Method for the rapid determination of norepinephrine,dopamine, and serotonin in the same brain region

A method is presented for the fluorometric analysis of norepinephrine, dopamine and serotonin. This procedure is a combination of an unpublished catecholamine assay developed by Hogans and of the o-phthaldialdehyde serotonin reaction reported by Maickel and Miller [9]. This procedure should greatly facilitate the correlation of neurotransmitter levels in brain regions with changes in behavior produced by experimental manipulations.     

Effects of spermine and spermidine on single brain stem neurones

Spermine and spermidine were applied by iontophoresis to neurones in the brain stem of decerebrate cats and urethane-anaesthetized rats. The main effect was depression of spontaneous activity (35%); excitatory effects (18%) were more common in the rat. The direction of the effect was always the same for both compounds. It is suggested that they may participate in processes which influence neuronal activity.     

A simple,continuous fluorometric assay for HIV protease

Novel fluorogenic substrates for human immunodeficiency viral protease have been developed based on the principle of fluorescence energy transfer. Starting from a p24/p15 cleavage site-derived hexapeptide substrate, Ac-Thr-Ile-Nle-Nle-Gln-Arg-NH₂, incorporation of 2-aminobenzoic acid in place of the acetyl group as the donor and p-NO₂-Phe at the P1′ position as acceptor gave the intramolecularly quenched fluorogenic substrate. Cleavage of the substrate by HIV protease released the fluorescent N-terminal tripeptide from its close apposition to the quenching nitrobenzyl group, resulting in enhanced fluorescence. An automated assay based on 96=well microtiter plates and a fluorometric plate reader have been developed, which allow high throughput of compounds in the search for HIV protease inhibitors.     

Involvement of brain histamine in Δ9-tetrahydrocannabinol tolerance and withdrawal

The involvement of brain histamine (HA) in Δ⁹-tetrahydrocannabinol (Δ⁹-THC) tolerance and dependence was studied in rats. Rats treated for 5 days with Δ⁹-THC (2–6 mg/kg, IV) developed tolerance to the hypothermic effects of the drug. Tolerance also developed over the 5 day period to the decrease in brain regional HA concentrations observed after an acute injection of Δ⁹-THC. Administration of the tricyclic antidepressant drug clomipramine hydrochloride to tolerant rats induced a withdrawal-like behavioural syndrome. Accompanying this behaviour was a fall in HA concentrations of the midbrain, cortex, medulla oblongata/pons and the cerebellum. Administration of Δ⁹-THC, but not of the Δ⁹-THC vehicle, prior to clomipramine challenge attenuated both the intensity of the withdrawal-like syndrome and the reductions in brain regional HA concentration.     

Histamine and histamine methyltransferase in the gastric mucosa of man,pig, dog and cow

Summary High histamine concentrations and histamine methyl transferase activity were demonstrated in the gastric mucosa of man, dog, pig and cow. Modified methods for the determination of histamine and histamine methyltransferase were developed. Histamine was identified by its fluorescence spectrum, by thin-layer chromatography in 8 different solvent systems and by bioassay. Histamine methyltransferase from pig antral mucosa was purified 6-fold by ultra-centrifugation and fractional precipitation by ammonium sulfate. ItsK _m for histamine as substrate was 2.3×10^–5 M, for S-adenosylmethionine 4.3×10^–5 M, the pH-optimum was found to be pH 7.4. Nicotinamide in concentrations up to 1×10^–2 M had no effect on the activity of the enzyme.We thank the Deutsche Forschungsgemeinschaft for a grant (Sonderforschungs-bereich 37).     

Influence of histamine depletion in rat brain on the rotatory nystagmus

Summary Decrease of histamine in brain after decaborane prevents nystagmus caused by horizontal rotation. This occurs independently of the decrease of serotonin and noradrenaline. Inhibition of cholinacetyltransferase did not prevent nystagmus.     

