Exogenous TRH Research Articles

Cardiac Delivery of Interference RNA for Thyrotropin-Releasing Hormone Inhibits Hypertrophy in Spontaneously Hypertensive Rat

See related article, pp 103–109 Thyrotropin-releasing hormone (TRH) is a tripeptide synthesized in the paraventricular nucleus of the hypothalamus and is best known for its classic role in stimulating the release of thyroid-stimulating hormone (TSH) and prolactin from the anterior pituitary gland. By stimulating the release of TSH, TRH regulates thyroxine (T4) and triiodothyronine (T3) production and secretion into the bloodstream. The presence of a TRH system in the paraventricular and preoptic nuclei of the hypothalamus suggests a direct role of TRH in cardiovascular physiology. Indeed, blood pressure (BP), heart rate, and contractility increase after intracerebroventricular1 or intravenous2 administration of TRH in rats. Although administration of exogenous TRH at pharmacological doses has helped to identify the potential roles of TRH, further elucidation of endogenous TRH function and mediation by its 2 receptors in rodents (in humans, only 1 TRH receptor has been identified thus far) have been hampered because of the lack of receptor subtype-selective antagonists. Furthermore, these observations are consistent with the fact that the heart is a major target organ for thyroid hormones, and that there are marked changes in cardiac function and structure in patients with hypothyroidism or hyperthyroidism.3 Spontaneously hypertensive rats (SHR) are used as a model of human essential hypertension. It presents an increase in both TRH content and TRH precursor mRNA abundance in the preoptic area, with a higher cerebrospinal fluid TRH concentration and TRH receptor number in this area.4 This model demonstrates a gradual progression of myocardial hypertrophy, fibrosis, left ventricular (LV) dysfunction, and heart failure. Persistent hypertension begins when rats reach ≈2 months of age and is followed by a relatively long period of stable, compensated hypertrophy. By 18 to 24 months, male SHR develop heart failure and exhibit a marked upregulation of genes that encode …

Corticotropin releasing hormone (CRH), melanocortins (MC) and thyrotropin releasing hormone (TRH) stimulate pigmentation and melanocyte function in the human hair follicle

Melanocortins (MC) such as adrenocorticotropic hormon (ACTH) and α‐melanocyte‐stimulating hormone (αMSH) are neuropeptides derived from the prohormone proopiomelanocortin (POMC). It is now well established that the human skin is not only a target but also a local source for MCs. Almost all cutaneous cell types express MCs and MC‐receptor (MCRs) in vitro. The POMC peptides, their receptors and CRH the main inducer of POMC gene/protein expression in the pituitary and in skin ‐were also detected in human scalp hair follicles (HFs). MCs regulate pigmentation: stimulate melanogenesis, dendricity and proliferation in epidermal melanocytes, and similar effect was reported recently on cultured melanocytes derived from the outer root sheath of human HFs. TRH is the most proximal member of the hypothalamic‐pituitary‐thyroid (HPT) axis. In amphibian skin, TRH also regulates POMC release, which leads to skin darkening via αMSH‐MC1R interactions. Recently we found TRH and TRH‐R expression in human HFs. Therefore, we examined the – as yet ill‐explored – effect of exogenous CRH, ACTH, αMSH and TRH on melanin production, melanocytes number and dendricity in normal, microdissected, organ‐cultured human anagen scalp HFs. Melanin production was measured by quantitaive Masson‐Fontana histochemistry, and melanocyte dendricity were assessed by immunohistochemistry against the melanosome marker gp100 (Nki‐beteb). The melanin content increased significantly after αMSH (10–100 nM) and TRH(1–10 ng/ml) addition to the medium. Total Nki‐beteb immunoreactivity was significantly elevated in the HF melanocytes treated with αMSH, thus confirming the Masson‐Fontana results. TRH, CRH (10 nM–100 pM) and MC (10–100 nM) also stimulated dendrite formation of HF pigmentary unit melanocytes. CRH, MCs and TRH as well were able to compensate the significant decline of the melanin content caused by catagen induction with TGFβ2. This provides unequivocal evidence that CRH, ACTH and a‐MSH stimulate normal human hair follicle pigmentation in situ. We also show for first time that TRH, a key regulator of pigmentation in amphibian skin, might play similar role in human HF.

