The distribution of Histidyl-Proline Diketopiperazine [cyclo(His-Pro)] in discrete rat hypothalamic nuclei

The distribution of Histidyl-Proline Diketopiperazine [cyclo(His-Pro)], a metabolite of thyrotropin-releasing hormone (TRH), was determined by specific RIA in Palkovits micropunch pools derived from discrete hypothalamic nuclei. Highest concentrations of cyclo(His-Pro) were identified in the anterior nucleus (3.5 ng/mg protein) and the paraventricular nucleus (2.95 ng/mg protein), while lower concentrations of cyclo(His-Pro) were seen in the other 6 nuclei. In contrast, TRH concentrations were highest in the ventromedial nucleus pars medialis (3.2 ng/mg protein) and arcuate nucleus (2.7 ng/mg protein). This qualitatively different distribution of cyclo(His-Pro) from that of TRH suggests that not all of cyclo(His-Pro) is derived exclusively from TRH.     

Thyrotropin-releasing hormone: its distribution and metabolism during development in bullfrog (Rana catesbeiana)

The development changes in the metabolism of thyrotropin-releasing hormone (TRH), cyclo (His-Pro) formation from TRH, and the levels of endogenous TRH in frog brain and skin were determined. The results indicated that TRH concentrations were considerably higher in brain than in skin, and in both of these structures TRH content increased significantly following metamorphosis to adulthood. This increase in TRH concentration is probably a reflection of a marked decrease in TRH-metabolism in adult frogs compared to tadpoles. However, the formation of cyclo (His-Pro) from TRH increased during the developmental period reaching to a maximum in adulthood. The possible role of cyclo (His-Pro) in the amphibian developmental process is discussed in relation to our recent observation showing cyclo (His-Pro) inhibition of prolactin secretion.     

High affinity specific binding of the thyrotrophin releasing hormone metabolite histidylproline to rat brain membranes

Histidylproline a metabolite of Thyrotrophin Releasing Hormone specifically binds to both high and low affinity sites in fresh rat brain membranes. Characterisation of the high affinity site under optimal conditions demonstrated an equilibrium dissociation constant (KD) of approximately 9nM and maximum binding capacity of approximately 120 fmols/mg protein. Kinetic analysis of [3H]-His-Pro binding is limited by low binding density, instability of the high affinity site and rapid degradation of the radioligand. The thiol blocking reagent pHydroxymercuriphenylsulphonic acid (HMPS) inhibited [3H]-His-Pro degradation but also reduced binding of the peptide to membranes. The results are discussed with reference to the lack of specific binding sites in brain for the proposed neuropeptide and TRH metabolite cyclo(His-Pro).     

Distribution of histidyl-proline diketopiperazine [cyclo (His-Pro)] and thyrotropin-releasing hormone (TRH) in the primate central nervous system

The concentrations of cyclo (His-Pro) and its precursor, thyrotropin-releasing hormone (TRH) were measured in 47 different loci of monkey brain using specific radioimmunoassays. Cyclo (His-Pro) concentrations were higher than those of TRH in all loci excepting the hypothalamus, where TRH concentration was found to be the highest of all the loci and twice those of cyclo (His-Pro). The high levels of cyclo (His-Pro) were seen within the cerebellar system (inferior olivary nucleus>nucleus interpositus>fastigial nucleus>posterior vermis). The great variations in TRH-cyclo (His-Pro) ratios among different loci suggest that other factors in addition to TRH concentration must play roles in determining the unique distribution pattern of cyclo (His-Pro) in the primate brain.     

Cyclo (His-Pro): Mapping hypothalamic sites for its hypothermic action

Hypothalamic loci of Sprague-Dawley rats were individually injected with cyclo (His-Pro) to determine the sites where that metabolite of thyrotropin-releasing hormone acts to produce hypothermia. There was almost always a positive hypothermic response in the preoptic-anterior hypothalamic area (POA/AHA); injection into the posterior or middle hypothalamic areas or into the hippocampus caused no significant decrease in core temperature. The fact that only injection into the POA/AHA evoked hypothermia suggests that this area is a major hypothalamic site of action of cyclo (His-Pro) in modulating thermoregulation in the rat.     

