Thyrotropin-releasing Hormone Analog Research Articles

Altered Glucose and Insulin Responses to Brain Medullary Thyrotropin-Releasing Hormone (TRH)-Induced Autonomic Activation in Type 2 Diabetic Goto-Kakizaki Rats

Insulin secretion is impaired in type 2 diabetes (T2D). The insulin and glucose responses to central autonomic activation induced by excitation of brain medullary TRH receptors were studied in T2D Goto-Kakizaki (GK) rats. Blood glucose levels in normally fed, pentobarbital-anesthetized GK and nondiabetic Wistar rats were 193 and 119 mg/100 ml in males and 214 and 131 mg/100 ml in females. Intracisternal injection (ic) of the stable TRH analog RX 77368 (10 ng) induced significantly higher insulin response in both genders of overnight-fasted GK rats compared with Wistar rats and slightly increased blood glucose in female Wistar rats but significantly decreased it from 193 to 145 mg/100 ml in female GK rats. RX 77368 (50 ng) ic induced markedly greater glucose and relatively weaker insulin responses in male GK rats than Wistar rats. Bilateral vagotomy blocked ic RX 77368-induced insulin secretion, whereas adrenalectomy abolished its hyperglycemic effect. In adrenalectomized male GK but not Wistar rats, ic RX 77368 (50 ng) dramatically increased serum insulin levels by 6.5-fold and decreased blood glucose levels from 154 to 98 mg/100 ml; these changes were prevented by vagotomy. GK rats had higher basal pancreatic insulin II mRNA levels but a lower response to ic RX 77368 (50 ng) compared with Wistar rats. These results indicate that central-vagal activation-induced insulin secretion is susceptible in T2D GK rats. However, the dominant sympathetic-adrenal response to medullary TRH plays a suppressing role on vagal-mediated insulin secretion. This unbalanced vago-sympathetic activation by medullary TRH may contribute to the impaired insulin secretion in T2D.

Thyrotropin-Releasing Hormone (TRH) Analogues That Exhibit Selectivity to TRH Receptor Subtype 2

Thyrotropin-releasing hormone (TRH) analogues in which the C-2 position of the imidazole ring of the centrally placed histidine residue is substituted with various alkyl groups were synthesized and studied as agonists for TRH receptor subtype 1 (TRH-R1) and subtype 2 (TRH-R2). Several analogues were found to be selective agonists for TRH-R2 exhibiting no activation of TRH-R1. For example, analogue 4 (R= c-C3H5) was found to activate TRH-R2 with a potency (EC50) of 0.41 microM but did not activate TRH-R1 (potency > 100 microM). This study describes the first discovery of TRH-R2-specific agonists and provides impetus to design predominately CNS-effective TRH peptides.

Novel Neuroprotective Tripeptides and Dipeptides

It has long been recognized that thyrotropin-releasing hormone (TRH) and certain TRH analogues are neuroprotective in a variety of animal models of CNS trauma. In addition to these neuroprotective actions, TRH and most TRH analogues have other physiological actions that may not be desirable for treatment of acute injury, such as analeptic, autonomic, and endocrine effects. We have developed a series of dual-substituted TRH analogues that have strong neuroprotective actions, but are largely devoid of these other physiological actions. In addition, we have developed a family of cyclized dipeptides (diketopiperazines), structurally somewhat related to a metabolic product of TRH, that appear even more effective as neuroprotective agents in vitro and in vivo, and may have nootropic properties. Here, we review these novel tripeptide and dipeptide compounds.

Thyrotropin-releasing hormone in the dorsal vagal complex stimulates pancreatic blood flow in rats