Tissue Deiodinase Activity during Prolonged Critical Illness: Effects of Exogenous Thyrotropin-Releasing Hormone and Its Combination with Growth Hormone-Releasing Peptide-2

Prolonged critical illness is characterized by reduced pulsatile TSH secretion, causing reduced thyroid hormone release and profound changes in thyroid hormone metabolism, resulting in low circulating T(3) and elevated rT(3) levels. To further unravel the underlying mechanisms, we investigated the effects of exogenous TRH and GH-releasing peptide-2 (GHRP-2) in an in vivo model of prolonged critical illness. Burn-injured, parenterally fed rabbits were randomized to receive 4-d treatment with saline, 60 microg/kg.h GHRP-2, 60 microg/kg.h TRH, or 60 microg/kg.h TRH plus 60 microg/kg.h GHRP-2 started on d 4 of the illness (n = 8/group). The activities of the deiodinase 1 (D1), D2, and D3 in snap-frozen liver, kidney, and muscle as well as their impact on circulating thyroid hormone levels were studied. Compared with healthy controls, hepatic D1 activity in the saline-treated, ill animals was significantly down-regulated (P = 0.02), and D3 activity tended to be up-regulated (P = 0.06). Infusion of TRH and TRH plus GHRP-2 restored the catalytic activity of D1 (P = 0.02) and increased T(3) levels back within physiological range (P = 0.008). D3 activity was normalized by all three interventions, but only addition of GHRP-2 to TRH prevented the rise in rT(3) seen with TRH alone (P = 0.02). Liver D1 and D3 activity were correlated (respectively, positively and negatively) with the changes in circulating T(3) (r = 0.84 and r = -0.65) and the T(3)/rT(3) ratio (r = 0.71 and r = -0.60). We conclude that D1 activity during critical illness is suppressed and related to the alterations within the thyrotropic axis, whereas D3 activity tends to be increased and under the joint control of the somatotropic and thyrotropic axes.

Thyrotropin-releasing hormone (protirelin) inhibits potassium-stimulated glutamate and aspartate release from hippocampal slices in vitro

Excess excitatory amino acid release is involved in pathways associated with seizures and neurodegeneration. Thyrotropin-releasing hormone (TRH; protirelin), a brain-derived tripeptide, has shown efficacy in the treatment of such disorders, yet its mechanism of neuroprotection is poorly understood. Using superfused hippocampal slices, we tested the hypothesis that TRH could inhibit evoked glutamate/aspartate release in vitro. Rat hippocampal slices were first equilibrated in oxygenated Krebs buffer (KRB) (120 min) then superfused for 10 min with KRB (control), or KRB containing 0.1, 1, or 10 μM TRH respectively, prior to and during 5 min depolarization with high potassium KRB (50 mM [K +] ± TRH). Fractions (1 min) were collected during the 5 min stimulation and for an additional 10 min thereafter and analyzed for glutamate and aspartate by HPLC. TRH had no effect on baseline glutamate/aspartate release, while all three TRH doses significantly ( P < 0.05) inhibited peak 50 mM [K +]-stimulated glutamate/aspartate release, and glutamate remained below control ( P < 0.05) at 15 min post stimulation. A 5 min pulse of TRH (10 μM) had no affect on basal glutamate/aspartate release, whereas the TRH pre-pulsed slices failed to release glutamate/aspartate by [K +]-stimulation given 15 min later. These results are the first to show a potent and prolonged inhibitory effect of TRH on evoked glutamate/aspartate release in vitro. These initial studies suggest that exogenous and/or endogenous TRH may function, in part, to modulate excess glutamate release in specific CNS loci. Additional studies are in progress to fully understand the mechanism of this potent effect of TRH and its implication in various CNS disorders.

Abnormalities of hypothalamic-pituitary-thyroid axis in patients with primary empty sella.

