Thyrotropin-releasing Hormone System Research Articles

Cardiac Thyrotropin-releasing Hormone Inhibition Improves Ventricular Function and Reduces Hypertrophy and Fibrosis After Myocardial Infarction in Rats

BackgroundCardiac thyrotropin-releasing hormone (TRH) is a tripeptide with still unknown functions. We demonstrated that the left ventricle (LV) TRH system is hyperactivated in spontaneously hypertensive rats and its inhibition prevented cardiac hypertrophy and fibrosis. Therefore, we evaluated whether in vivo cardiac TRH inhibition could improve myocardial function and attenuate ventricular remodeling in a rat model of myocardial infarction (MI). Methods and ResultsIn Wistar rats, MI was induced by a permanent left anterior descending coronary artery ligation. A coronary injection of a specific small interfering RNA against TRH was applied simultaneously. The control group received a scrambled small interfering RNA. Cardiac remodeling variables were evaluated one week later. In MI rats, TRH inhibition decreased LV end-diastolic (1.049 ± 0.102 mL vs 1.339 ± 0.102 mL, P < .05), and end-systolic volumes (0.282 ± 0.043 mL vs 0.515 ± 0.037 mL, P < .001), and increased LV ejection fraction (71.89 ± 2.80% vs 65.69 ± 2.85%, P < .05). Although both MI groups presented similar infarct size, small interfering RNA against TRH treatment attenuated the cardiac hypertrophy index and myocardial interstitial collagen deposition in the peri-infarct myocardium. These effects were accompanied by attenuation in the rise of transforming growth factor-β, collagen I, and collagen III, as well as the fetal genes (atrial natriuretic peptide, B-type natriuretic peptide, and beta myosin heavy chain) expression in the peri-infarct region. In addition, the expression of Hif1α and vascular endothelial growth factor significantly increased compared with all groups. ConclusionsCardiac TRH inhibition improves LV systolic function and attenuates ventricular remodeling after MI. These novel findings support the idea that TRH inhibition may serve as a new therapeutic strategy against the progression of heart failure.

Angiotensin II requires an intact cardiac thyrotropin-releasing hormone (TRH) system to induce cardiac hypertrophy in mouse

Cardiac tyhrotropin-releasing hormone (TRH) is overexpressed in the hypertrophied left ventricle (LV) of spontaneously hypertensive rats (SHR) and its inhibition prevents both hypertrophy and fibrosis. In a normal heart, the TRH increase induces fibrosis and hypertrophy opening the question of whether TRH could be a common mediator of left ventricular hypertrophy (LVH). We used angiotensin II (AngII) as an inductor of LVH to evaluate if the blockade of LV-TRH prevents hypertrophy and fibrosis in mice. We challenged C57BL/6 adult male mice with an infusion of AngII (osmotic pumps; 2 mg/kg.day) to induce LVH. Groups of mice were injected with an intracardiac siRNA-TRH or scrambled siRNA (siRNA-Con). Body weight, water intake and systolic arterial blood pressure (SABP) were measured daily. AngII significantly increased water intake and SABP (p < .05). Cardiac hypertrophy (heart weight/body weight) was evident in the group with the normal cardiac TRH system. In fact, it was found an AngII-induced increase of TRH precursor mRNA (p < .05) in conjunction with elevated TRH levels measured by immunohistochemistry and western blot. These changes were not observed in the AngII + siRNA-TRH group. Furthermore, AngII increased significantly (p < .05) BNP (hypertrophic marker), collagens I and III and TGF-β (fibrosis markers) expression in the group with the native cardiac TRH system. These increases were attenuated in the groups with the TRH system blocked despite the high blood pressure. Similar and stronger results were observed “in vitro” with NIH3T3 and H9C2 cell culture models, where, when the TRH system is blocked, AngII stimulus was not able to induce the markers of its fibrotic and hypertrophic effects, so we believe that these effects are independent of any other physiological modifications.Our results point out that cardiac TRH is required for AngII-induced hypertrophic and fibrotic effects.

Thyrotropin-releasing hormone overexpression induces structural changes of the left ventricle in the normal rat heart.