Effects of halothane,enflurane and pentobarbital on brain histamine dynamics in mice

Summary The effects of halothane, enflurane, ketamine and pentobarbital on brain histamine dynamics were examined in mice. Brain histamine and tele-methylhistamine, a predominant metabolite of brain histamine, were simultaneously measured by high-performance liquid chromatography with fluorescence detection. Anaesthesia with the four agents had no effect on brain histamine content. The tele-methylhistamine content significantly increased during 1 h and 2 h anaesthesia with halothane (0.051 mmol/l or 0.076 mol/1) and 2 h anaesthesia with enflurane (0.11 mol/l or 0.16 mol/1). Enflurane and pentobarbital significantly inhibited the histamine depletion induced by -fluoromethylhistidine (50 mg/kg, intraperitoneally), a specific inhibitor of histidine decarboxylase, suggesting that these agents decrease the histamine turnover. However, halothane and ketamine were ineffective in this respect. These results emphasize that various anaesthetics have different influences on brain histamine dynamics. Since there have been findings suggesting that brain histaminergic systems are involved in physiological functions such as regulation of blood pressure, body temperature and hormone secretion, changes in the brain histamine turnover should be given due attention with regard to physiological changes during anaesthesia. Send offprint requests to K. Saeki at the above address     

Effect of stress on brain histamine

Stress of short duration (5 min) resulted in a significant increase in plasma corticosterone level and a significant decrease in the midbrain histamine concentration in rats. Exposure to 15 min stress caused a significant elevation in the hypothalamic histamine level. Stress of longer duration (30 or 60 min) did not affect hypothalamic, cortical or midbrain histamine concentration although plasma corticosterone level remained elevated. Repeated exposures of rats to 15 min stress did not significantly alter histamine concentration of any of the brain regions studied. Plasma corticosterone concentration was only 28% of that observed in animals exposed to single 15 min stress. Present data suggest a role of brain histamine in response to stress.     

Influence of histamine on the serotonergic system of rat brain.

Both histamine and 4-methylhistamine, after intraventricular injection into normal rats, reduced the levels of serotonin and increased those of 5-hydroxyindoleacetic acid in hypothalamus; after injection into tranylcypromine-treated rats, head twitches were induced which were blocked by antiserotonin agents. 2-Pyridylethylamine, an agonist of histamine H1 receptors, neither influenced serotonin level in hypothalamus nor evoked behavioural changes. It is concluded that injected histamine may release serotonin from the hypothalamus and that this produces the behavioural changes.     

Interference with the fluorometric histamine assay by biogenic amines and amino acids

Naturally occurring low molecular weight biogenic amines and amino acids were analyzed for interference in the fluorometric histamine assay. Three mechanisms of interference were detected, mimicking of histamine, suppression of histamine fluorescence and generation of increasing histamine-like fluorescence during excitation or preirradiation of the o-phthaldialdehyde condensation product with daylight or UVA light. The latter effect was seen only with amino acids. Rat peritoneal lavage fluid contained no fluorescence-suppressing substances, but there were considerable amounts of histamine-mimicking compounds which could not be digested with a diamine oxidase, and of substances generating increasing histamine-like fluorescence when exposed to UV light after condensation with phthaldialdehyde. Although butanol extraction was highly effective in selecting histamine over interfering compounds, it was not sufficient to avoid interference. In contrast, Dowex 50 W-X8 ion exchange chromatography added little to separating histamine and interfering substances. Precautions are recommended to avoid the different types of interference when biological samples are analyzed for histamine content with the fluorometric assay.     

Involvement of brain endogenous histamine in the degeneration of dopaminergic neurons in 6-hydroxydopamine-lesioned rats

Previous studies have suggested that brain histamine is involved in the pathogenesis of Parkinson's disease (PD), but the role of endogenous histamine in the degeneration of dopaminergic neurons in the substantia nigra pars compact (SNpc) remains unclear. We aimed to investigate this issue by changing the brain histamine levels by giving histaminergic agents, and administrating histamine receptor antagonists in the PD animal model, i.e. the 6-hydroxydopamine (6-OHDA)-lesioned rat. In saline-treated animals, 6-OHDA infusion produced a progressive increase in apomorphine-induced turning rate and a loss of tyrosine hydroxylase immunoreactive (TH-ir) neurons in the SNpc. Histaminergic agents were given prior and daily for 1, 7 or 14 days after 6-OHDA infusion. Histidine (500 mg/kg, i.p.), a precursor of histamine, increased the turning rate (27% on day 7 and 26% on day 14, respectively; P<0.05) and also the loss of TH-ir neurons, but only on day 1 and 7 (67% vs 47% and 90.4% vs 74% loss, respectively; P<0.05). In contrast, alpha-fluoromethylhistidine (alpha-FMH, 25 microg, i.c.v.), an irreversible inhibitor of histidine decarboxylase (HDC), significantly decreased the turning rate (25% on day 7 and 26% on day 14, respectively; P<0.05) and prevented the loss of TH-ir neurons, also only on day 1 and day 7 (28% vs 47% and 58% vs 74% loss, respectively; P<0.05). In addition, the histamine H(1) receptor antagonist pyrilamine (5 microg, i.c.v.), but not the H(2) receptor antagonist cimetidine (5 microg, i.c.v.), also decreased the turning rate (38% on day 7 and 21% on day 14, respectively; P<0.05) and prevented the loss of TH-ir neurons on day 1 and day 7 (38% vs 51% and 60% vs 78% loss, respectively; P<0.05). On day 14 after 6-OHDA lesion, there were no significant differences in the number of TH-ir neurons among all the different treatment groups. Taken together, these findings indicate that endogenous histamine may accelerate the degeneration of dopaminergic neurons via its H(1) receptor, while attenuation of histamine transmission may play a protective role on it in the early stage of development of 6-OHDA lesioned PD rats.     