Primary empty sella (PES) is a very frequent neuroradiological finding in the general population, that can induce hypopituitarism. Some studies focused on the association of PES with GH deficiency (GHD) or hypogonadotropic hypogonadism (HH), while data regarding the involvement of hypothalamic-pituitary-thyroid (HPT) axis, despite sporadic reports of central hypothyroidism, or the occurrence of hypoadrenalism (HA) are scanty. In this study, thyroid function and TSH response to exogenous TRH injection (TRH/TSH) were investigated in 43 patients [10 men and 33 women; aged (mean +/- SD), 48+/-12 yr] with PES: 22 patients had total and 21 partial PES. Forty healthy subjects (9 men and 31 women; aged 46+/-12 yr) were enrolled as a control group. Central hypothyroidism was found only in 2/43 cases, whereas one patient showed primary hypothyroidism. In euthyroid patients, mean serum TSH levels were significantly lower than controls (TSH: 1.0+/-0.7 vs 1.4+/-0.6 mU/l, p<0.01) and 79% of them showed abnormal TRH/TSH responses (TRH test was performed in 34 euthyroid patients: 17 cases with total and 17 cases with partial PES), but mean serum free T4 (FT4) and free T3 (FT3) values were not significantly lower than controls (FT4: 15.9+/-0.4 vs 15.0+/-2.1 pmol/l, p=NS; FT3: 5.3+/-1.2 vs 5.8+/-1.5 pmol/l, p=NS). Moreover, no significant differences were evident in mean serum TSH, FT4 and FT3 between patients with total and partial PES (TSH: 1.1+/-0.7 vs 0.9+/-0.8 mU/l, p=NS; FT4: 16.3+/-2.6 vs 15.7+/-2.2 pmol/l, p=NS; FT3: 5.4+/-1.3 vs 5.2+/-0.8 pmol/l, p=NS) and the TRH/TSH peak was impaired or exaggerated/delayed in 9 and 3 patients with total and in 12 and 3 cases with partial PES. No significant differences in the prevalence of abnormal TRH/TSH responsiveness were found between patients with partial or total PES (chi2=1.6, p=NS). Other impairment of pituitary function was detected in 23/43 patients: GHD was present in 15 cases, HH in 11 and central HA in 5 patients. Isolated or combined hypopituitarism was present in 17 and in 6 patients, respectively. In conclusion, pituitary dysfunction is very frequent in patients with PES, but central hypothyroidism occurs rarely. The entity of arachnoid herniation into the sellar fossa does not play a significant role on the degree of HPT axis dysfunction.

Evidence suggesting that cadmium induces a non-thyroidal illness syndrome in the rat.

The effect of in vivo administration of cadmium chloride on the pituitary-thyroidal axis was assessed in 200 g body weight Wistar rats. A dose of 2.5 mg/kg body weight was injected i.v. 24 h before the experiments were initiated. Plasma thyroxine (T4) and tri-iodothyronine (T3) concentrations in cadmium-treated rats were significantly (P < 0.01) decreased, whereas plasma TSH failed to increase in response to low T4 and T3. However, the TSH response to TRH and the pituitary content of TSH in these rats were both normal. Cadmium induced a significant (P < 0.01) decrease in 4-h thyroidal 131I uptake and in thyroid/plasma radioactivity ratio. The in vitro conversion of T4 to T3 in the pituitary was significantly (P < 0.01) blocked by cadmium whereas there was no in vivo effect. Parameters of peripheral T4 kinetics in cadmium-treated rats, such as metabolic clearance rate (P < 0.01), fractional turnover rate (P < 0.01), absolute disposal rate (P < 0.05), urinary clearance (P < 0.05) and faecal clearance (P < 0.05), were all decreased by cadmium. The lack of response of TSH to low plasma T4 and T3 and the normal response to exogenous TRH in this and in other non-thyroidal illness syndromes produced by other pathologies suggest a decreased stimulation of pituitary thyrotrophs by endogenous TRH.