Thyrotropin-releasing hormone (TRH) hyperactivity has been observed in the left ventricle of spontaneously hypertensive rats. Its long-term inhibition suppresses the development of hypertrophy, specifically preventing fibrosis. The presence of diverse systemic abnormalities in spontaneously hypertensive rat hearts has raised the question of whether specific TRH overexpression might be capable of inducing structural changes in favor of the hypertrophic phenotype in normal rat hearts. We produced TRH overexpression in normal rats by injecting into their left ventricular wall a plasmid driving expression of the preproTRH gene (PCMV-TRH). TRH content and expression of preproTRH, collagen type III, brain natriuretic peptide, β-myosin heavy chain, Bax-to-Bcl-2 ratio, and caspase-3 were measured. The overexpression maneuver was a success, as we found a significant increase in both tripeptide and preproTRH mRNA levels in the PCMV-TRH group compared with the control group. Immunohistochemical staining against TRH showed markedly positive brown signals only in the PCMV-TRH group. TRH overexpression induced a significant increase in fibrosis, evident in the increase of collagen type III expression accompanied by a significant increase in extracellular matrix expansion. We found a significant increase in brain natriuretic peptide and β-myosin heavy chain expression (recognized markers of hypertrophy). Moreover, TRH overexpression induced a slight but significant increase in myocyte diameter, indicating the onset of cell hypertrophy. We confirmed the data "in vitro" using primary cardiac cell cultures (fibroblasts and myocytes). In conclusion, these results show that a specific TRH increase in the left ventricle induced structural changes in the normal heart, thus making the cardiac TRH system a promising therapeutic target.

The acute response of the amygdalar TRH system to psychogenic stressors varies dependent on the paradigm and circadian condition

Central administration of thyrotropin releasing hormone (TRH) reduces anxiety; amygdalar TRH expression is inversely proportional to the anxious behavior displayed in the elevated plus maze performed during the dark phase (EPM-D). To better understand the role of TRH in amygdala function, we evaluated the expression of TRH and the elements involved in its transmission in various stressful paradigms and how they associated with behavior. Wistar male rats were exposed to restraint (RES), EPM, or the open field test (OFT) and sacrificed 0–60min afterwards; OFT, RES and EPM were performed during the light (L), and OFT during the dark phase. Restraint increased amygdalar levels of proCRH mRNA, without change in proTRH. All paradigms augmented corticosterone release, highest after OFT-L that also enhanced proCRH mRNA levels and decreased those of proTRH. OFT-D activated the TRH system. Levels of anxiety or locomotion were similar in animals tested in light or dark phases but their association with biochemical parameters differed. ProTRH expression and TRH release correlated positively with decreased anxiety in EPM-L and in OFT-D. No association with anxiety was detected in OFT-L where proCRH and proTRH expression correlated with locomotion supporting their involvement in arousal. The responses of TRH amygdalar systems appeared modulated by the extent of the stress response and by the circadian conditions. Increased proTRH expression of animals exposed to OFT-D was specifically observed in the cortical nucleus of the amygdala, area involved in processing fear stimuli; these TRH neurons may thus be part of a circuit with anxiolytic properties.

Cardiac Thyrotropin-Releasing Hormone Mediates Left Ventricular Hypertrophy in Spontaneously Hypertensive Rats

Local thyrotropin-releasing hormone (TRH) may be involved in cardiac pathophysiology, but its role in left ventricular hypertrophy (LVH) is still unknown. We studied whether local TRH is involved in LVH of spontaneously hypertensive rats (SHR) by investigating TRH expression and its long-term inhibition by interference RNA (TRH-iRNA) during LVH development at 2 stages (prehypertrophy and hypertrophy). SHR and their control rats (WKY) were compared. Cardiac hypertrophy was expressed as heart/total body weight (HW/BW) ratio. TRH content (radioimmuno assay), preproTRH, TRH receptor type I, brain natriuretic peptide (BNP), and collagen mRNA expressions (real-time polymerase chain reaction) were measured. For long-term inhibition of TRH, TRH-iRNA was injected into the left ventricle (LV) wall for 8 weeks. Hearts were processed for morphometric studies and immunohistochemical analysis using antibodies against α-smooth muscle actin and collagen type III. LV preproTRH-mRNA abundance was similar in both strains at 7 weeks of age. At the hypertrophic stage (18 weeks old), however, there was a 15-fold increase in SHR versus WKY, consistent with a significant increase in tripeptide levels and the expression of its receptor. Specific LV-TRH inhibition at the prehypertensive stage with TRH-iRNA, which decreased >50% preproTRH expression and tripeptide levels, prevented LVH development as shown by the normal HW/BW ratio observed in TRH-iRNA-treated SHR. In addition, TRH-iRNA impeded the increase in BNP and type III collagen expressions and prevented the increase in cardiomyocyte diameter evident in mismatch iRNA-treated adult SHR. These results show for the first time that the cardiac TRH system is involved in the development of LVH in SHR.