Single and repeated air blast stress and brain histamine

Exposure of rats to air blasts for 1, 5 and 15 min resulted in a significant increase in plasma corticosterone level and in the hypothalamic histamine concentration. Midbrain histamine content was increased after 1 and 5 min of exposure but cortical histamine increased following 1 min of exposure only. Stress of longer duration (30 mins did not significantly affect histamine concentration in any of the three brain regions investigated, although plasma corticosterone level remained very significantly (14.5-fold) elevated. Repeated exposure of rats to air blasts of 15 min duration resulted in a significant elevation of hypothalamic histamine concentration while midbrain and cortical histamine was not significantly altered. Plasma corticosterone level was again very significantly (10-fold) increased. Present results suggest the involvement of brain histamine in the response to stress.     

Uptake and disposition of putrescine, spermidine, and spermine by rabbit lungs

The pulmonary uptake and accumulation of the three polyamines in intact, ventilated, and perfused rabbit lungs was investigated. Lungs were perfused using Krebs-Ringer bicarbonate buffer with albumin in which putrescine, spermidine, or spermine were included at an initial concentration of 10(-3), 10(-2), 10(-1), 5, 10, or 20 mM. At a 5 mM concentration of spermidine and spermine, the uptake by isolated lungs reached a steady state equilibrium in 20-30 min of perfusion. This did not occur for putrescine, which showed linear uptake throughout the entire period of the 60-min perfusion. The lung uptake of putrescine for all perfusate concentrations was greater than that of spermidine or spermine, but all three showed concentration-dependent linear uptake. In the presence of harmaline (1 mM) and ouabain (1 mM), isolated perfused rabbit lungs showed a decrease in uptake of putrescine although no effect was seen for spermidine and spermine. Perfusate containing decreased sodium showed no effect on putrescine uptake by isolated rabbit lungs, but there was a significant increase in the uptake of spermidine and spermine. Significant uptake of all three polyamines was also observed when incubated separately with rabbit lung slices for 60 min. HPLC analysis of lung, the perfusate samples, lung slices, and the incubation medium after a 60-min incubation did not indicate the presence of metabolites of these polyamines. Likewise, the analysis of the lung homogenate incubated with polyamines did not show any metabolites confirming the absence of detectable pulmonary metabolism.(ABSTRACT TRUNCATED AT 250 WORDS)     

An anti-immobility effect of spermine in the forced swim test in mice

BackgroundSpermine is one of the naturally occurring ligands that influence the function of the N-methyl-d-aspartate receptor. Similar to other endogenous polyamines present in micromolar concentration in the brain, it may play a role in the modulation of depression. Thus, the present study investigated the suggested antidepressant effect of spermine.MethodsThe mouse forced swim test (FST) was used as a reliable tool that allowed us to determine the antidepressant activity.ResultsSpermine, administered intracerebroventricularly (icv), significantly and dose-dependently reduced the immobility time in the FST within the dose range of 5–20 μg without changing the spontaneous locomotor activity. The pre-treatment of the animals with ifenprodil (an antagonist of the polyamine binding site of the NMDA receptor), given intraperitoneally at a dose of 20 mg/kg, thoroughly reversed the anti-immobility effect of spermine (5 μg, icv).ConclusionOur preliminary study revealed the anti-immobility activity of centrally administered spermine in the FST in mice, with a probable involvement of the polyamine-binding site at the NMDA receptor complex.     

