Analeptic and antianaleptic effects of naloxone and naltrexone in rabbits.

Naloxone (5 mg/kg), but not naltrexone, shortened the duration of anaesthesia in rabbits pretreated with pentobarbital. This analeptic effect was blocked by atropine, but not by methylatropine; it thus appears that a central cholinergic mechanism is involved. In contrast, smaller doses of both naloxone and naltrexone attenuated the arousal property of thyrotropin releasing hormone (TRH). Naloxone, but not naltrexone, also antagonized the analeptic property of d-amphetamine. In conscious animals naloxone potentiated, whereas naltrexone attenuated, the excitatory effects of TRH and d-amphetamine.     

Thydrotropin releasing hormone: antagonism of pentobarbital in rodents

Thyrotropin releasing hormone (TRH) antagonizes the behavioral and temperature reducing effects of pentobarbital in rodents. The hormone is effective whether given before or after the barbiturate. This antagonism by TRH of the effects of pentobarbital probably does not depend upon thyroid hormone release as L-triiodothyronine administration is ineffective.     

Potentiation of the behavioral effects of pentobarbital, chlordiazepoxide and ethanol by thyrotropin-releasing hormone

Effects of pentobarbital, chlordiazepoxide and ethanol were studied alone and in combination with thyrotropin-releasing hormone (TRH), IM, on punished behavior. Key-peck responses of pigeons were maintained by food presentation under a fixed-interval 3-min schedule in which every 30th response produced shock. Moderate doses of pentobarbital, chlordiazepoxide and ethanol increased punished responding to 150-200% of control values while the higher doses of these drugs almost completely eliminated responding. TRH (0.01-1 mg/kg) had little effect on punished responding and 3 mg/kg produced 50% decreases. Although the lower doses of TRH were without effect when given alone, doses of 0.03 mg/kg and greater markedly potentiated the rate-increasing effects of pentobarbital, chlordiazepoxide and ethanol. Increases in punished responding of 350% were obtained with combinations of TRH and these drugs. The rate-decreasing effects of the sedative-hypnotic and anxiolytic compounds were not reversed by TRH. Potentiation of the behavioral effects of sedative-hypnotic and anxiolytic drugs by TRH suggests that TRH may play an important role in modulating the behavioral effects of these compounds and that combinations of neuroactive peptides with certain psychotherapeutic agents may be of some therapeutic value.     

Thyrotropin-releasing hormone (TRH) and its analog (DN-1417): interaction with pentobarbital in choline uptake and acetylcholine synthesis of rat brain slices

TRH or its analog DN-1417 (gamma-butyrolactone-gamma-carbonyl-L-histidyl-L-proliamide) given 15 min after intravenous (i.v.) administration of pentobarbital (30 mg/kg) markedly shortened the pentobarbital-induced sleeping time in rats. This effect was almost completely abolished by intracerebroventricular pretreatment with atropine methylbromide (20 micrograms/rat), thereby suggesting the involvement of cholinergic mechanism. The action mechanism was investigated using rat brain slices. TRH (10(-6)-10(-4)M) or DN-1417 (10(-7)-10(-5)M) caused significant increases in the uptake of [3H]-choline into striatal slices. TRH(10(-4)M) or DN-1417(10(-5)M) also stimulated the conversion of [3H]-choline to [3H]-acetylcholine in striatal slices. A 30% reduction of acetylcholine synthesis from [3H]-choline in hippocampal slices and a 40% reduction of [3H]-choline uptake in slices of cerebral cortex, hippocampus and hypothalamus were observed in rats pretreated with pentobarbital (60 mg/kg, i.v.). TRH or DN-1417 (20 mg/kg, i.v.) given 15 min after the administration of pentobarbital markedly reversed both of the pentobarbital effects. Direct application of pentobarbital (5 X 10(-4)M) to slices in vitro also caused a 20-40% reduction of [3H]-choline uptake of cerebral cortex, hippocampus and diencephalon. A concomitant application of TRH(10(-4)M) or DN-1417(10(-5)M) and pentobarbital abolished the pentobarbital effect. These results provide neurochemical evidence that the antagonistic effects of TRH and DN-1417 on pentobarbital-induced narcosis are closely related to alterations in the rat brain choline uptake and acetylcholine synthesis, which are considered to be measures of the activity of cholinergic neurons.     