Thyrotropin-Releasing Hormone Precursor Gene Knocking Down Impedes Melanocortin-Induced Hypertension in Rats

To the Editor: Recently, da Silva et al1 reported that endogenous melanocortin may cause elevation of arterial blood pressure (ABP) in spontaneously hypertensive rats. We invite authors to consider that the hypothalamic thyrotropin-releasing hormone (TRH) system may be involved, because spontaneously hypertensive rats show an hyperactivity of this system,2 and intracerebroventricular (ICV) injections of a prepro-TRH antisense oligonucleotide (AS) decreases both elevated TRH content and ABP independent of thyroid status.3 Leptin effects include increases in sympathetic activity …

The Rotating Ring-Disk Electrochemistry of the Copper(II) Complex of Thyrotropin-releasing Hormone.

Thyrotropin-releasing Hormone (TRH) forms an electroactive Cu(II) complex in aqueous solution. Rotating ring-disk electrochemistry reveals oxidation at the disk electrode and reduction at the ring electrode. The plot of limiting current vs. square root of rotation frequency deviates from the Levich equation, indicating both preceding and following chemical reactions. The reaction following the oxidation is a multiple-electron ECE-type of process that has been seen before in Cu(II)-peptide electrochemistry. The preceding reaction is unusual. The deviation from diffusion-controlled behavior is more pronounced at higher initial concentration of Cu(II) and peptide. We propose that a non-electroactive dimer, Cu(II)(2)-TRH(2), is in a slow equilibrium with the electroactive Cu(II)-TRH. Simulation of the RRDE behavior of the postulated Cu(II)-TRH system has succeeded in matching experimental data. Capillary electrophoresis indicates that there is a negative charge on the dimer. It is suggested that a hydroxo-bridge may link the two Cu(II) centers. Calculations verify that bi-nuclear Cu(II)(2)-TRH(2) complexes are possible.

Effect of immobilization on in vitro thyrotropin-releasing hormone release from brain septum in wild-type and corticotropin-releasing hormone knock-out mice.

There is considerable evidence linking alcohol consumption, sedation, and thyrotropin-releasing hormone (TRH) in the brain septum. We have shown that ethanol in clinically relevant concentrations can in vitro induce TRH release from the septum by a mechanism involving neuronal swelling. Corticotropin-releasing hormone-deficient (CRH-KO) mice serve as an interesting model to help us understand the role of CRH in the regulation of different neuroendocrine systems. The aim of this study was to compare TRH release activity in the brain septum at basal and stress conditions in CRH-KO mice and their wild-type (WT) littermates. Experimental mice were decapitated immediately or 3 h after single (2 h) or repeated (seven times for 2 h daily) immobilization stress. The brain septum was immediately cut out and incubated to measure basal-, ethanol-, and hyposmosis-stimulated TRH release in vitro. Ethanol in isosmotic medium or hyposmotic medium stimulated TRH release from mice septal explants from WT and CRH-KO mice. The response was disturbed immediately after immobilization and recovered 3 h later. Our results show that immobilization stress transiently affects the TRH system in brain septum. Inborn absence of CRH does not affect septal TRH and its response to ethanol before and 3 h after immobilization.