The biological activities of the neuropeptide histidyl-proline diketopiperazine

Histidyl-proline diketopiperazine [cyclo(His-Pro)] is a recently discovered neuropeptide which is produced by the metabolism of thyrotropin releasing hormone, TRH. This peptide mediates a number of central nervous system activities, some of which can also be observed with TRH, some of which are opposed by TRH and some which are not affected by TRH. Cyclo(His-Pro) also inhibits release of prolactin from pituitary by a process inhibited by TRH.     

Cyclo(His-Pro) and the development of tolerance to the hypothermic effect of ethanol

Acute intraperitoneal administration of ethanol to rats causes a dose-dependent transient hypothermia. On repeated exposure, however, rats develop tolerance to hypothermic effects of ethanol. Cyclo(His-Pro), an endogenous brain peptide, modifies both acute and chronic themomodulatory effects of alcohol. For example, a) acute pretreatment of rats with increasing amounts of cyclo(His-Pro) produces a progressive decrease in ethanol hypothermia, and b) chronic cyclo(His-Pro) administration augments the development of tolerance to hypothermic effects of alcohol. While the mechanism of cyclo(His-Pro) action is not clear, these data are interpreted to suggest that this peptide may play important roles in ethanol intoxication, preference, tolerance, and/or addiction.     

Ontogeny of receptors for thyrotropin-releasing hormone in the rat brain

Developmental changes in the thyrotropin-releasing hormone (TRH) receptor concentrations of hypothalamic and certain extrahypothalamic structures in the rat were evaluated from 2 days after birth until sexual maturity. The maturational pattern of TRH receptors in hypothalamus, striatum and amygdala follow that of the pituitary and all show the same kinetic properties. The developmental pattern also coincides with that of other components of the hypothalamic-pituitary-thyroid axis. In all tissues, the concentration of receptors increased in early life to reach a maximum between 10 and 30 days of age and then decreased gradually to adult values. Development of TRH receptors reflected changes in the number of receptor sites but not in binding affinity, which remained constant and suggests that the TRH receptor is identical in all 4 tissues studied. These results indicate the parallel development of pituitary and central nervous system TRH receptors.     

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 mg/kg 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 (Bmax). 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.     

Regulation of cholecystokinin receptors in the ventromedial nucleus of the hypothalamus: sex steroid hormone effects

Quantitative autoradiography was used to analyze cholecystokinin receptor (CCK-R) binding in the ventromedial hypothalamic nucleus (VMH) of gonadectomized rats treated with estrogen or testosterone. In the rostral VMH, sex steroids did not affect binding levels. In the caudal VMH, in both gonadectomized male and female rats, the levels of CCK-R binding were decreased 24 h after injection of 50 μg of estrogen benzoate but were not changed after injection of 300 μg of testosterone propionate. We hypothesize that these changes in CCK-R binding in the VMH reflect ligand-induced down-regulation that result from an estrogen-facilitation of stimulated CCK release in the VMH.     

Effects of hypothalamic lesion on pancreatic autonomic nerve activity in the rat

The effects of hypothalamic lesions and intravenous glucose infusion on the efferent activity of vagal and splanchnic nerves to the pancreas were studied in anesthetized rats. Lesions of the ventromedial hypothalamic (VMH), the dorsomedial hypothalamic (DMH) and the paraventricular (PVN) nuclei increased vagal and reduced splanchnic nerve activity. Lesion of the lateral hypothalamic area (LHA) decreased pancreatic vagal nerve activity, and produced either increased or decreased activity of pancreatic splanchnic nerve. Intravenous glucose infusion increased activity of the vagal nerve and reduced that of the splanchnic nerve. These glucose responses were influenced by hypothalamic lesions only slightly or not at all. The findings suggest that hypothalamic modulation of pancreatic hormone secretion involves both the parasympathetic and sympathetic nervous systems, and provide evidence that not only the VMH and the LHA but also the DMH and the PVN are involved in this mechanism.     