Thyrotropin-releasing hormone (TRH) and its analog (DN-1417): interaction with pentobarbital in oxygen consumption and cyclic AMP formation of rat cerebral cortex slices

Effects of TRH or its analog DN-1417 (gamma-butyrolactone-gamma-carbonyl-L-histidyl-L- prolinamide ) and pentobarbital, alone or in combination, on oxygen consumption and cyclic AMP formation in rat cerebral cortex slices were investigated. The oxygen consumption of rat cerebral cortex slices as measured with a Warburg apparatus, increased linearly over time (0 to 60-min incubation at 37C). Addition of pentobarbital (1 to 7 x 10-4M) inhibited oxygen consumption, in a concentration-dependent manner, up to 45% of control. A concomitant application of DN-1417 (10-5M) or TRH (10-4M) and pentobarbital (5 x 10-4M) led to a partial recovery of the pentobarbital effect. The similar anti-pentobarbital effects were observed with the addition of carbachol (10-4M) or dibutyryl cyclic AMP (10-3M), but not norepinephrine (10-4M) or dopamine (10-4M). DN-1417, TRH, carbachol, norepinephrine or dopamine at 10-4M stimulated cyclic AMP formation in the cerebral cortex slices. Addition of pentobarbital (1 to 7 x 10-4M) inhibited the cyclic AMP formation, in a concentration-dependent manner. DN-1417, TRH or carbachol at 10-4M but not norepinephrine or dopamine at 10-4M significantly reversed the reduction of cyclic AMP formation induced by pentobarbital (5 x 10-4M). Atropine (10-4M) almost completely abolished DN-1417-, TRH- and carbachol-induced cyclic AMP formation in the presence and absence of pentobarbital.     

Effect of thyrotropin-releasing hormone (TRH) on local cerebral glucose utilization, by the autoradiographic 2-deoxy[14C]glucose method, in conscious and pentobarbitalized rats

Effects of TRH and pentobarbital alone, and in combination, on local cerebral glucose utilization of rats were studied by the autoradiographic 2-deoxy[14C]glucose method. TRH (5 mg/kg i.v.) reduced the rate of cerebral glucose utilization slightly in the whole brain. Locally, significant depression was observed in the following structures: frontal and visual cortices, hippocampus Ammon's horn and dentate gyrus, medial and lateral geniculate bodies, nucleus accumbens, caudate-putamen, substantia nigra, pontine gray matter, superior colliculus, superior olivary nucleus, vestibular nucleus, lateral lemniscus and cerebellar cortex. Pentobarbital (30 mg/kg i.v.) produced a marked and diffuse reduction in the rate of glucose utilization throughout the brain. TRH given 15 min after the administration of pentobarbital markedly shortened the pentobarbital sleeping time and caused some reversal of the depression in local cerebral glucose utilization produced by pentobarbital. These effects were almost completely abolished by pretreatment with intracerebroventricular injection of atropine methyl bromide (20 microgram/rat). These results indicate that although TRH acts to cause a reduction in the rate of cerebral glucose utilization, it reverses the depression induced by pentobarbital, via a cholinergic mechanism, in a number of structures, some of which are related to monoaminergic systems and the reticulo-thalamo-cortical activating system.     

Respiratory and cardiovascular effects of thyrotropin-releasing hormone as modified by isoflurane, enflurane, pentobarbital and ketamine

Thyrotropin-releasing hormone (TRH) possesses significant arousing and cardio-respiratory stimulant actions. The effects of a 2 mg/kg i.v. bolus dose of TRH on respiration and systemic hemodynamics were compared in conscious, freely-moving rats and during anesthesia with 4 different anesthetics. Fifty-four male Sprague-Dawley rats weighing 285 +/- 4 g (mean +/- S.E.M.) were divided into 5 groups: conscious, enflurane (2%), isoflurane (1.4%), pentobarbital (8 mg/kg/h i.v.), and ketamine (60 mg/kg/h i.v.). Anesthetized rats were intubated and breathed oxygen or anesthetic/oxygen spontaneously. Aortic blood pressure, heart rate, cardiac output, respiratory rate, arterial blood pH, blood gases, lactate and glucose were measured, and data were collected over a 20 min baseline period and for 130 min post-TRH. TRH increased respiratory rate in all groups; concomitant changes in arterial PCO2 indicated increased minute ventilation in the inhalation agent groups but not in the i.v. anesthetic groups or in the awake group. Significant respiratory depression in the enflurane group was rapidly reversed by TRH. The respiratory stimulant and arousing effects of TRH were smallest with ketamine anesthesia. The hemodynamic responses to TRH were consistent with a pattern of sympathoadrenalmedullary activation and were relatively uniform across groups despite anesthetic-induced alterations in baseline values. TRH or its analogues may prove useful as an analeptic in clinical anesthesia.     