Thyrotropin-Releasing Hormone Decreases Leptin and Mediates the Leptin-Induced Pressor Effect

Leptin, an adipocyte-released hormone, modifies food intake and energy expenditure regulating hypothalamic-pituitary-thyroid axis function. We previously reported that thyrotropin-releasing hormone (TRH) precursor gene overexpression induces hypertension in the normal rat and that spontaneously hypertensive rats have central TRH hyperactivity with increased TRH synthesis and release and an elevated TRH receptor number. In both models, intracerebroventricular antisense (AS) treatment against the TRH precursor produced a dose-dependent reduction of the increased diencephalic TRH content while normalizing high arterial blood pressure. In this article, we report that male Wistar rats that were made hypertensive by intracerebroventricular injection of a eucaryotic expression plasmid containing the pre-TRH cDNA showed decreased leptin plasma levels and that pre-TRH AS treatment reversed this phenomenon. In addition, male and female spontaneously hypertensive rats showed lower levels of circulating leptin than did sex-matched Wistar-Kyoto control rats. This difference also was abated by the pre-TRH AS treatment. Conversely, 20 microg ICV leptin induced a long-lasting pressor effect (18 +/- 5 mm Hg, n=6, P<0.01, >60 minutes) that was not observed in pre-TRH AS pretreated rats (2 +/- 3 mm Hg, n=6) but persisted in rats used as controls that were treated with inverted oligonucleotide (20 +/- 6 mm Hg, n=4, P<0.01). These data suggest that in rats with TRH-induced hypertension, leptin is decreased, inducing compensatory adiposity. We propose that because leptin produces central TRH synthesis and release, obesity may induce hypertension through TRH system activation and that the TRH-leptin interaction may thus contribute to the strong association between hypertension and obesity.

Differential Responses of Thyrotropin-Releasing Hormone (TRH) Neurons to Cold Exposure or Suckling Indicate Functional Heterogeneity of the TRH System in the Paraventricular Nucleus of the Rat Hypothalamus

Thyrotropin-releasing hormone (TRH) is released from the median eminence upon neural stimulation such as cold or suckling exposure. Concomitant with the cold- or suckling-induced release of TRH is a rapid and transient increase in the expression of proTRH mRNA in the paraventricular nucleus (PVN) of the hypothalamus. We employed two strategies to determine whether TRH neurons responding to cold exposure are different from those responding to suckling. First, we attempted to identify a marker of cellular activation in TRH neurons of the PVN. Cold induced c-fos expression in about 25% of TRH neurons of the PVN, but no induction was observed by suckling. Moreover, we explored the expression of a variety of immediate early genes including NGFI-A, fra-1 and c-jun, or CREB phosphorylation but found none to be induced by suckling. The number of cells expressing high levels of proTRH mRNA was counted and compared to total expressing cells. An increased number of cells expressing high levels of proTRH mRNA was observed when both stimuli were applied to the same animal, suggesting that different cells respond separately to each stimulus. We therefore analyzed the distribution of responsive TRH neurons as defined by the cellular level of proTRH mRNA. The proTRH mRNA signal was analyzed within three rostrocaudal zones of the PVN and within six mediolateral columns. Results showed that in response to cold, all areas of the PVN of the lactating rat present increased proTRH mRNA levels, including the anterior zone where few hypophysiotropic TRHergic cells are believed to reside. The distribution of the proTRH mRNA expressing cells in response to cold was quite comparable in female and in male rats. In contrast, the response after suckling was confined to the middle and caudal zones. Our results provide evidence of a functional specialization of TRH cells in the PVN.

Antisense inhibition of thyrotropin-releasing hormone reduces arterial blood pressure in spontaneously hypertensive rats.

Thyrotropin-releasing hormone (TRH) plays an important role in central cardiovascular regulation. Recently, we described that the TRH precursor gene overexpression induces hypertension in the normal rat. In addition, we published that spontaneously hypertensive rats (SHR) have central extrahypothalamic TRH hyperactivity with increased TRH synthesis and release and an elevated TRH receptor number. In the present study, we report that intracerebroventricular antisense (AS) treatment with a phosphorothioate oligonucleotide against the TRH precursor gene significantly diminished up to 72 hours and in a dose-dependent manner the increased diencephalic TRH content, whereas normalized systolic blood pressure (SABP) was present in the SHR compared with Wistar-Kyoto (WKY) rats. Although basal thyrotropin was higher in SHR compared with WKY rats and this difference disappeared after antisense treatment, no differences were observed in plasma T4 or T3 between strains with or without AS treatment, indicating that the effect of the AS on SABP was independent of the thyroid status. Because the encephalic renin-angiotensin system seems to be crucial in the development and/or maintenance of hypertension in SHR, we investigated the effect of antisense inhibition of TRH on that system and found that TRH antisense treatment significantly diminished the elevated diencephalic angiotensin II (Ang II) content in the SHR without any effect in control animals, suggesting that the Ang II system is involved in the TRH cardiovascular effects. To summarize, the central TRH system seems to be involved in the etiopathogenesis of hypertension in this model of essential hypertension.