THYROID HORMONES AND PROLACTIN IN PREMATURE INFANTS WITH AND WITHOUT RDS AFTER PREANTAL TRH TREATMENT. † 979

Since 1989 TRH plus glucocorticoids combination has been studying in humans. Aim: to evaluate the incidence and severity of RDS and chronic lung disease (CLD) after prenatal TRH plus betamethasone treatment. Hormonal studies included free T3 (FT3) [nd/dl], free T4 (FT4) [ng/dl], TSH[μU/ml] and prolactin (PRL) [ng/ml]. The correlation between RDS and thyroid function and PRL was analysed. Methods: study group (TRH+B) 58 newborns with mean 31.9 wga and mean birth weight 1795g. Control group(B) 68 neonates of 31.5 mean wga and 1714g mean birthweight. Hormonal analysis was performed on cord blood, and day 1 and 7 of life. For statistical analysis, Statistica 4.5 for Windows was used. Results: 1) no statistical differences in mean TSH level was found in any sample between study and control group (10.70 vs 10.62 in cord; 7.34 vs 5.48 after day 1; 3.33 vs 4.38 after day 7). 2) FT3 concentration did not differ in particular samples for TRH +B and B group (0.22 vs 0.25 in cord; 0.33 vs 0.31 after day 1 and 0.34 vs 0.33 after day 7). 3) There was statistically significant difference in FT4 level, in cord blood (1.26 in TRH +B vs 1.18 in B group; p< 0.05), but not in other samples (1.64 v. 1.63 after day 1 and 1.42 vs 1.28 after day 7). 4) PRL level was similar in study and control goup in each succeeding samples (97.07 vs 112.59; 108.88 vs 93.27; 73.64 vs 72.58). RDS and thyroid function. 5) Mean TSH level in cord blood was higher in group with RDS (12.90 vs 9.28). On day 1 it was lower in RDS group (5.21 vs 7.27; p < 0.01). On day 7 there was no statistical difference between both groups (4.28 in RDS and 2.85 in control group). 6) FT3 level was lower in RDS than in control group in each sample (0.23 vs 0.26; 0.24 * vs 0.39*; 0.31 vs 0.37; *p <0.001). 7) FT4 concentration was slightly higher in RDS group in cord blood (1.24 vs 1.19). On day 1 and 7 FT4 level was higher in control group (1.68 vs 1.59 and 1.44 vs 1.26). 8) Prolactin level was lower in RDS group in each following samples: cord blood; 99.82 vs 113.62; day 1: 79.55 vs 116.60, p<0.01; and day 7: 66.43 vs 81.02. Conclusions: fetal pituitary-thyroid axis is sensitive to exogenous TRH. Low thyroid hormones concentration ininfants with RDS may have an important role in protectig sick neonate from higher oxygen consumption in situations when it's delivery to organs and tissues is imparied. The rate of T4 to T3 conversion is low in these situations, with overproduction of rT3. We suggest that hormonal disturbances in infants with RDS are transient and contribute to a low T3 syndrome.

Effect of haemoglobin and endogenous erythropoietin on hypothalamic-pituitary thyroidal and gonadal secretion: an analysis of anaemic (high EPO) and polycythaemic (low EPO) patients.

Correction of anaemia with recombinant human erythropoietin (rHu-EPO) improves the responsiveness of thyroidal and gonadal axes to exogenous TRH and GnRH in chronic haemodialysis patients, but the mechanisms remain to be fully elucidated. In order to assess the influences of endogenous erythropoietin on the hypothalamo-hypophyseal thyroidal and gonadal axes, we studied the response of polycythaemic and anaemic patients, in comparison to normal controls, after the administration of exogenous TRH and GnRH. Exogenous hypothalamic factors, 500 micrograms TRH and 100 micrograms GnRH, were administered as a bolus and blood samples were obtained over a 3-hour period at 30, 60, 90, 120 and 180 minutes. Five male polycythaemic patients (low EPO), three male anaemic patients (high EPO) and six normal age and sex matched controls were studied. Blood samples were centrifuged immediately and the serum was stored at -20 degrees C until assayed for total T4, free T4, free T3, TSH, prolactin, growth hormone (TRH test), and FSH, LH, testosterone (GnRH test). Haematological parameters and biochemical profiles were also measured. After TRH administration, both patient groups showed a normal TSH response; however, their free T4 and free T3 secretion was blunted compared to controls. Normal basal PRL levels increased in an exaggerated fashion, whereas, when compared to chronic renal failure patients on chronic haemodialysis, we did not see a paradoxical GH response or a basal GH increase in these 5 patients. GnRH administration in the study groups elicited a normalization in the LH response without an increase in testosterone levels; however, an exaggerated FSH response was found in the polycythaemic patients (low EPO). Thus by investigating the role of low endogenous EPO levels in non-anaemic and anaemic patients with high EPO levels, our study suggests that the underlying chronic disease state may be the major contributing factor in the regulation of the hypothalamo-hypophyseal thyroid and gonadal axes, rather than the EPO levels.

Antithyrotropin-releasing hormone serum inhibits secretion of glucagon from isolated perfused rat pancreas: an experimental model for positive feedback regulation of glucagon secretion.

TRH is synthesized in the islets of Langerhans and was found in the perfusate of isolated rat pancreas. In the present study, designed to determine the role of endogenous TRH, we first characterized chromatographically the identity of immunoreactive TRH with synthetic pGlu-His-Pro-NH2. Since endogenous TRH secretion may mask the effects of exogenous TRH, we performed, in parallel to dose-response studies, immunoneutralization experiments using anti-TRH serum to neutralize the endogenous TRH secretion from isolated perfused rat pancreas. The data indicate that exogenous TRH enhances basal glucagon secretion; inversely, anti-TRH serum inhibits glucose plus arginine-induced glucagon secretion and produces a concomitant slight inhibition of somatostatin secretion. The present study shows a physiological contribution for endogenous TRH as a local modulator of intraislet hormone regulation; from these observations, we postulate a direct effect of pancreatic TRH on glucagon-containing (alpha) cell secretion, which, in turn, may produce the fluctuation in somatostatin secretion. Local TRH secretion provides a model for positive feedback regulation of glucagon secretion, frequently associated with diabetes.