Central administration of thyrotropin-releasing hormone (TRH) enhanced pancreatic blood flow in animal models. TRH nerve fibers and receptors are localized in the dorsal vagal complex (DVC), and retrograde tracing techniques have shown that pancreatic vagal nerves arise from the DVC. However, nothing is known about the central sites of action for TRH to elicit the stimulation of pancreatic blood flow. Effect of microinjection of a TRH analog into the DVC on pancreatic blood flow was investigated in urethane-anesthetized rats. After measuring basal flow, a stable TRH analog (RX-77368) was microinjected into the DVC and pancreatic blood flow response was observed for 120 min by laser Doppler flowmetry. Vagotomy of the several portions, or pretreatment with atoropine methyl nitrate or N G-nitro- l-arginine-methyl ester was performed. Microinjection of RX-77368 (0.1–10 ng) into the left or right DVC dose-dependently increased pancreatic blood flow. The stimulation of pancreatic blood flow by RX-77368 microinjection was eliminated by the same side of cervical vagotomy as the microinjection site or subdiaphragmatic vagotomy, but not by the other side of cervical vagotomy. The TRH-induced stimulation of pancreatic blood flow was abolished by atropine or N G-nitro- l-arginine-methyl ester. These results suggest that TRH acts in the DVC to stimulate pancreatic blood flow through vagal-cholinergic and nitric oxide dependent pathways, indicating that neuropeptides may act in the specific brain nuclei to regulate pancreatic function.

Neuroprotective effects of novel small peptides in vitro and after brain injury

Thyrotropin-releasing hormone (TRH) and TRH analogues have been reported to be neuroprotective in experimental models of spinal cord injury and head injury. We have previously shown that a diketopiperazine structurally related to the TRH metabolite cyclo-his-pro reduces neuronal cell death in vitro and in vivo. Here we report the neuroprotective activity of other cyclic dipeptides in multiple in vitro models of neuronal injury and after controlled cortical impact (CCI) in mice. Using primary neuronal cultures, three novel dipeptides were compared to the previously reported diketopiperazine as well as to vehicle controls; each of the compounds reduced cell death after direct physical trauma or trophic withdrawal. Two of these peptides also protected against glutamate toxicity and β-amyloid-induced injury; the latter also strongly inhibited glutamate-induced increases in intracellular calcium. Treatment with each of the test compounds resulted in highly significant improvement of motor and cognitive recovery after CCI, as well as markedly reducing lesion volumes as shown by high field magnetic resonance imaging. DNA microarray studies following fluid percussion induced traumatic brain injury (TBI) in rats showed that treatment with one of these dipeptides after injury significantly down-regulated expression of mRNAs for cell cycle proteins, aquaporins, cathepsins and calpain in ipsilateral cortex and/or hippocampus, while up-regulating expression of brain-derived neurotrophic factor, hypoxia-inducible factor and several heat-shock proteins. Many of these mRNA expression changes were paralleled at the protein level. The fact that these small peptides modulate multiple mechanisms favoring neuronal cell survival, as well as their ability to improve functional outcome and reduce posttraumatic lesion size, suggests that they may have potential utility in clinical head injury.

Central thyrotropin-releasing hormone increases hepatic cyclic AMP through vagal-cholinergic and prostaglandin-dependent pathways in rats

Central neuropeptides play roles in many physiologic regulations through the autonomic nervous system. We have demonstrated that central thyrotropin-releasing hormone (TRH), one of neuropeptides, induces a stimulation of hepatic proliferation through vagal-cholinergic pathways. Since cAMP is known to play an important role in the hepatic proliferation, effect of central TRH on hepatic cAMP was investigated. Rats were intracisternally injected with either a TRH analog, RX-77368 (1–100 ng), or saline. The liver was removed 2–72 h after the TRH analog and hepatic cAMP content was determined by radioimmunoassay. In some experiments, pretreatment with hepatic vagotomy, atropine methyl nitrate, or 6-hydroxydopamine (6-OHDA) was performed. Hepatic cAMP was dose-dependently increased by intracisternal TRH analog (5–100 ng) with a peak response occurring 12 h postinjection. The central TRH-induced increase in hepatic cAMP was abolished by vagotomy, atropine and indomethacin, but not by 6-OHDA. Intravenous injection of the TRH analog (10 ng) did not affect hepatic cAMP. These results demonstrate that TRH acts in the brain to increase hepatic cAMP through vagal-cholinergic and prostaglandin-dependent pathways, suggesting that central TRH modulates hepatic functions through cAMP-mediated signaling pathways.