Effects of repeated cocaine administration on the thyrotropin-releasing hormone level and receptors in the rat brain

The effects of single and repeated administration of cocaine on the thyrotropin-releasing hormone (TRH) level and receptors in discrete rat brain structures were evaluated. Male Wistar rats received saline or cocaine (15 mg/kg i.p., once an hour within 3 h, for 8 days). The animals were killed by decapitation at 45 min and 72 h (chronic group only) after the last injection. A radioimmunoassay (RIA) study showed that a single dose of cocaine increased the TRH level in the striatum by 68%, but had no significant effect on the peptide content in the nucleus accumbens, hippocampus, amygdala, septum, hypothalamus, frontal and prefrontal cortex at 45 min after the drug injection. Repeated administration of cocaine increased the TRH level in the striatum by 89% at 45 min, and in the hippocampus by 26% at 72 h after the last dose. No changes in the TRH level were found in other brain structures. In vitro cocaine (10 −6−10 −4 M) inhibited the K +-stimulated release in a concentration-dependent manner, but had no effect on the basal release of TRH from the striatum and nucleus accumbens of naive rats. Acute cocaine decreased the B max of TRH receptors in the striatum, but had no effect on the density and affinity of TRH receptors in other brain regions. Repeated administration of cocaine evoked a long-lasting decrease in the B max of TRH receptors in the striatum (by c. 30%), whereas an increase in that parameter was observed in the frontal cortex. The B max and affinity of TRH receptors following repeated cocaine remained unchanged in the nucleus accumbens. The results obtained indicate that cocaine affects the TRH system mainly in the striatum, and to a lesser extent in the nucleus accumbens, cortex and hippocampus. Furthermore, the above changes do not resemble those induced by amphetamine, which points to certain differences in adaptation of the TRH neuronal system to these psychostimulants. On the other hand, the increase in the hippocampal TRH level during both chronic cocaine and morphine withdrawal is a common feature of the mechanism of dependence on these drugs.

The TRH neuronal phenotype forms embryonic cell clusters that go on to establish a regionalized cell fate in forebrain.

How neurons diversify in developing brain to produce discrete cell fates in their appropriate regions remains a fundamental question. Embryonic Xenopus was previously used to identify juxtaposed embryonic cells that first express proopiomelanocortin mRNA in forebrain and pituitary, supporting the idea that this neuropeptide phenotype is induced locally (Hayes and Loh, 1990, Development 110: 747-757). To begin to examine how a more widespread population of forebrain cells is set up, the present focus is on the thyrotropin-releasing hormone (TRH) phenotype. Serial section in situ hybridization histochemistry produced the unexpected finding that the adult-like TRH system spanning forebrain and comprising over six different telencephalic and diencephalic nuclei, is preceded by an embryonic TRH cell population that is initially localized and then highly regionalized in the area from which the adult pattern develops. Thus, the first TRH cells, detected in vivo after 35h (stage 29/30), were confined to discrete anterior or posterior bilateral clusters in embryonic forebrain or hindbrain. Thereafter, the TRH cell clusters in diencephalon, but not hindbrain, expanded to form rows, extending anteriorly into telencephalon and bifurcating posteriorly around the infundibulum. By 80 h (stage 42), after extensive brain morphogenesis, these forebrain rows showed regional differences in levels of TRH mRNA corresponding to the specific brain nuclei that have been shown to contain TRH cells in adult. These findings show that subsets of phenotype-specific forebrain cells first form a regionalized neuronal cell fate before distinct brain nuclei form. This in turn points to the testable hypothesis in Xenopus that certain neuronal cell fates in forebrain may be dictated by cell lineage or local induction.