Differential Responsiveness of the Pituitary-Thyroid Axis to Thyrotropin-Releasing Hormone in Mouse Lines Selected to Differ in Central Nervous System Sensitivity to Ethanol*

Long-sleep (LS) and short-sleep (SS) mice are genetic lines that differ in central nervous system sensitivity to ethanol. The possible role of TRH in mediating the difference in the thyroid status between these two lines was investigated. An increase in TRH gene expression in the paraventricular nucleus and TRH peptide levels in the hypothalamus between postnatal days 8-14 in both SS and LS mice coincided with increased circulating levels of thyroxine during this critical period of central nervous system development. No significant differences in TRH biosynthesis were observed between LS and SS mice during this time. Exogenous administration of TRH to LS and SS mice on day 8, when endogenous serum thyroxine levels were equivalent, resulted in a greater increase in serum thyroxine in SS mice (150%) than LS mice (51%). The differential response to the TRH stimulation test was also present on day 14 (SS, 43%; LS, 18%). The differential responsiveness of the pituitary-thyroid axis to exogenous TRH paralleled the differential increase in endogenous serum thyroxine observed between day 8 and 14 in these mice. Administration of TRH to day 20 and adult (60 days) LS and SS mice resulted in nearly equivalent (approximately 75%) increases in free thyroxine serum levels, yet the magnitude of thyroxine release was 50% greater in SS mice, due perhaps to between-line differences within the thyroid glands. It is unlikely that dissimilar endogenous levels of TRH account for the intrinsic difference in the thyroid status in LS and SS mice. Instead, the increased pituitary-thyroid responsiveness to TRH in SS mice during the second postnatal week may translate into increased functional capacity of the thyroid gland in adult SS relative to LS mice.

Relationships among LH, FSH and prolactin secretion, storage and response to secretagogue and hypothalamic GNRH content in ovariectomized pony mares administered testosterone, dihydrotestosterone, estradiol, progesterone, dexamethasone or follicular fluid

Thirty-five ovariectomized pony mares were used to study the relationships among luteinizing hormone (LH), follicle stimulating hormone (FSH) and prolactin (PRL) concentrations in blood (secretion), in pituitary (storage) and in blood after secretagogue administration, as well as the content of gonadotropin releasing hormone (GnRH) in hypothalamic areas, under various conditions of steroidal and nonsteroidal treatment. Five mares each were treated daily for 21 d with vegetable shortening (controls), testosterone (T; 150 μg/kg of body weight, BW), dihydrotestosterone (DHT; 150 μg/kg BW), estradiol (E 2; 35 μg/kg BW), progesterone (P 4; 500 μg/kg BW), dexamethasone (DEX; 125 μg/kg BW) or charcoal-stripped equine follicular fluid (FF; 10 ml). Secretagogue injections (GnRH and thyrotropin releasing hormone, TRH, at 1 and 4 μg/kg of BW, respectively) were given one d prior to treatment and again after 15 d of treatment. Relative to controls, treatment with T, DHT and DEX reduced (P <. 05) LH secretion, storage and response to exogenous GnRH, whereas treatment with E 2 increased (P < .05) these same characteristics. Treatment with P 4 reduced (P < .05) only LH secretion. Treatment with T, DHT, E 2 and DEX reduced (P < .05) FSH secretion, whereas treatment with P 4 increased (P < .05) it and FF had no effect (P > .1). All treatments increased (P < .05) FSH storage, whereas only treatment with T and DHT increased (P < .05) the FSH response to exogenous GnRH. Other than a brief increase (P < .05) in PRL secretion in mares treated with E 2, secretion of PRL did not differ (P > .1) among groups. Only treatment with E 2 increased (P < .01) PRL storage, yet treatment with T or DHT (but not E 2) increased (P < .05) the PRL response to exogenous TRH. Content of GnRH in the body and pre-optic area of the hypothalamus was not affected (P > .1) by treatment, whereas treatment with T, E 2 and DEX increased (P < .1) GnRH content in the median eminence. For LH, secretion, storage and response to exogenous GnRH were all highly correlated (r ≥ .77; P < .01). For FSH, only storage and response to exogenous GnRH were related (r = .62; P < .01). PRL characteristics were not significantly related to one another. Moreover, the amount of GnRH in the median eminence was not related (P > .1) to any LH or FSH characteristic.