Neuropeptide-dopamine interactions. II. Cyclo (His-Pro) augmentation of amphetamine- but not apomorphine-induced stereotypic behavior

Histidyl-proline diketopiperazine (cyclo [His-Pro]) not only exists in the basal ganglia of rodents, monkeys, and humans, but also exhibits a variety of biologic activities, some of which appear to be mediated via dopaminergic mechanisms. We investigated the potential modulation by cyclo (His-Pro) of amphetamine- and apomorphine-induced stereotypic behavior, a behavior that is associated with the activation of postsynaptic dopamine receptor. Administration of amphetamine to rats resulted in a dose-dependent increase in stereotypic behavior that was further augmented if animals were pretreated with cyclo (His-Pro). Although apomorphine also led to a dose-related progression in the stereotypic behavior, the apomorphine effects were not modified by cyclo (His-Pro) pretreatment. We conclude that cyclo (His-Pro) either acts indirectly at the presynaptic dopamine site or modulates other neurotransmitters to potentiate actions of amphetamine.     

Neuropeptide-dopamine interactions. I. Dopaminergic mechanisms in cyclo(His-Pro)-mediated hypothermia in rats

Administration of cyclo(His-Pro) to rats produces a dose-dependent hypothermia that is attenuated by dopaminergic antagonists. Chronic treatment with cyclo(His-Pro) potentiates hypothermia induced by apomorphine. These results suggest that cyclo(His-Pro) acts via a dopaminergic mechanism to modulate body temperature.     

Localization and Characterization of Dopamine D1 Receptors in Sheep Hypothalamus and Striatum

Dopamine receptors are pharmacologically grouped as D₁ and D₂ receptors. Previous research in the ewe has shown that central D₁ receptors may have a role in facilitating prolactin release. The aims of this study were therefore to localize and characterize D₁ binding sites in the hypothalamus of sheep. For comparison, a known D₁ receptor-rich tissue (striatum) was also studied. The bioactivities of several D₁ analogues were also assessed for their efficacy in sheep tissue. In vitro autoradiography with [¹²⁵I]-SCH23982 was used to localize D₁ binding sites. The ventromedial hypothalamic nucleus (VMH) displayed moderate levels of specific binding, localized to the medial portion of the nucleus. Low levels of specific binding were seen in the preoptic area, supraoptic nucleus and anterior hypothalamic area. The suprachiasmatic nucleus, median eminence and arcuate nucleus did not show specific binding. As expected the striatum displayed high levels of specific binding. The VMH, preoptic area, median eminence, striatum and anterior pituitary were examined with radioligand binding studies to quantify and characterize D₁ binding sites. Scatchard analysis gave K_D 1.04 nM and B_max 127.4 fmol/mg protein for VMH and K_D 1.99 nM and B_max 454.6 fmol/mg protein for striatum. While specific binding occurred in the preoptic area and median eminence this binding did not show saturation characteristics. Specific binding was not observed in the anterior pituitary. Affinities determined by competitive binding studies showed that the binding sites in both VMH and striatum have the characteristics of a D₁ receptor, that is, high affinity for the D₁ agonists and antagonists, low affinity for dopamine and the serotonergic antagonist ketanserin and extremely low affinity for the D₂ agonists and noradrenaline. Adenylate cyclase studies showed that in the striatum dopamine and the D₁ agonists, fenoldopam and SKF38393, were able to cause significant dose-dependent increases in adenylate cyclase activity. In contrast the D₁ agonist, SKF82958, was inactive in this system. The D₁ antagonists SCH23390 and SCH39166, but not SKF83566, abolished the adenylate cyclase response to 50 μM dopamine. In the VMH the D₁ agonist SKF38393, but not dopamine, stimulated adenylate cyclase activity. In conclusion, these results demonstrate that D₁ binding sites exist within the hypothalamus in the VMH and that these binding sites have the characteristics of D₁ receptors. These receptors are a potential site of action for dopamine in facilitating prolactin release. In addition, the results show that at least for some dopamine analogues, receptor binding affinity does not always correlate with biological activity.     

The involvement of pineal gland and melatonin in immunity and aging: II. Thyrotropin-releasing hormone and melatonin forestall involution and promote reconstitution of the thymus in anterior hypothalamic area (AHA)-lesioned mice.