Effects of ether and pentobarbital anesthesia on thyroid function in the rat.

Studies were performed to examine the effect of two anesthetic agents, ether and pentobarbital, on the hypothalamic-pituitary-thyroid function in vivo. In non-anesthetized animals, plasma thyrotropin (TSH) increased rapidly from basal values of 0.1, a peak of 0.49 microng/ml, 25 min after exposure to the cold. Anesthesia with ether during exposure to the cold completely prevented the rise in TSH. During pentobarbital anesthesia, the rise in TSH after exposure to cold was reduced by more than 90%. Even a three minute period of ether anesthesia prior to cold exposure reduced the peak response to cold as well as delayed this response when compared to the untreated group. During two hours of anesthesia with ether, the TSH concentration declined in animals which were fed a low iodine diet at essentially the same rate as in animals on the same diet given an injection of 3 microng of triiodothyronine. Pentobarbital did not suppress TSH at room temperature. The release of thyrotropin after injection of synthetic thyrotropin-releasing hormone (TRH) was greater in animals anesthetized with pentobarbital than in controls and was slightly reduced in ether-anesthetized animals. This difference was observed when thyrotropin was given intraperitoneally or intravenously and the slope of the dose-response curves to TRH showed a flattening of the curve of rats treated with ether and a steeper slope of response in animals anesthetized with pentobarbital. We conclude that pentobarbital inhibited TSH response to cold but did not reduce the resting levels. Ether inhibited the rise of TSH in the cold and lowered the basal levels of TSH in animlas at room temperature. Pentobarbital increased the response to TRH and ether may have reduced the response to TRH.     

The effects of centrally administered neuropeptides on the development of gastric lesions in the rat

Centrally administered neuropeptides were investigated for their effects on the development of gastric lesions in rats. Thyrotropin releasing hormone (TRH), vasoactive intestinal peptide (VIP) and gonadotropin releasing hormone (LHRH) produced gastric lesions acutely, with TRH demonstrating the most pronounced effect in terms of incidence and severity. Ten-fold higher doses of the same peptides administered intravenously produced none or very few gastric lesions. Moreover, pretreatment with atropine partially inhibited their production. Corticotropin releasing factor (CRF) exhibited only mild ulcerogenic effects, and the gastric lesions induced with this peptide developed more slowly than with TRH, VIP and LHRH. Although ulcerogenic in their own right, none of these four neuropeptides significantly potentiated the potent ulcerogenic effects of cold-restraint stress. Since other neuropeptides, including somatostatin, human pancreatic growth hormone releasing factor (hpGRF), substance P, bombesin, and neurotensin, had no demonstrable effects on gastric mucosa, we can conclude that the lesions were not a general effect of intracisternal administration of neuropeptides. The results suggest that within the central nervous system, there are several neuropeptides that play a significant role in the development of gastric lesions via, at least in part, vagal-dependent mechanisms.     

Spontaneously hypertensive rats exhibit supersensitivity to the hypertensive and hyperthermic effects of thyrotropin releasing hormone

The effect of thyrotropin releasing hormone (TRH) on colonic temperature and systolic blood pressure of age-matched spontaneously hypertensive (SHR) and normotensive Wistar-Kyoto (WKY) rats was determined. Administration of TRH produced dose-dependent increases in body temperature and systolic blood pressure. TRH-induced changes in both responses were of greater magnitude in SHR rats compared to WKY rats. The results provide the first evidence that SHR rats exhibit supersensitivity to non-neuroendocrinological effects of TRH and that TRH may play a role in the pathophysiology of elevated blood pressure.     

Behavioral effects of thyrotropin releasing hormone in frontal decorticated rats

A low dose intracerebroventricular injection of thyrotropin releasing hormone (TRH, 100 ng) changed many behavioral responses in the rat. TRH increased locomotion, scratching, body shaking, piloerection, and rearing, but decreased sniffing, and resting. Ablation of frontal neocortex further enhanced the TRH effects on locomotion and resting. A dose effect of TRH (0, 5, 10, 50, 100 ng) to increase general activity was established and the effect was further enhanced by decortication. In our test situations decortication had no effect by itself. Since the TRH effects became much more pronounced without the frontal neocortex it appears that the cortex exerts a powerful inhibitory effect to moderate the TRH effects. The TRH effect does not depend upon the frontal cortex, actually a cortical function is to dampen the TRH effects on various behavioral responses.     

