake, BW gain, visceral white adipose tissue mass, adipocyte diameter, serum glucose, leptin and thyroid hormone concentrations, brown adipose tissue mass or uncoupling protein-1 expression, and insulin tolerance. Vm24 did reduce basal and glucose-stimulated serum insulin concentrations, serum C-peptide concentration, increased QUICKI, and tended to lower HOMA-IR. Vm24 treatment did not change the activation of insulin receptor substrate-1, but enhanced protein-kinase B activation and membrane glucose-transporter 4 (GLUT4) protein levels in skeletal muscle. SIGNIFICANCE: In conclusion, in male rats, long-term blockade of Kv1.3 channels with Vm24 does not reduce weight gain and visceral adiposity induced by HFF diet; instead, it reduces serum insulin concentration, and enhances GLUT4 mobilization in skeletal muscle.

Differential Arc protein expression in dorsal and ventral striatum after moderate and intense inhibitory avoidance training.

Intense training refers to training mediated by emotionally arousing experiences, such as aversive conditioning motivated by relatively high intensities of foot-shock, which produces a strong memory that is highly resistant to extinction. Intense training protects memory consolidation against the amnestic effects of a wide variety of treatments, administered systemically or directly into brain structures. The mechanisms of this protective effect are unknown. To determine a potential neurobiological correlate of the protective effect of intense training, rats were trained in a one-trial step-through inhibitory avoidance task using different intensities of foot-shock (0.0, 0.5, 1.0, and 2.0mA). Some rats from each group were sacrificed 45min after training for immunohistochemical Arc protein detection in dorsal and ventral striatum; other rats were tested for extinction during six consecutive days, starting 48h after training. The results showed that training with 1.0 and 2.0mA produced optimal retention scores, which were significantly higher than those of the 0.5 and 0.0mA groups. Also, a higher resistance to extinction was obtained with 2.0mA than with the other intensities. A high number of neurons expressed Arc in ventral, but not in dorsal striatum in both the 1.0 and 2.0mA groups, with a larger area of Arc signal in the latter group. We conclude that an increased Arc expression may be related to enhanced synaptic plasticity in the ventral striatum, suggesting that it may be one of the physiological substrates of enhanced learning.

A screen for modulators reveals that orexin-A rapidly stimulates thyrotropin releasing hormone expression and release in hypothalamic cell culture.

In the paraventricular nucleus of the mammalian hypothalamus, hypophysiotropic thyrotropin releasing hormone (TRH) neurons integrate metabolic information and control the activity of the thyroid axis. Additional populations of TRH neurons reside in various hypothalamic areas, with poorly defined connections and functions, albeit there is evidence that some may be related to energy balance. To establish extracellular modulators of TRH hypothalamic neurons activity, we performed a screen of neurotransmitters effects in hypothalamic cultures. Cell culture conditions were chosen to facilitate the full differentiation of the TRH neurons; these conditions had permitted the characterization of the effects of known modulators of hypophysiotropic TRH neurons. The major end-point of the screen was Trh mRNA levels, since they are generally rapidly (0.5-3h) modified by synaptic inputs onto TRH neurons; in some experiments, TRH cell content or release was also analyzed. Various modulators, including histamine, serotonin, ß-endorphin, met-enkephalin, and melanin concentrating hormone, had no effect. Glutamate, as well as ionotropic agonists (kainate and N-Methyl-d-aspartic acid), increased Trh mRNA levels. Baclofen, a GABAB receptor agonist, and dopamine enhanced Trh mRNA levels. An endocannabinoid receptor 1 inverse agonist promoted TRH release. Somatostatin increased Trh mRNA levels and TRH cell content. Orexin-A rapidly increased Trh mRNA levels, TRH cell content and release, while orexin-B decreased Trh mRNA levels. These data reveal unaccounted regulators, which exert potent effects on hypothalamic TRH neurons in vitro.

Glucocorticoids curtail stimuli-induced CREB phosphorylation in TRH neurons through interaction of the glucocorticoid receptor with the catalytic subunit of protein kinase A.

PURPOSE: Corticosterone prevents cold-induced stimulation of thyrotropin-releasing hormone (Trh) expression in rats, and the stimulatory effect of dibutyryl cyclic-adenosine monophosphate (dB-cAMP) on Trh transcription in hypothalamic cultures. We searched for the mechanism of this interference. METHODS: Immunohistochemical analyses of phosphorylated cAMP-response element binding protein (pCREB) were performed in the paraventricular nucleus (PVN) of Wistar rats, and in cell cultures of 17-day old rat hypothalami, or neuroblastoma SH-SY5Y cells. Cultures were incubated 1h with dB-cAMP, dexamethasone and both drugs combined; their nuclear extracts were used for chromatin immunoprecipitation; cytosolic or nuclear extracts for coimmunoprecipitation analyses of catalytic subunit of protein kinase A (PKAc) and of glucocorticoid receptor (GR); their subcellular distribution was analyzed by immunocytochemistry. RESULTS: Cold exposure increased pCREB in TRH neurons of rats PVN, effect blunted by corticosterone previous injection. Dexamethasone interfered with forskolin increase in nuclear pCREB and its binding to Trh promoter; antibodies against histone deacetylase-3 precipitated chromatin from nuclear extracts of hypothalamic cells treated with tri-iodothyronine but not with dB-cAMP + dexamethasone, discarding chromatin compaction as responsible mechanism. Co-immunoprecipitation analyses of cytosolic or nuclear extracts showed protein:protein interactions between activated GR and PKAc. Immunocytochemical analyses of hypothalamic or SH-SY5Y cells revealed diminished nuclear translocation of PKAc and GR in cells incubated with forskolin + dexamethasone, compared to either forskolin or dexamethasone alone. CONCLUSIONS: Glucocorticoids and cAMP exert mutual inhibition of Trh transcription through interaction of activated glucocorticoid receptor with protein kinase A catalytic subunit, reducing their nuclear translocation, limiting cAMP-response element binding protein phosphorylation and its binding to Trh promoter.