Serum Pattern of Thyroxine (T4), 3,3‘,5‐Triiodothyronine (T3) and 3,3’,5‘‐Triiodothyronine (rT3) in Fed and Fasted Cocks Following TRH Stimulation

Food deprivation for 27 and 75h in cocks decreased serum levels of T3 maximally by 61.1% and increased the level of rT3 by 51.4% and that of T4 by 22.4%. Injection of a single dose of TRH (10 micrograms/kg b.w.) increased serum levels of all 3 iodothyronines in both fed and food-deprived animals. In fasted, TRH-treated birds the peak of serum rT3 was 80.5 and 73.3% above control levels in 27- and 75-hr food-deprived cocks, respectively (fed birds: 14.6 and 9.7%); the relevant data for T3 were: 78.3 and 40.4% (fed birds: 95.7 and 127.5%); for T4: 30.0 and 27.0% (fed birds: 9.2 and 6.1%). The greater relative increment of serum rT3 than T3 in food-deprived and TRH-treated cocks was supported by a fall of the serum T3/rT3 ratio to 78.7 and 27.9% in fed cocks and 27- and 75-hr fasted animals, respectively. We conclude, that food deprivation modifies the response of the hypophysis-thyroid axis to exogenous TRH in the sense that during fasting the production of rT3 is enhanced relative to T3.

Hypothalamic regulation of pulsatile thyrotopin secretion.

To determine the mechanism underlying pulsatile TSH secretion, 24-h serum TSH levels were measured in three groups of five healthy volunteers by sampling blood every 10 min. The influence of an 8-h infusion of dopamine (200 mg), somatostatin (500 micrograms), or nifedipine (5 mg) on the pulsatile release of TSH was tested using a cross-over design. The amount of TSH released per pulse was significantly lowered by these drugs, resulting in significantly decreased mean basal TSH serum levels. However, pulses of TSH were still detectable at all times. The TSH response to TRH (200 micrograms) tested in separate experiments was significantly lowered after 3 h of nifedipine infusion compared to the saline control value. Nifedipine treatment did not alter basal, pulsatile, or TRH-stimulated PRL secretion. The persistence of TSH pulses under dopamine and somatostatin treatment and the blunted TSH responses to nifedipine infusion support the hypothesis that pulsatile TSH secretion is under the control of hypothalamic TRH. The 24-h TSH secretion pattern achieved under stimulation with exogenous TRH in two patients with hypothalamic destruction through surgical removal of a craniopharyngioma provided further circumstantial evidence for this assumption. No TSH pulses and low basal TSH secretion were observed under basal conditions (1700-2400 h), whereas subsequent repetitive TRH challenge (25 micrograms/2 h to 50 micrograms/1 h) led to a pulsatile release of TSH with fusion of TSH pulses, resulting in a TSH secretion pattern strikingly similar to the circadian variation. These data suggest that pulsatile and circadian TSH secretions are predominantly controlled by TRH.

Anterolateral Hypothalamic Deafferentation Inhibits Histamine-Induced Prolactin Secretion and Potentiates TRH-Induced Thyrotropin Secretion in Male Rats

We studied the effect of histamine on serum prolactin and thyrotropin (TSH) levels in male rats with anterolateral hypothalamic deafferentation of hypothalamic connections or anterolateral cut (ALC). The success of ALC was confirmed by immunohistochemistry of somatostatin (SRIF) in the medial basal hypothalamus. ALC did not affect basal prolactin or TSH levels. Thyrotropin-releasing hormone (TRH, 200 ng/rat, i.p.) did not affect prolactin secretion either in sham-operated or ALC rats. In sham-operated rats intracerebroventricularly administered histamine increased significantly prolactin levels. Hypothalamic deafferentation abolished the effect of histamine on prolactin levels. TRH increased significantly serum TSH levels both in sham-operated controls and ALC rats. In the latter, however, the TSH-secretory response to TRH was significantly (p less than 0.05) larger compared to the controls. Intracerebroventricularly infused histamine (2 micrograms/rat) did not change the TRH-induced TSH secretion in either group of rats. These results show that (1) the effect of histamine on prolactin secretion is mediated through nerve tracts which are destroyed by ALC, and (2) cutting of afferent TRH (through sensitization) and SRIF fibers (through lacking inhibition) entering medial basal hypothalamus may both contribute to the enhanced TSH response to exogenous TRH.

