Thyrotropin-releasing Hormone In Mice Research Articles

Cardiovascular and body weight regulation changes in transgenic mice overexpressing thyrotropin-releasing hormone (TRH).

Thyrotropin-releasing hormone (TRH) plays several roles as a hormone/neuropeptide. Diencephalic TRH (dTRH) participates in the regulation of blood pressure in diverse animal models, independently of the thyroid status. The present study aimed to evaluate whether chronic overexpression of TRH in mice affects cardiovascular and metabolic variables. We developed a transgenic (TG) mouse model that overexpresses dTrh. Despite having higher food consumption and water intake, TG mice showed significantly lower body weight respect to controls. Also, TG mice presented higher blood pressure, heart rate, and locomotor activity independently of thyroid hormone levels. These results and the higher urine noradrenaline excretion observed in TG mice suggest a higher metabolic rate mediated by sympathetic overflow. Cardiovascular changes were impeded by siRNA inhibition of the diencephalic Trh overexpression. Also, the silencing of dTRH in the TG mice normalized urine noradrenaline excretion, supporting the view that the cardiovascular effects of TRH involve the sympathetic system. Overall, we show that congenital dTrh overexpression leads to an increase in blood pressure accompanied by changes in body weight and food consumption mediated by a higher sympathetic overflow. These results provide new evidence confirming the participation of TRH in cardiovascular and body weight regulation.

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

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

Potentiation of cyclo (N-methyl-Tyr-Arg)-induced antinociceptive activity by thyrotropin-releasing hormone in mice.

The effect of thyrotropin-releasing hormone (TRH) and its metabolite, cyclo(His-Pro) (C.HP), on cyclo(N-methyl-Tyr-Arg) (C.NMTA)-induced antinociception as measured by the tail-pressure test in mice has been examined. C.NMTA-induced antinociception was significantly potentiated by simultaneously intracerebroventricular or intraperitoneal injection of TRH (approximately 20-50%) in a dose-dependent manner, whereas the effect of morphine was not influenced significantly by TRH. C.HP had no significant effect on the antinociceptive response induced by C.NMTA or morphine. It is concluded that the mechanism of C.NMTA-induced antinociception may be involved in TRH neuronal system in the brain.

1-(Phenyl)isoquinoline carboxamides: a novel class of subtype selective inhibitors of thyrotropin-releasing hormone (TRH) receptors

We report the synthesis of and binding to the two subtypes of mouse thyrotropin-releasing hormone (TRH) receptors, TRH-R1 and TRH-R2, of several 1-(phenyl)isoquinoline carboxamide analogues. These analogues showed a degree of selectivity for binding at TRH-R2. These are the first ligands reported that show selective binding to these receptors.

Regulator of G Protein Signaling 4 Suppresses Basal and Thyrotropin Releasing-Hormone (TRH)-Stimulated Signaling by Two Mouse TRH Receptors, TRH-R1 and TRH-R2

Regulator of G Protein Signaling 4 Suppresses Basal and Thyrotropin Releasing-Hormone (TRH)-Stimulated Signaling by Two Mouse TRH Receptors, TRH-R1 and TRH-R2

Regulator of G protein signaling 4 suppresses basal and thyrotropin releasing-hormone (TRH)-stimulated signaling by two mouse TRH receptors, TRH-R(1) and TRH-R(2).

We cloned the mouse TRH receptor type 2 (mTRH-R2) gene, which is 92% identical with rat TRH-R2 and 50% identical with mTRH-R1 at the amino acid level, and identified an intron within the coding sequence that is not present in the TRH-R1 gene structure. Similar to its rat homolog, mTRH-R2 binds TRH with an affinity indistinguishable from mTRH-R1, signals via the phosphoinositide pathway like mTRH-R1, but exhibits a higher basal signaling activity than mTRH-R1. We found that regulator of G protein signaling 4 (RGS4), which differentially inhibits signaling by other receptors that couple to Gq, inhibits TRH-stimulated signaling via mTRH-R1 and mTRH-R2 to similar extents. In contrast, other RGS proteins including RGS7, RGS9, and GAIP had no effect on signaling by mTRH-R1 or mTRH-R2 demonstrating the specificity of RGS4 action. Interestingly, RGS4 markedly inhibited basal signaling by mTRH-R2. Inhibition of basal signaling of mTRH-R2 by RGS4 suggests that modulation of agonist-independent signaling may be an important mechanism of regulation of G protein-coupled receptor activity under normal physiologic circumstances.