A stereotactic electrolytic lesion of the anterior hypothalamic area in mice produces a rapid involution of the thymus and a reduction of lymphocytes in the peripheral blood. This effect on the thymus and blood lymphoid compartment can be prevented by postoperational administration of thyrotropin-releasing hormone (TRH) or melatonin. These activities of TRH or melatonin are antagonized by the opioid receptor blocker naltrexone. They do not seem to depend on stimulation of the thyroid gland or of the endogenous opioid system but rather on a direct activity of TRH on thymic targets or binding sites on lymphocytes.     

Hypothalamic control of liver glycogen metabolism in adult and aged rats

The regulatory function of the hypothalamus in the metabolism of liver glycogen was investigated by analyzing the changes in the activities of the rate-limiting enzymes implicated in glycogen breakdown (glycogen phosphorylase) and synthesis (glycogen synthetase), after electrical stimulation of the ventromedial hypothalamic nucleus (VMH) and the lateral hypothalamic nucleus (LH) of rats. Electrical stimulation of the VMH induced rapid and marked increase in the content of the active form of phosphorylase in the liver, but it did not affect the synthetase. Stimulation of the LH, on the other hand, caused an increase in the level of the active form of synthetase, but produced little change in the phosphorylase activity. The concentration of insulin in the portal blood decreased significantly on stimulation of the VMH, but did not change appreciably on stimulation of the LH. The ability of the hypothalamus to control liver glycogen metabolism was found to be impaired in 2-year-old rats: the magnitude and duration of the early increase in active phosphorylase in response to VMH stimulation were reduced, and the response of synthetase to LH stimulation was slow.The results are discussed from the viewpoint that the VMH and LH are the centers for controlling irect neural and neural-hormonal regulations of liver glycogen metabolism. It is concluded that the VMH-splanchnic nerve pathway is an important neural component for controlling glycogenolysis and glucose output by the liver, and that the LH-vagal pathway is an opposing neural component for controlling glycogenesis in the liver.     

Failure of cyclo (His-Pro) to exhibit natriuretic activity

Administration of exogenous cyclo (His-Pro) to dogs has been reported to elicit natriuresis. In contrast, our data fail to show any natriuretic activity of cyclo (His-Pro) in dogs or rats. The possible reasons underlying this discrepancy are discussed.     

Neural network of glucose monitoring system

Glucose-sensitive neural elements exist in the hypothalamus, the nucleus of the solitary tract (NTS) and autonomic afferents from visceral organs such as liver and gastrointestinal tract. Glucose affects neural activity through these central and peripheral chemosensors. Glucose is generally suppressive in the liver, the NTS and the lateral hypothalamic area (LHA), and generally excitatory in the small intestine and ventromedial hypothalamic nucleus (VMH). The hypothalamus is involved in the control of pancreatic hormone secretion through autonomic efferent nerves. Stimulation or lesion of the hypothalamus induces various changes in pancreatic autonomic nerve activity. The VMH, the dorsomedial hypothalamic nucleus and the paraventricular nucleus have inhibitory effects on vagal nerve activity and excitatory effects on splanchnic nerve activity. The LHA is excitatory to the vagal nerve, and both excitatory and inhibitory to the splanchnic nerve. These findings suggest that the neural network of the glucose monitoring system, which also analyzes and integrates information concerning other metabolites and peptides in the blood and cerebrospinal fluid, contributes to regulation of peripheral metabolism and endocrine activity as well as feeding behavior. The physiological function and input-output organization of this network are discussed.     

The biphasic changes in splenic natural killer cell activity following ventromedial hypothalamic lesions in rats

The cytotoxic activity of natural killer (NK) cells in the spleen was measured by a standard 4-h chromium release assay following electrical lesioning of the ventromedial hypothalamic nucleus (VMH) in rats. The splenic NK cell activity of VMH lesioned rats was found to be significantly suppressed at effector: target cell ratios of 100:1 and 50:1 compared with that of sham lesioned rats on day 4 after the lesion. On the other hand, on day 49, the VMH lesioned animals that had become hyperphagic and obese showed an enhancement of splenic NK cell activity compared with sham lesioned animals. The mechanisms of the biphasic change in NK cell activity following the VMH lesions are discussed.