Novel small peptides with neuroprotective and nootropic properties

The tripeptide thyrotropin-releasing hormone (TRH) and/or related analogues have shown neuroprotective activity across multiple animal trauma models as well as in a small clinical trial of spinal cord injury. The metabolic product of TRH (cyclo-his-pro) retains physiological activity. We have developed a number of novel cyclic dipeptides that are structurally similar to cyclo-his-pro, and have examined their neuroprotective activity across multiple in vitro models of neuronal injury and after traumatic brain injury (TBI) in rodents. Four such compounds were found to reduce cell death after trophic withdrawal or traumatic injury in primary neuronal cultures; two also protected against glutamate or beta-amyloid neurotoxicity. All compounds significantly improved motor and cognitive recovery after controlled cortical impact injury in mice, and markedly reduced lesion volumes as shown by high field magnetic resonance imaging. Further, compound 35b, which is being developed for clinical trials, also showed considerable neuroprotection after fluid percussion induced TBI in rats, and improved cognitive function after daily administration in chronically brain injured rats. At a mechanistic level, the drugs attenuate both apoptotic and necrotic cell death in primary neuronal cultures, markedly reduce intracellular calcium accumulation after injury, and limit changes in mitochondrial membrane potential and associated cytochrome c release. In addition, microarray studies show that 35b reduces transcriptional changes after injury for a number of genes (and proteins) that may be associated with secondary injury, including cell cycle genes, aquaporins and cathepsins. It also upregulates brain-derived neurotrophic factor (BDNF), heat shock proteins (HSP) and hypoxia inducible factor (HIF). Thus, these novel dipeptides have multipotential actions that make them candidates for the treatment of both acute and chronic neurodegeneration.

Central vagal stimulation activates enteric cholinergic neurons in the stomach and VIP neurons in the duodenum in conscious rats

The influence of central vagal stimulation induced by 2 h cold exposure or intracisternal injection of thyrotropin-releasing hormone (TRH) analog, RX-77368, on gastro-duodenal enteric cholinergic neuronal activity was assessed in conscious rats with Fos and peripheral choline acetyltransferase (pChAT) immunoreactivity (IR). pChAT-IR was detected in 68%, 70% and 73% of corpus, antrum and duodenum submucosal neurons, respectively, and in 65% of gastric and 46% of duodenal myenteric neurons. Cold and RX-77368 induced Fos-IR in over 90% of gastric submucosal and myenteric neurons, while in duodenum only 25–27% of submucosal and 50–51% myenteric duodenal neurons were Fos positive. In the stomach, cold induced Fos-IR in 93% of submucosal and 97% of myenteric pChAT-IR neurons, while in the duodenum only 7% submucosal and 5% myenteric pChAT-IR neurons were Fos positive. In the duodenum, cold induced Fos in 91% of submucosal and 99% of myenteric VIP-IR neurons. RX-77368 induces similar percentages of Fos/pChAT-IR and Fos/VIP-IR neurons. These results indicate that increased central vagal outflow activates cholinergic neurons in the stomach while in the duodenum, VIP neurons are preferentially stimulated.

Taltirelin Improves Motor Ataxia Independently of Monoamine Levels in Rolling Mouse Nagoya, a Model of Spinocerebellar Atrophy

To examine the relationship between motor ataxia and monoamine levels in the central nervous system, the contents and concentrations of noradrenaline (NA), dopamine (DA) and serotonin (5-HT) in the cerebellum, brain stem and spinal cord were measured in rolling mouse Nagoya (RMN), a murine model of spinocerebellar atrophy. The tissue weight of the cerebellum and spinal cord, but not that of the brain stem was significantly lower in RMN than in the control group. In RMN, the NA content of the brain stem and spinal cord, but not the cerebellum were decreased relative to the control, and the concentration of NA in the spinal cord was also lower, but not significant. The DA and 5-HT contents in each tissue did not differ from those of the control, but the concentrations of monoamines, except for DA, were elevated in the brain stem and spinal cord in RMN. In particular, the concentrations of NA, DA and 5-HT in the cerebellum were significantly increased in RMN. Repeated administration of tartilerin hydrate, an analog of thyrotropin-releasing hormone, improved the ataxia of RMN, and elicited no obvious changes in either monoamine content or concentration of cerebellum, brain stem and spinal cord. These results indicate that the concentration of DA, as well as NA and 5-HT, increased in the RMN cerebellum, and that tartilerin improves the motor function of these mice via mechanisms other than changes in the levels of NA, DA and 5-HT in the central nervous system.