Thyrotropin releasing hormone: autonomic effects upon cardiorespiratory function in endotoxic shock

These studies define potential sites and mechanisms by which thyrotropin releasing hormone (TRH) stimulates cardiorespiratory function in normotensive rats as well as in rats subjected to endotoxic shock. Changes in mean arterial pressure, pulse pressure, heart rate, and respiratory rate were determined in conscious animals following injection of TRH into the lateral, third, or fourth ventricular spaces. Injections of TRH into the third ventricular space resulted in a greater increase in cardiorespiratory variables than did fourth ventricular injection. In endotoxin-treated rats, the cardiorespiratory effects of intracerebroventricular (icv) TRH and its analog MK 771 were assessed. TRH and MK 771 were shown to act within the brain to reverse endotoxic shock hypotension; at the doses used, the pressor effects of these two tripeptides were achieved through selectively different actions upon heart rate and pulse pressure. Adrenal demedullated and sham-operated control rats subjected to endotoxic shock were injected with icv and intravenous (iv) TRH in order to evaluate the potential involvement of sympatho-medullary function in cardiorespiratory responses. The cardiovascular effects of icv TRH were dependent upon adrenal medullary integrity; effects of iv TRH were not. Doses of iv TRH which effectively reverse shock neither altered nociceptive latencies nor interfered with analgesic responses to morphine. Collectively, these studies reinforce the potential therapeutic utility of TRH and its analogs in the treatment of shock and indicate potential sites and mechanisms which mediate these salutary effects.     

The effects of thyrotropin-releasing hormone on cyclic AMP accumulation in rabbit cerebral cortical tissue in the presence and absence of CNS depressants

The $\text{in vitro}$ effects of thyrotropin-releasing hormone on cAMP accumulation in cortical brain slices from rabbits is reported. Incubation of cortical tissue at three concentrations of thyrotropin-releasing hormone (1,2,5nM) had no discernible effects on baseline cAMP levels. When cortical tissue was incubated in the presence of pentobarbital (.5mM) or if cortical tissue was taken from animals pretreated with α-methyl-p-tyrosine (α-MPT), the baseline cAMP accumulation was depressed. This depression could be eliminated by the addition of TRH to the incubation media. Where cortical tissue from atropine-pretreated animals was used or when atropine was added to the incubation media, there was an increase in baseline cAMP accumulation which was unaffected by addition of TRH. These results show that TRH can modify cAMP accumulation in mammalian cortical brain tissue but this ability may only become evident in situations where normal cAMP concentration has been depressed.     

Phylogenetic Distribution and Significance of the Hypothalamic Releasing Hormones

SYNOPSIS. The mammalian hypothalamic releasing factors regulatingthyroid, gonadal and adrenal function as well as growth hormonesecretion have been isolated, characterized and their nucleotidesequences determined. In general, their hypophysiotropic effectsare replicated in lower vertebrates though thyrotropin releasinghormone (TRH) does not appear to stimulate thyroid functionin amphibia and fish. The releasing factors, or peptides structurallyrelated to these substances, are found throughout the CNS ofall vertebrates where they likely function as neurotransmittersor neuromodulators. High concentrations of TRH and other neuralpeptides includingsauvagine, which is related to corticotropinreleasing factor (CRF) and has CRF-like activity, are foundin amphibian skin, a neural crest derived tissue. mRNA extractedfrom the skin of Xenopus laevis was cloned and led to the identityof the DNA sequence of pre-pro TRH. Molecular variants of somatostatinhave been recognized from studies on the pancreatic islets ofthe anglerfish and catfish. Within mammalian species there isheterogeneity of growth hormone releasing factor (GRF), the44 (and 40) amino acid peptides isolated from a human(h) pancreatictumor. In the teleost brain-pituitary, 2 distinct hGRF-likeneuronal systems are present. Additionally, various molecularforms of hGRF exist in the fish brain showing structural changesfrom the human variety.     