Changes in brain thyrotropin-releasing hormone (TRH) of seizure-prone El mice

We examined the anticonvulsant effects of DN-1417 an analog of the thyrotropin-releasing hormone (TRH) in seizure-prone El mice. Changes in both immunoreactive TRH (IR-TRH) and TRH receptor binding activity in discrete brain regions of El mice were also measured before and after sensitization and during the postictal period, and they were compared with those in the ddY mice. Intraperitoneal injection of DN-1417 with 150 and 450 mg/kg significantly increased the El mouse seizure threshold in a dose-dependent manner. IR-TRH in the hippocampus of El mice, which was significantly lower than in ddY mice, significantly increased after sensitization. During the postictal period, however, it slowly decreased again and then gradually recovered to the preconvulsive level without any change in TRH receptor binding. In the striatum of El mice, although TRH receptor binding was significantly higher than in ddY mice, it was not affected by sensitization. These findings indicate that the hippocampal TRH system may play an inhibitory role in El mouse seizures whereas the striatal TRH system may be important for its seizure susceptibility.

Bulbospinal and intraspinal thyrotropin releasing hormone systems: Modulation of spinal cord pain transmission

Recent research indicates that pain processing by spinal nociceptive neurons is modulated by: 1) descending bulbospinal pathways originating in the rostral ventral medulla (RVM) and 2) short intraspinal peptide systems located in the dorsal horn. Immunohistochemical studies have identified both bulbospinal and intraspinal thyrotropin releasing hormone (TRH) systems, however, the role of these two TRH systems in pain modulation has not been defined. The purpose of the present study was to explore the role of the bulbospinal and intraspinal TRH systems in pain modulation. Three TRH mediated neural network were examined in electrophysiologic experiments; each model predicting specific testable outcomes. Model A represented a direct TRH projection from RVM to spinal cord. This model predicted that the effect of RVM stimulation and TRH micropressure administration on dorsal horn unit activity would be in the same direction, either both inhibitory effects or both excitatory. Of 44 dorsal horn units inhibited by RVM stimulation, 82% were excited by TRH, therefore, Model A was rejected. Model B interposed an intraspinal neuron between the bulbospinal TRH projection and the dorsal horn nociceptive unit. This model predicted that desensitization of spinal TRH receptor systems should block the effect of RVM stimulation on dorsal horn neuron. In TRH desenisitization experiments, no attenuation or blockade of RVM stimulation was observed, therefore, the effect of RVM stimulation did not appear mediated by TRH and Model B was rejected. Model C, which described an intraspinal TRH system facilitating pain transmission was supported by several lines of evidence including: 1) prominent increase in unit activity after TRH micropressure application and 2) the ability of TRH to magnify the effect of noxious stimulation on unit activity while having little or no effect on baseline firing rate. It is concluded that TRH interneurons may either directly or indirectly, via modulation of transmitter release from primary afferents, facilitate pain transmission in spinal cord while the modulation of pain transmission by the bulbospinal TRH system could not be clearly demonstrated under the present experimental conditions.

Reduction of rat striatal thyrotropin-releasing hormone receptors produced by repeated methamphetamine administration

It has been reported previously that repeated, but not continuous, administration of methamphetamine (MAP) to animals produces progressive and sustained enhancement of MAP-induced behavior (behavioral sensitization), which may be related to functional changes in central dopamine (DA) systems. To investigate the possible involvement of thyrotropin-releasing hormone (TRH), a neuromodulator of DA, both immunoreactive TRH (IR-TRH) levels and specific TRH binding were examined in rat brain regions after MAP administration either repeatedly (4 mglkg intraperitoneally once a day for 14 consecutive days) or continuously (about 4 mg/kg/day for 13 consecutive days). Although no significant changes were observed in IR-TRH levels in any regions of the brain following repeated MAP injections, specific TRH binding in the striatum significantly decreased. Scatchard analysis revealed that the decrease was due to a reduction in the maximum number of binding sites (B max ). Pretreatment with haloperidol prior to each MAP injection prevented this decrease. Continuous MAP administration had no effect on regional specific TRH binding. These results suggest that repeated MAP administration caused lasting dysfunction in the brain TRH system, which may be implicated in the behavioral sensitization.

Changes in brain thyrotropin-releasing hormone (TRH) of El mice.