Evaluation of early gastric mucosal permeability induced by central thyrotropin-releasing hormone administration

Accumulating evidence suggests that central thyrotropin-releasing hormone (TRH) administration induces gastric erosion 4 h after administration through the vagal nerves. However, early changes in the gastric mucosa during these 4 h have not been described. To assess early changes in the gastric mucosa after intracisternal injection of a stable TRH analog, pGlu-His-(3,3'-dimethyl)-ProNH2 (RX-77368), we measured the blood-to-lumen 51Cr-labeled EDTA clearance and examined the effects of vagotomy, atropine, omeprazole, and hydrochloric acid (HCl) on RX-77368-induced mucosal permeability. A cytoprotective dose of RX-77368 (1.5 ng) did not increase mucosal permeability. However, higher doses significantly increased mucosal permeability. Permeability peaked within 20 min and gradually returned to control levels in response to a 15-ng dose (submaximal dose). Increased mucosal permeability was not recovered after a 150-ng dose (ulcerogenic dose). This increase in permeability was inhibited by vagotomy or atropine. Intragastric perfusion with HCl did not change the RX-77368 (15 ng)-induced increase in permeability, but completely inhibited the recovery of permeability after the peak. Pretreatment with omeprazole did not change the RX-77368 (15 ng)-induced increase in permeability, but quickened the recovery of permeability after the peak. These data indicate that the RX-77368-induced increase in permeability is mediated via the vagal-cholinergic pathway and is not a secondary change in RX-77368-induced acid secretion. Inhibited recovery of permeability on exposure to an ulcerogenic RX-77368 dose or on exposure to HCl plus a submaximal dose of RX-77368 may be crucial for the induction of gastric mucosal lesions by central RX-77368 administration.

Centrally Acting and Metabolically Stable Thyrotropin-Releasing Hormone Analogues by Replacement of Histidine with Substituted Pyridinium

Metabolically stable and centrally acting thyrotropin-releasing hormone (TRH) analogues were designed by replacing the central histidine with substituted pyridinium moieties. Their analeptic and acetylcholine-releasing actions were evaluated to assess their potency as central nervous system (CNS) agents. A strong experimental connection between these two CNS-mediated actions of the TRH analogues was obtained in subject animals. The analogue 3-(aminocarbonyl)-1-(3-[2-(aminocarbonyl)pyrrolidin-1-yl]-3-oxo-2-[[(5-oxopyrrolidin-2-yl)carbonyl]amino]propyl)pyridinium (1a) showed the highest (TRH-equivalent) potency and longest, dose-dependent duration of action from a series of homologous compounds in antagonizing pentobarbital-induced narcosis when administered intravenously in its CNS-permeable prodrug form (2a) obtained via reduction of the pyridinium moiety to the nonionic dihydropyridine. The maximum change in hippocampal acetylcholine concentration upon perfusion of the pyridinium-containing tripeptides into the hippocampus of rats was also achieved with 1a. No binding to the endocrine TRH receptor was measured for the TRH analogues reported here; therefore, our design afforded a novel lead for centrally acting TRH analogues. We have also demonstrated the benefits of the prodrug approach on the pharmacokinetics and brain uptake/retention of pyridinium-containing TRH analogues (measured by in vivo microdialysis sampling) upon systemic administration.

Protective effect of central thyrotropin-releasing hormone analog on cerulein-induced acute pancreatitis in rats