Dissociation of tolerance to the analgesic and hypothermic effects of morphine by using thyrotropin releasing hormone

The effects of thyrotropin releasing hormone (TRH) on tolerance to the analgesic and hypothermic effects of morphine were determined in male Swiss Webster mice. The tolerance to morphine was induced by SC implantation of a morphine pellet containing 75 mg morphine free base for 3 days. Subcutaneous injections of TRH (4 mg/kg) twice a day inhibited tolerance to the analgesic effect of morphine, as evidenced by a greater degree of analgesia in TRH treated mice as compared with similarly treated vehicle injected controls. The same treatment, however, failed to modify tolerance to the hypothermic effect of morphine. These effects were produced with alterations in brain or plasma levels of morphine. It is concluded that tolerance to the two pharmacological effects of morphine may involve separate mechanism.     

Thyrotropin releasing hormone antagonizes beta endorphin hypothermia and catalepsy.

Injection of 30 μg β endorphin intraventricularly (ivt) in rats produced an alteration of body temperature, a state of catalepsy, and an increase in antinociceptive latencies. Subsequent ivt injections of 20 μg of thyrotropin releasing hormone (TRH) reversed the ongoing changes in body temperature and catalepsy produced by β endorphin. Since TRH antagonized these effects in hypophysectomized rats, it is implied that these effects of TRH are independent of pituitary-thyroid involvement. In contrast to the above, TRH did $\text{not}$ alter the antinociception produced by β endorphin in either sham-control or hypophysectomized rats. The failure of TRH to antagonize all three of these opiate effects, as well as the inability of TRH to displace bound dihydromorphine from synaptic plasma membranes, suggests that the level of TRH-β endorphin interaction is not at the opiate receptor.     

An improved synthesis of thyrotropin releasing hormone (TRH) and crystallization of the tartrate

An improved synthesis of thyrotropin releasing hormone (TRH), pGlu-His-Pro-NH₂, is reported. Z-pGlu-ONB (N-hydroxy-5-norbornene-2,3-dicarboximide ester) was reacted with H-His-OH to yield a crystalline Z-pGlu-His-OH which was coupled with H-Pro-NH₂ by the $\text{HONB}\text{DCC}$ method to give Z-pGlu-His-Pro-NH₂ as a fine crystal. Hydrogenation of this protected tripeptide yielded pure TRH nearly quantitatively. The optical purity of TRH thus obtained was confirmed by the method L- and D- amino acid oxidase digestion. The crystallization of TRH was achieved as a tartrate, and the properties of the crystalline TRH-tartrate are described.     

Neuroanatomical dissociation of thyrotropin-releasing hormone induced shaking behavior and thermogenic mechanisms

To more clearly characterize the neuroanatomical substrates mediating thyrotropin-releasing hormone (TRH) induced shaking and antagonism of pentobarbital hypothermia, TRH was microinjected into 140 individual sites of the rat forebrain and brainstem. Previously determined threshold dosages of 10 ng TRH for the temperature response and 50 ng TRH for the shaking response were used. A clear distinction in regional sensitivity between the two TRH-induced effects was observed. The shaking response was most consistently observed with microinjection of TRH into the floor of the 4th ventricle and the periventricular posterior diencephalon, including the posterior hypothalamus and rostral periventricular grey. In contrast, the temperature response was most effectively induced by TRH administered in the interpeduncular nucleus and the rostral preoptic region located medial to, and including the diagonal band of Broca. The sensitivity of some brain areas to nanogram doses of TRH supports the possibility that TRH may have a physiological function in the initiation of shaking behavior and/or thermogenesis. If such a function does exist, the brain regions identified in this study as most sensitive to exogenous TRH are likely neuroanatomical substrates for endogenous TRH.     

Effect of bombesin on basal and stimulated secretion of some pituitary hormones in humans

The effect of bombesin (5 ng/kg/min X 2.5 h) on basal pituitary secretion as well as on the response to thyrotropin releasing hormone (TRH; 200 micrograms) plus luteinizing hormone releasing hormone (LHRH; 100 micrograms) was studied in healthy male volunteers. The peptide did not change the basal level of growth hormone (GH), prolactin, thyroid-stimulating hormone (TSH), luteinizing hormone (LH) and follicle-stimulating hormone (FSH). On the contrary, the pituitary response to releasing hormones was modified by bombesin administration. When compared with control (saline) values, prolactin and TSH levels after TRH were lower during bombesin infusion, whereas LH and FSH levels after LHRH were higher. Thus bombesin affects in man, as in experimental animals, the secretion of some pituitary hormones.