Molecular cloning and functional expression of a human thyrotropin-releasing hormone (TRH) receptor gene

In this study, we isolated genomic DNA fragments coding for the human thyrotropin-releasing hormone (TRH) receptor. Analysis of the nucleotide sequence revealed that the human TRH receptor gene had an exon-intron structure comprising at least two exons. A polypeptide encoded by the gene consisted of 398 amino acid residues with putative seven transmembrane domains. It showed high homology as a whole amino acid sequence with the rat and mouse TRH receptors except for considerable variation in the C-terminal region. Chromosomal mapping study indicated that the human TRH receptor gene was assigned to chromosome 8. Chinese hamster ovary (CHO) cells transfected with a DNA fragment containing the coding regions of the human TRH receptor bound with [ 3H]TRH. This binding was inhibited by adding unlabeled TRH in a dose-dependent fashion. Scatchard analysis indicated that the transfected CHO cells expressed a single class of high affinity binding sites at a dissociation constant ( K d) of approximately 1 nM. These results demonstrated that the isolated gene encoded a specific TRH receptor with high affinity.

TPA (12- O-tetradecanoylphorbol-13-acetate enhances the central hypoglycemic action of thyrotropin-releasing hormone in mice

This study examined the effect of the calcium- and phospholipid-dependent protein kinase C (PKC) activator, 12-O-tetradecanoylphorbol-13-acetate (TPA) on the plasma glucose responses to central thyrotropin-releasing hormone (TRH) injection in mice in order to evaluate the involvement of PKC in the mechanism of TRH action in the central nervous system (CNS). TRH (0.1–10 μg), as well as the neuroactive TRH analogs, CG 3509, CG 3703, DN 1417, RX 77368, [Nva 2]-TRH, KPC-TRH, and TRH-Gly (0.1–10 μg), injected centrally in normoglycemic mice reduced the circulating glucose levels in a dose-dependent manner. TPA (0.1–1 μg), administered centrally together with TRH (1 μg) or the TRH analogs strongly enhanced the hypoglycemic response. Similar doses of TPA had no effect on plasma glucose when administered alone or together with TRH analogs devoid of central hypoglycemic action, i.e. [Glu 1]-TRH, [Phe 2]-TRH, and [Gly 3]-TRH (1 μg). Central injection of a TPA analog lacking PKC-stimulating activity, 4-α-phorbol (0.1–1 μg) had no effect on the hypoglycemic response to coadministered TRH. These results, demonstrating a specific effect of TPA in enhancing the hypoglycemic response to central TRH or its neuroactive, though not inactive, analogs are consistent with a possible role for PKC in the mechanism of TRH action in the CNS.

Effects of acute and long-term treatments with thyrotropin-releasing hormone on locomotor activity and jumping behavior in mice

The acute and chronic effects of thyrotropin-releasing hormone (TRH) on ambulation and, in combination with apomorphine, on jumping behavior were investigated in mice. A single administration of TRH (1–10 mg/kg SC) produced an initial hyperactivity in a dose-dependent manner. Following administration of TRH (1–10 mg/kg SC) for 21 successive days, the stimulatory effect on locomotion progressively increased. Haloperidol exerted a biphasic action on hyperlocomotion induced by acute and repeated TRH, i.e., stimulation at lower doses (0.01–0.02 mg/kg SC) and inhibition at higher doses (0.05–1 mg/kg SC). When TRH was administered in combination with low doses of apomorphine, locomotor activity was inhibited but jumping behavior occurred. The inhibitory effect of low doses of apomorphine on locomotion was shifted from doses of 0.1–0.25 mg/kg SC of apomorphine for acute TRH (10 mg/kg) to 0.25–0.35 mg/kg for repeated TRH (10 mg/kg), whereas the stimulatory effect of higher doses of apomorphine (0.5–1 mg/kg SC) on locomotion tended to decrease with repeated TRH. Jumping behavior induced by the combined treatment of TRH and apomorphine was proportional to the dose of TRH but exhibited an inverted-U response to the dose of apomorphine. Chronic TRH (10 mg/kg) in combination with apomorphine (0.1–1 mg/kg SC) also produced jumping behavior, but the dose-response curve for apomorphine was shifted to the right. The present results suggest that repeated treatment with TRH in mice produces hyperlocomotion, despite attenuation of both pre- and postsynaptic receptor activity, and that the inhibitory effect of repeated TRH on presynaptic receptors may be more potent than that on postsynaptic receptors.