Advances in TRH signaling.

The activity of the hypothalamus-pituitary-thyroid axis (HPT) is coordinated by hypophysiotropic thyrotropin releasing hormone (TRH) neurons present in the paraventricular nucleus of the hypothalamus. Hypophysiotropic TRH neurons act as energy sensors. TRH controls the synthesis and release of thyrotropin, which activates the synthesis and secretion of thyroid hormones; in target tissues, transporters and deiodinases control their local availability. Thyroid hormones regulate many functions, including energy homeostasis. This review discusses recent evidence that covers several aspects of TRH role in HPT axis regulation. Knowledge about the mechanisms of TRH signaling has steadily increased. New transcription factors engaged in TRH gene expression have been identified, and advances made on how they interact with signaling pathways and define the dynamics of TRH neurons response to acute and/or long-term influences. Albeit yet incomplete, the relationship of TRH neurons activity with positive energy balance has emerged. The importance of tanycytes as a central relay for the feedback control of the axis, as well as for HPT responses to alterations in energy balance, and other stimuli has been reinforced. Finally, some studies have started to shed light on the interference of prenatal and postnatal stress and nutrition on HPT axis programing, which have confirmed the axis susceptibility to early insults.

Neonatal Maternal Separation Alters, in a Sex-Specific Manner, the Expression of TRH, of TRH-Degrading Ectoenzyme in the Rat Hypothalamus, and the Response of the Thyroid Axis to Starvation.

Hypothalamic-pituitary-thyroid (HPT) axis activity is important for energy homeostasis, and is modified by stress. Maternal separation (MS) alters the stress response and predisposes to metabolic disturbances in the adult. We therefore studied the effect of MS on adult HPT axis activity. Wistar male and female pups were separated from their mothers 3 h/d during postnatal day (PND)2-PND21 (MS), or left nonhandled (NH). Open field and elevated plus maze tests revealed increased locomotion in MS males and anxiety-like behavior in MS females. At PND90, MS females had increased body weight gain, Trh expression in the hypothalamic paraventricular nucleus, and white adipose tissue mass. MS males had increased expression of TRH-degrading enzyme in tanycytes, reduced TSH and T3, and enhanced corticosterone serum concentrations. MS stimulated brown adipose tissue deiodinase 2 activity in either sex. Forty-eight hours of fasting (PND60) augmented serum corticosterone levels similarly in MS or NH females but more in MS than in NH male rats. MS reduced the fasting-induced drop in hypothalamic paraventricular nucleus-Trh expression of males but not of females and abolished the fasting-induced increase in Trh expression in both sexes. Fasting reduced serum concentrations of TSH, T4, and T3, less in MS than in NH males, whereas in females, TSH decreased in MS but not in NH rats, but T4 and T3 decreased similarly in NH and MS rats. In conclusion, MS produced long-term changes in the activity of the HPT axis that were sex specific; response to fasting was partially blunted in males, which could affect their adaptive response to negative energy balance.

60 YEARS OF NEUROENDOCRINOLOGY: TRH, the first hypophysiotropic releasing hormone isolated: control of the pituitary-thyroid axis.

This review presents the findings that led to the discovery of TRH and the understanding of the central mechanisms which control hypothalamus-pituitary-thyroid axis (HPT) activity. The earliest studies on thyroid physiology are now dated a century ago when basal metabolic rate was associated with thyroid status. It took over 50 years to identify the key elements involved in the HPT axis. Thyroid hormones (TH: T4 and T3) were characterized first, followed by the semi-purification of TSH whose later characterization paralleled that of TRH. Studies on the effects of TH became possible with the availability of synthetic hormones. DNA recombinant techniques facilitated the identification of all the elements involved in the HPT axis, including their mode of regulation. Hypophysiotropic TRH neurons, which control the pituitary-thyroid axis, were identified among other hypothalamic neurons which express TRH. Three different deiodinases were recognized in various tissues, as well as their involvement in cell-specific modulation of T3 concentration. The role of tanycytes in setting TRH levels due to the activity of deiodinase type 2 and the TRH-degrading ectoenzyme was unraveled. TH-feedback effects occur at different levels, including TRH and TSH synthesis and release, deiodinase activity, pituitary TRH-receptor and TRH degradation. The activity of TRH neurons is regulated by nutritional status through neurons of the arcuate nucleus, which sense metabolic signals such as circulating leptin levels. Trh expression and the HPT axis are activated by energy demanding situations, such as cold and exercise, whereas it is inhibited by negative energy balance situations such as fasting, inflammation or chronic stress. New approaches are being used to understand the activity of TRHergic neurons within metabolic circuits.

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.