Thyroid Hormones Modulate Thyrotropin-Releasing Hormone Biosynthesis in Tissues Outside the Hypothalamic-Pituitary Axis of Male Rats*

In the present study we have examined the in vivo effects of thyroid hormone and TRH on secretory tissue concentrations of TRH and TRH-Gly (pGlu-His-Pro-Gly), a TRH precursor. Within secretory granules, TRH-Gly is converted to TRH through alpha-amidation of the C-terminal proline residue, using Gly as the NH2 donor. Using specific RIA, we measured the TRH-Gly immunoreactivity (TRH-Gly-IR) and TRH-IR concentrations in tissues from the reproductive and gastrointestinal systems, adrenals, and other internal organs in euthyroid, hypothyroid, and T4-treated 250-g Sprague-Dawley male rats. TRH-Gly-IR concentrations were more than 2-fold higher than TRH-IR concentrations within the adrenal, pancreas, bowel, and stomach at the time of death. Untreated hypothyroidism and exogenous TRH significantly increased adrenal TRH-Gly-IR levels. Pancreatic TRH-Gly levels increased about 2-fold in hypothyroid rats. Incubation at 60 C significantly increased TRH-Gly-IR levels in the pancreas, adrenal, bowel, stomach, and epididymis by 14-, 3-, 6-, 6-, and 6-fold, respectively. Also after 60 C incubation increases in the TRH-Gly-IR/TRH-IR ratio of 2.7-, 4-, and 1.7-fold were observed in the pancreas, epididymis, and bowel, respectively. Pooled tissue extracts were fractionated by cation exchange and reverse phase HPLC for characterization of TRH-Gly-IR. Both chromatographic methods revealed a major peak of TRH-Gly-IR coeluting with synthetic TRH-Gly. Incubation at 60 C caused 13.5-, 4.1-, 1.5-, and 5-fold increments in the TRH-Gly-IR for adrenal, pancreas, prostate, and thyroid, respectively, compared to the immediately extracted control aliquots. Cation exchange and reverse phase HPLC also revealed production of higher mol wt TRH precursor peptides after incubation at 60 C for 4 or 20 h. Only the TRH-Gly-IR peak coeluting with pGlu-His-Pro-Gly was converted into TRH by rat brain alpha-amidating enzyme. The data suggest that biosynthesis of TRH occurs in rat extrahypothalamic tissues and may be modulated by thyroid status, iv TRH, and selective thermal inactivation of enzymes that convert prepro-TRH to TRH.

Physiological regulation of thyrotropin

The pattern of TSH secretion in man is pulsatile in addition to the well known circadian variation. The mechanism triggering TSH pulses remains unclear to date. Infusions of somatostatin or dopamine rapidly lowering basal TSH levels without suppressing the pulsatile pattern suggest that an episodic disinhibition exerted by a physiological inhibitor is not a likely cause. On the same basis, thyroid hormones do not appear to be candidates, since they similarly inhibit basal TSH levels after a time lag of several hours but again do not suppress pulsatile release of the hormone. In contrast, bolus injections of dexamethasone completely abolish pulsatile release of TSH for several hours despite a normal sensitivity of the pituitary to exogenous TRH, suggesting a hypothalamic action of the drug. The hypothesis that pulsatile TSH release might be governed by a pulsatile mode of a hypothalamic stimulator is supported by the observation that an infusion of nifedipine, a calcium channel blocker, which in vitro selectively inhibits the TRH effect on TSH butnot prolactin secretion, exerts a comparable effect when it is infused in vivo.

Islet secretion of immunoreactive thyrotropin-releasing hormone and the 'paracrine-like' effects of its exogenous administration.

In order to know more about the secretory pattern of islet TRH in response to glucose and its possible physiological relevance, the release of this hormone as well as that of insulin, glucagon, and somatostatin was radioimmunologically measured. Whereas the secretion of immunoreactive insulin and somatostatin by incubated rat islets is known to be dose-dependently stimulated by glucose, that of glucagon and TRH was inhibited by glucose. Similarly, palmitate dose-dependently inhibited islet glucagon and TRH release. Exogenous TRH exerted strong and dose-dependent effects on islet secretion of the other hormones at the same concentration range at which its hypophysiotropic effects are produced (10(-10) to 10(-8) mol/l). It inhibited the insulin response to glucose and blocked that of glucagon, whereas it enhanced glucose-induced stimulation of somatostatin. These results are suggestive of a possible paracrine inhibitory role of islet TRH, either directly exerted on the secretion of insulin and glucagon or partially mediated through the stimulation of somatostatin release.

Absence of receptors for thyrotropin (TSH)-releasing hormone in human TSH-secreting pituitary adenomas associated with hyperthyroidism.