The antinociceptive effects of histidyl-proline diketopiperazine and thyrotropin-releasing hormone in the mouse

Intracerebroventicular (i.c.v.) injection of histidyl-proline diketopiperazine [cyclo (His-Pro)], an active metabolite of thyrotropin-releasing hormone (TRH) in mice produced an antinociceptive effect in a dose-dependent manner as measured in four antinociceptive tests; tail-pressure, tail-flick, hot-plate and acetic acid writhing. The antinociceptive effect of cyclo (His-Pro) reached a maximum at 5 min and lasted for 30 min. The ED 50 values for the tests were 760.0 (598.4–965.2), 540.0 (442.6–658.8), 595.0 (487.7–725.9) and 370.0 (286.8–477.3) nmol/mouse and the slope functions were 1.61, 1.56, 1.57 and 1.66, respectively. Pretreatment with naloxone (0.5, 2 and 8 mg/kg), an opioid antagonist, administered subcutaneously antagonized the antinociceptive effect of cyclo (His-Pro). TRH injected i.c.v. to mice also exerted a dose-dependent antinociceptive action as measured in three antinociceptive tests; tail-pressure, hot-plate and acetic acid writhing. The antinociceptive effect of TRH was only seen at 5 min. The ED 50 values for each test were 112.0 (47.5–264.3), 19.2 (10.4–35.5) and 0.2 (0.1–0.4) nmol/mouse and the slope functions were 8.89, 4.14 and 3.94, respectively. TRH was without effect in the tail-flick test. In contrast to cyclo (His-Pro), TRH-induced antinociception was not antagonized by pretreatment with naloxone (0.5, 2 and 8 mg/kg). The data suggest that the two peptides have a different mechanism of antinociceptive action in relation in the involvement of the opioid system in the central nervous system.

Interaction of neurotensin with prostaglandin E2 and prostaglandin synthesis inhibitors: effects on colonic temperature in mice.

Neurotensin (NT) administered intracisternally (i.c.) to adult mice produced a marked hypothermia while prostaglandin E2, administered by the same route, produced hyperthermia. When administered concurrently the effects of the two substances were neutralized. The prostaglandin synthesis inhibitors, indomethacin and acetylsalicylic acid, were injected subcutaneously 30 min prior to i.c. administered NT and/or thyrotropin-releasing hormone (TRH). Both inhibitors failed to potentiate the hypothermia induced by NT or alter its antagonism by TRH in mice kept at 26 degrees C. When mice were kept at 6 degrees C, pretreatment with indomethacin, but not acetylsalicylic acid, potentiated NT-induced hypothermia and prevented its antagonism by TRH. Because indomethacin inhibits synthesis of prostaglandins within the central nervous system (CNS) as well as in peripheral organs while acetylsalicylic acid acts only in the periphery, it appears that NT-induced hypothermia in a cold environment is enhanced by a reduction of prostaglandins in the CNS.

The effect of thyrotropin releasing hormone on pentobarbital tolerance

The effects of thyrotropin releasing hormone (TRH) on the responses produced by acute and chronic administration of pentobarbital (PB) were investigated in male Swiss Webster mice. Intraperitoneal injection of TRH antagonized the hypothermic and hypnotic effect of PB (75 mg/kg, i.p.). Pentobarbital hypnosis was also antagonized by TRH in mice rendered tolerant to PB by implantation of a pellet (each containing 75 mg of free acid form of the drug) for two days. Injections of TRH (5 mg/kg) once a day or twice a day for two days did not modify the development of tolerance to the hypothermic and hypnotic effect of PB. It is concluded that TRH can modify only the acute responses of PB in naive and PB-tolerant mice, however, in the dosage regimen used it does not affect the processes involved in the genesis of the tolerant state.

Hyperthyroidism induced by thyrotropin-releasing hormone in mice.

To test whether repeated administration of thyrotropin—releasing hormone (TRH) results in hyperthyroidism, we administered TRH, 1 μg every 12 hr, to 6 groups of mice for 1–10 days. No increase in thyroid size, 131I uptake or serum T4 was noted in any group of animals when compared to saline injected controls. When the experiment was repeated with a higher dose, 10 ¼g of TRH, hyperthyroidism evidenced by a mild increase in serum T4 was detected only after 2 doses of TRH; it was not present in any other group despite continued administration of TRH at 12–hr intervals for up to 7 days. In another experiment, where TRH (10 ¼g) was administered at 2–hr intervals around the clock for 1–10 doses, serum T4 increased after the first dose and the increment was maintained at comparable or higher levels during subsequent injections. Thyroid 131I uptake and thyroid size were not significantly affected after 1–5 doses of TRH; however, significant increase in both was noted after administration of 10 doses. These result...