The present study showed that DN-1417 had a dose-dependent anticonvulsant activity on El mouse seizure. This finding is consistent with other reports using the kindling model of epilepsy. Since both the El mouse and kindling preparations have been regarded as complex partial seizure with secondary generalization, endogenous brain TRH, as well as exogenous TRH, may act as an anticonvulsant substance to such a seizure type of epilepsy. Moreover, this study showed IR-TRH of the El mouse changed significantly in the striatum or hippocampus genetically or postictally without a change in the TRH receptor binding. A transient decrease in hippocampal IR-TRH after convulsion shown in this study may suggest an increased release of TRH during and after the seizure. Further studies are required to clarify the relationship between a change in the brain TRH system and seizure susceptibility in the El mouse.

Long-term increase in striatal thyrotropin-releasing hormone receptor binding caused by amygdaloid kindling.

Our previous finding that intracerebroventricular (i.c.v.) administration of both thyrotropin-releasing hormone (TRH) and its analogue, gamma-butyrolactone-gamma-carbonyl-L-histidyl-L-prolinamide citrate (DN-1417), suppressed seizure development of amygdaloid (AM) kindling and kindled AM seizures leads to a new hypothesis that endogenous TRH may be an antiepileptic substance in the brain. In this study, we examined postictal chronological changes in both immunoreactive TRH (IR-TRH) and TRH receptor binding activity in discrete brain regions of AM-kindled rats to study the relationship of the brain TRH system to kindling-induced seizure susceptibility. AM-kindled rats were decapitated 30 min, 24 h, 48 h, 7 days, and 21 days after the last kindled convulsion. IR-TRH increased markedly in the AM/pyriform cortex and hippocampus 24 and 48 h after the last convulsion, and returned to the control (unstimulated, sham-operated) value within 3 weeks after the convulsions ended. In contrast, a significant increase in the striatal TRH binding sites was evident 24 h after the cessation of convulsions which lasted 21 days. A lasting change in the striatal TRH neural system may be related to kindling-induced seizure susceptibility.

A thyrotropin-releasing hormone-containing system in the rat dorsal horn separate from serotonin

In this study, we report the identification of a thyrotropin-releasing hormone (TRH)-containing system in the dorsal horn of the rat spinal cord. This system is distinct from the TRH and serotonin (5-hydroxytryptamine, 5-HT) cotransmitter supraspinal system that has projections to the intermediolateral (IML) and ventral columns. Spinal cord sections from untreated rats, and those treated with colchicine or 5,7-dihydroxytryptamine (5,7-DHT) were processed using peroxidase-antiperoxidase (PAP) immunocytochemistry with nickel intensification. Results of the 5,7-DHT treatment were verified by quantifying TRH and 5-HT by radioimmunoassay (RIA) and high performance liquid chromatography (HPLC), respectively. Prominent immunocytochemical staining for TRH in the dorsal horn was seen in varicose fibers mainly in lamina II and superficial lamina III of the dorsal horn of the spinal cord of control rats. A few fibers were seen ascending into lamina I. A moderate number of fibers that were immunoreactive for 5-HT were primarily in laminae I and II. The distribution of TRH- and 6-HT-containing neurites in the IML and the ventral horn agreed with previously published reports. Rats treated with colchicine showed many small round TRH immunoreactive cells that were limited to laminae II/III of the dorsal horn. TRH immunoreactivity in the dorsal horn and IML was resistant to the effects of the selective serotonin neurotoxin, 5,7-DHT, while the ventral horn was depleted of TRH staining. Serotonin was almost completely eliminated in all spinal cord laminae. Quantitative biochemical studies showed significant, but non-parallel reductions of TRH and 5-HT in cervical, thoracic and lumbar spinal cord. These studies demonstrate the existence of TRH-containing cell bodies and terminals in the dorsal horn of the rat spinal cord. These findings (1) provide evidence that a TRH-containing system exists in the dorsal horn of the rat and that it is distinct from the descending medullary raphe system that contains 5-HT; (2) suggest that a population of TRH-containing fibers that project to the IML may not contain 5-HT; and (3) confirm previously published results that 5-HT and TRH coexist in terminals in the ventral horn of the spinal cord.