Central neuropeptides play a role in many physiological functions through the autonomic nervous system. We have recently demonstrated that central injection of a thyrotropin-releasing hormone (TRH) analog increases pancreatic blood flow through vagal and nitric oxide-dependent pathways. In this study, the central effect of a TRH analog on experimental acute pancreatitis was investigated in rats. Acute pancreatitis was induced by two intraperitoneal injections of cerulein (40 μg/kg) at 1-h interval. Either stable TRH analog, RX 77368 (5–100 ng), or saline was injected intracisternally 15 min before the first cerulein injection under ether anesthesia. Serum amylase level was measured before and 5 h after the first cerulein injection. Pancreatic wet/dry weight ratio and histological changes were also evaluated. Intracisternal TRH analog inhibited cerulean-induced elevation of serum amylase level, increase in pancreatic wet/dry weight ratio and pancreatic histological changes, such as interstitial edema, inflammation and vacuolization. The pancreatic cytoprotection induced by central TRH analog was abolished by subdiaphragmatic vagotomy and N G-nitro- l-arginine-methyl ester ( l-NAME), but not by 6-hydroxydopamine (6-OHDA). Intravenous administration of the TRH analog did not influence cerulein-induced acute pancreatitis. These results indicate that the TRH analog acts in the central nervous system to protect against acute pancreatitis through vagal and nitric oxide-dependent pathways.

Pyroglutamyl peptidase I and prolyl endopeptidase in human semen: increased activity in necrozoospermia

Thyrotropin-releasing hormone (TRH) and its analogues have been reported to have important functions in human semen. In the present paper, we have characterized the activity of the TRH-degrading enzymes pyroglutamyl peptidase I and prolyl endopeptidase in the fluid and prostasomes of human semen and in subcellular fractions of the corresponding sperm. Enzymatic activities were measured fluorimetrically using β-naphthylamine derivatives as substrate. Activity associated with both enzymes was detected in seminal fluid and in the prostasome fraction, as well as in soluble and particulate sperm subcellular fractions. Pyroglutamyl-peptidase I activity presented highest levels in the particulate sperm fraction, whereas the activity of prolyl endopeptidase was maximal in the soluble sperm fraction. In addition, we compared the activity of both enzymes in different seminal fractions in normozoospermic, fertile men and in subfertile patients with different abnormalities revealed by spermiogram analysis (astenozoospermia, necrozoospermia and teratozoospermia). The activities of pyroglutamyl peptidase I and prolyl endopeptidase in necrozoospermia were found to be higher in the corresponding soluble and particulate sperm fractions, respectively, with respect to those measured in normozoospermic semen. The results of the present study indicate that these enzymes may participate in regulating the levels of seminal TRH analogues and in mediating sperm death associated with necrozoospermia.

Effect of central thyrotropin-releasing hormone on pancreatic blood flow in rats

Central neuropeptides play a role in physiological regulation through the autonomic nervous system. Thyrotropin-releasing hormone (TRH) is a neuropeptide distributed throughout the central nervous system and acts as a neurotransmitter to regulate gastric and hepatic functions through vagal-cholinergic pathways. In this study, the central effect of TRH on pancreatic blood flow was investigated in urethane-anesthetized rats. Pancreatic blood flow was determined by laser Doppler flowmetery. After measurement of basal blood flow, a stable TRH analog, RX 77368 (1–50 ng) or saline was injected intracisternally. Pancreatic blood flow was observed for 120 min thereafter. In some experiments, pretreatnent with atropine methyl nitrate (0.15 mg/kg, i.p.), N G-nitro- l-arginine-methyl ester (10 mg/kg, i.v.), or 6-hydroxydopamine (6-OHDA;180 mg/kg, i.p.), or subdiaphragmatic vagotomy was performed. Intracisternal injection of TRH analog dose-dependently increased pancreatic blood flow with a peak response occurring 30 min after injection. The stimulatory effect of TRH analog on pancreatic blood flow was blocked by vagotomy, atropine, and N G-nitro- l-arginine-methyl ester, but not by 6-hydroxydopamine. Intravenous administration of the TRH analog did not influence pancreatic blood flow in the same animal model. These results indicate that TRH acts in the central nervous system to stimulate pancreatic blood flow through vagal-cholinergic and nitric oxide-dependent pathways.