In patients with TSH-secreting pituitary adenomas associated with hyperthyroidism, TSH secretion usually does not respond to exogenous TRH stimulation. To determine the basis for this unresponsiveness, we studied TRH binding to the membranes of two such TSH-secreting pituitary adenomas. The patients, a 28-yr-old man and a 60-yr-old woman, had clinical and biochemical hyperthyroidism with increased serum TSH levels (15.7 and 14 mU/L, respectively; normal range, 1.1-7.2) and alpha-subunit to TSH molar ratios greater than 1 (2.4 and 1.7, respectively). Neither patients had an increase in serum TSH in response to TRH (200 micrograms, iv). Immunocytochemistry of the two adenomas, removed by transsphenoidal surgery showed a pure population of thyrotropic cells. Binding studies were performed by incubation of tumor cell membrane suspensions with increasing amounts of [3H]TRH. Two PRL-secreting adenomas and one normal human pituitary were used as controls. The characteristics of the TRH-binding sites from the control tissues were similar to those previously reported (Kd, 56, 30, and 49 nM; maximum binding, 187, 46, and 94 fmol/mg protein, respectively). In contrast, no specific TRH binding was found in the two TSH-secreting adenomas. We conclude that the unresponsiveness of TSH after TRH administration is related to the absence of specific TRH-binding sites in these thyrotropic tumors.

Assessment of the relationship between serum thyroid hormone levels and peripheral metabolism in patients with anorexia nervosa.

It has been observed that basal and/or TRH-stimulated serum TSH levels occasionally conflict with the actual values of circulating thyroid hormones in patients with anorexia nervosa. In the present study sixteen female patients with anorexia nervosa during self-induced starvation displayed clinical findings suggesting hypothyroidism, e.g., cold intolerance, constipation, bradycardia, hypothermia and hypercholesterolemia in association with decreased serum total T3 (62.8 +/- 5.2 ng/dl) and T4 (6.6 +/- 0.3 micrograms/dl). Markedly decreased T3 correlated positively with average heart rate (r = 0.5655, P less than 0.025) and negatively with total cholesterol (r = -0.7413, P less than 0.005). This result may suggest that peripheral metabolic state of the underweight anorexics depends considerably upon the serum T3 concentration. Despite decreased total thyroid hormones, free T4 assayed by radioimmunoassay was normal in all five cases examined (1.4 +/- 0.2 ng/dl) and the free T4 index in fifteen cases was normal except in one case. Basal TSH was not increased and TSH response to exogenous TRH was not exaggerated in any. These results may be compatible with a theory that free T4 has a dominant influence on pituitary TSH secretion. Furthermore, glucocorticoids may also have some influence on depressed TSH response, because an inverse correlation between increased plasma cortisol and the sum of net TSH increase after TRH was observed in twelve cases examined. In conclusion, it is suggested that normal sensitivity of peripheral tissues and pituitary thyrotroph to different circulating thyroid hormones is maintained in anorexia nervosa patients even during severe self-induced starvation, and that the metabolic state in these patients is considerably under the influence of circulating T3.

The pituitary TSH response to TRH is inversely related to the plasma TSH concentration and directly related to the pituitary TSH content during hypothyroidism in the rat.

We studied the effects of degree and duration of hypothyroidism on the pituitary TSH concentration and the pituitary TSH secretory response to TRH. Varying degrees of hypothyroidism were achieved in thyroparathyroidectomized rats (THYREX) by continuous sc infusion of T3 (0.2, 0.3, 0.4, or 0.5 microgram/100 g X day) or T4 (0.6, 1.2, or 1.8 microgram/100 g X day). While T3 was more potent than T4, both resulted in a dose-dependent suppression of the post-thyroidectomy rise in TSH. After 7 or 14 days of severe hypothyroidism (nonreplaced THYREX rats) the pituitary TSH secretory response to TRH (250 ng/100 g body weight, iv) was found to be decreased when compared to that of euthyroid rats. Decreasing the degree of hypothyroidism increased the pituitary secretory response to TRH and the pituitary TSH content. The results indicate that in the hypothyroid rat: severe hypothyroidism results in a blunted pituitary TSH response to TRH through 14 days after thyroidectomy, at 7 and 14 days after thyroidectomy the pituitary TSH response to exogenous TRH is inversely related to the basal plasma TSH concentration, the pituitary TSH concentration increases with the duration of hypothyroidism, the pituitary TSH content is increased by low rates of thyroid hormone replacement, and the pituitary TSH response to exogenous TRH is directly related to the pituitary TSH content.