Expression of thyrotropin-releasing hormone receptor in immortalized beta-cell lines and rat pancreas

Thyrotropin-releasing hormone (TRH), a hypothalamic tripeptide, is expressed in pancreatic islets at peak levels during the late gestation and early neonate period. TRH increases insulin production in cultured beta-cells, suggesting that it might play a role in regulating pancreatic beta-cell function. However, there is limited information on TRH receptor expression in the pancreas. The aim of the present study was to explore the distribution of the TRH receptor in the pancreas and its function in pancreatic beta-cells. TRH receptor type 1 (TRHR1) gene expression was detected by RT-PCR and verified by Northern blotting and immunoblotting in the beta-cell lines, INS-1 and betaTC-6, and the rat pancreatic organ. The absence of TRH receptor type 2 expression in the tissue and cells indicated the tissue specificity of TRH receptor expression in the pancreas. The TRHR1 signals (detected by in situ hybridization) were distributed not only in islets but also in the surrounding areas of the pancreatic ductal and vasal epithelia. The apparent dissociation constant value for the affinity of [(3)H]3-methyl-histidine TRH (MeTRH) is 4.19 in INS-1 and 3.09 nM in betaTC-6. In addition, TRH induced epidermal growth factor (EGF) receptor phosphorylation with a half-maximum concentration of approximately 50 nM, whereas the high affinity analogue of TRH, MeTRH, was 1 nM. This suggested that the affinity of TRH ligands for the TRH receptor influences the activation of EGF receptor phosphorylation in betaTC-6 cells. Our observations suggested that the biological role of TRH in pancreatic beta-cells is via the activation of TRHR1. Further research is required to identify the role of TRHR1 in the pancreas aside from the islets.

Thyrotropin-releasing hormone in the dorsal vagal complex stimulates hepatic blood flow in rats.

Central administration of thyrotropin-releasing hormone (TRH) enhances hepatic blood flow in animal models. TRH nerve fibers and receptors are localized in the dorsal vagal complex (DVC), and retrograde tracing techniques have shown that hepatic vagal nerves arise mainly from the left DVC. However, nothing is known about the central sites of action for TRH to elicit the stimulation of hepatic blood flow. The effect of microinjection of a TRH analogue into the DVC on hepatic blood flow was investigated in urethane-anesthetized rats. After measuring basal flow, a stable TRH analogue (RX-77368) was microinjected into the DVC and hepatic blood flow response was observed for 120 minutes by laser Doppler flowmetry. Either left or right cervical vagotomy or hepatic branch vagotomy was performed 2 hours before the peptide. Microinjection of RX-77368 (0.5-5 ng) into the left DVC dose-dependently increased hepatic blood flow. The stimulation of hepatic blood flow by RX-77368 microinjection into the left DVC was eliminated by left cervical and hepatic branch vagotomy but not by right cervical vagotomy. By contrast, microinjection of RX-77368 into the right DVC did not significantly alter hepatic blood flow. These results suggest that TRH acts in the left DVC to stimulate hepatic blood flow through the left cervical and hepatic vagus, indicating that neuropeptides may act in the specific brain nuclei to regulate hepatic function.

Thyrotropin-releasing hormone in the dorsal vagal complex stimulates hepatic blood flow in rats

Central administration of thyrotropin-releasing hormone (TRH) enhances hepatic blood flow in animal models. TRH nerve fibers and receptors are localized in the dorsal vagal complex (DVC), and retrograde tracing techniques have shown that hepatic vagal nerves arise mainly from the left DVC. However, nothing is known about the central sites of action for TRH to elicit the stimulation of hepatic blood flow. The effect of microinjection of a TRH analogue into the DVC on hepatic blood flow was investigated in urethane-anesthetized rats. After measuring basal flow, a stable TRH analogue (RX-77368) was microinjected into the DVC and hepatic blood flow response was observed for 120 minutes by laser Doppler flowmetry. Either left or right cervical vagotomy or hepatic branch vagotomy was performed 2 hours before the peptide. Microinjection of RX-77368 (0.5-5 ng) into the left DVC dose-dependently increased hepatic blood flow. The stimulation of hepatic blood flow by RX-77368 microinjection into the left DVC was eliminated by left cervical and hepatic branch vagotomy but not by right cervical vagotomy. By contrast, microinjection of RX-77368 into the right DVC did not significantly alter hepatic blood flow. These results suggest that TRH acts in the left DVC to stimulate hepatic blood flow through the left cervical and hepatic vagus, indicating that neuropeptides may act in the specific brain nuclei to regulate hepatic function. (H epatology 2003;38:1500-1507.)

Involvement of calcitonin gene-related peptide and capsaicin-sensitive afferents in central thyrotropin-releasing hormone-induced hepatic cytoprotection

The involvement of capsaicin-sensitive afferent neurons and calcitonin gene-related peptide (CGRP) in central thyrotropin-releasing hormone (TRH)-induced hepatic cytoprotection was investigated in rats. Both systemic capsaicin pretreatment and intravenous administration of CGRP receptor antagonist, human CGRP-(8–37), completely abolished the protective effect of intracisternal TRH analog (RX-77368; p-Glu-His-(3,3′-dimethyl)-Pro-NH 2, 5 ng) against carbon tetrachloride (CCl 4)-induced acute liver injury, assessed by serum alanin aminotransferase levels and histological changes. These data demonstrate the involvement of capsaicin-sensitive afferent neurons and CGRP in central TRH-induced hepatic cytoprotection.

Studies on the transport of thyrotropin-releasing hormone (TRH) analogues in Caco-2 cell monolayers.

The transport mechanisms of thyrotropin-releasing hormone (TRH) and its pharmacologically active analogues ((3-methyl-His(2))TRH (MeTRH), taltirelin, montirelin, azetirelin) across Caco-2 cell monolayers were characterized. The results of kinetic analysis showed a linear relationship between the concentration (over the range 0.5-10 mM) and apical-to-basolateral transport rate of these agents. The permeability coefficients (P(app)) of these agents were not substantially different from each other, and their P(app) ratios of the basolateral-to-apical over the apical-to-basolateral transport were close to one (0.73-1.23). The cellular transport of [(3)H]MeTRH at low concentrations (3-15 nM) showed a linear relationship between the concentration and transport rate. The transport of [(3)H]MeTRH in Caco-2 cell monolayers was neither affected by TRH nor TRH analogues, and there was little difference in P(app) values between [(3)H]MeTRH and [(14)C]mannitol. The cell-per-medium ratio of [(3)H]MeTRH in the cellular uptake experiment was similar to the value of [14C]mannitol. A large excess of TRH and MeTRH did not significantly influence cell-per-medium ratios of [(3)H]MeTRH in Caco-2 cell monolayers. The k'(IAM) value, which represents lipophilicity, was decreased in the following order: montirelin > taltirelin > TRH > azetirelin, and the values varied from 0.234 to 1.028. These results indicate that a paracellular passive diffusion may be the major route for the transport of TRH and its analogues in Caco-2 cell monolayers.

Protective effect of central thyrotropin-releasing hormone on carbon tetrachloride-induced acute hepatocellular necrosis in rats

Background/Aims: Thyrotropin-releasing hormone (TRH) acts in the brain to stimulate hepatic proliferation and blood flow through vagal-muscarinic and prostaglandin-mediated pathways. Hepatic blood flow and prostaglandins are well recognized as cytoprotective factors for liver damage, and central TRH is known to play a role in gastric cytoprotection. The effect of central TRH on carbon tetrachloride (CCl 4)-induced acute hepatocellular necrosis was investigated in rats. Methods: Male fasted rats were injected with either TRH analog, RX 77368 (1–10 ng), or vehicle intracisternally, and CCl 4 (2.0 ml/kg) was injected subcutaneously 60 min later. Acute hepatocellular necrosis was assessed by serum hepatic enzymes and histological changes 24 h after CCl 4. Results: Intracisternal TRH dose-dependently inhibited elevation of serum alanine aminotransferase level induced by CCl 4. Intracisternal TRH reduced CCl 4-induced hepatic histological changes. The cytoprotective effect of central TRH on CCl 4-induced acute hepatocellular necrosis was abolished by hepatic branch vagotomy, atropine, indomethacin and N G-nitro- l-arginine methyl ester, but not by 6-hydroxydopamine. Intravenous TRH did not influence CCl 4-induced acute hepatocellular necrosis. Conclusions: These results suggest that the cytoprotective effect of central TRH on acute hepatocellular necrosis is mediated through vagal-muscarinic, and prostaglandin- and nitric oxide-dependent pathways.