Iontophoretic studies of histamine and histamine antagonists in the feline vestibular nuclei.

The activity of single neurons in the vestibular neuronal complex of midcollicular decerebrate, decerebellectomized cats were recorded and their responsiveness to iontophoretically applied histamine and other agents determined. The majority of the cells tested were inhibited by iontophoresis of histamine while 24% were excited by this agent. Neurons exhibiting inhibitory responses were widely distributed throughout the four vestibular nuclei and adjacent reticular formation whereas excitatory responses to histamine were obtained mainly in the region of the lateral vestibular nucleus. The H2-receptor blocking agents metiamide and cimetidine were examined as to their actions on spontaneously firing cells and cells affected by histamine. Metiamide was selective in blocking histamine-induced inhibition but not excitation while cimetidine was ineffective in blocking either response. These results suggest that histamine has both inhibitory and excitatory actions on brain stem neurons and metiamide is an effective antagonist of histamine-induced inhibition.     

Specific,irreversible antagonism of histamine receptors in the isolated guinea-pig vas deferens by 4(5)-[5-(4-azido-2-nitroanilino)ethyl] imidazole,a photoaffinity analog of histamine

Photolysis of 4(5)-[2-(4-azido-2-nitroanilino)ethyl] imidazole (AAH) in organ chambers containing isolated guinea-pig vas deferens resulted in an irreversible antagonism of histamine-induced contractions of the smooth muscle. A similar antognism was found in the presence of non-photolyzed AAH, and was reversible if not irradiated. After photolysis, AAH had no effect on tissue responses to KCl, norepinephrine and MgATP; responses to acetylcholine were only slightly reduced. The tissues were protected from the antagonistic effect of photolyzed AAH if diphenhydramine, an H₁-histamine receptor antagonist, was also present during photolysis. Conversely, histamine provided no such protection while cimetidine, an H₂-histamine receptor antagonist, was weakly protective. The results indicate that AAH is a specific and irreversible histamine photoaffinity antagonist. It satisfies the chemical and pharmacological criteria for such an antagonism. The antagonism appears to result from a covalent attachment of the compound to an H₁-histamine receptor-antagonist binding site.     

Effect of diphenhydramine on stress-induced changes in brain histidine decarboxylase activity,histamine and plasma corticosterone levels

Exposure of rats to platform stress induced a significant elevation in hypothalamic histamine levels. Air blast-stress resulted in a significant increase in hypothalamic histamine concentration and in histidine decarboxylase activity. No significant changes were noted either in the enzyme activity or in histamine levels in the midbrain or cortex of stressed rats. In the nonstressed rats, diphenhydramine (7.5 mg/kg intragastrically), a H₁-receptor antagonist, did not influence histidine decarboxylase activity or histamine concentration in any of the three brain regions investigated. However, diphenhydramine pretreatment prevented the increase in histidine decarboxylase activity induced by air blasts. In untreated rats, plasma corticosterone levels were significantly elevated following either platform stress (4.5-fold) or air blasts (7.8-fold). A significant increase was also noted in saline and diphenhydramine-treated animals following these stressors, however, the increase in saline or diphenhydramine treated rats following air blasts was significantly less than that seen in untreated stressed controls.     

Histamine and histamine receptors in pathogenesis and treatment of multiple sclerosis

Multiple sclerosis (MS) is an autoimmune disease associated with chronic inflammatory demyelination of the central nervous system (CNS). Due to disease complexity and heterogeneity, its pathogenesis remains unknown and despite extensive studies, specific effective treatments have not yet been developed. The factors behind the initiation of the inflammatory reactions in CNS have not been identified until now. MS is considered as a complex disease depending on genetic as well as environmental factors. Experimental autoimmune encephalomyelitis (EAE) is the preferential experimental rodent model for MS. Histamine [2-(4-imidazole) ethylamine] is a ubiquitous inflammatory mediator of diverse physiological processes including neurotransmission, secretion of pituitary hormones, and regulation of the gastrointestinal and circulatory systems which can modulate immune responses. Histamine functions are mediated through four G-protein coupled receptors that are named H1-H4 receptor. Histamine is implicated as an important factor in pathophysiology of MS and EAE. It has been shown that histamine can change the permeability of blood brain barrier, which leads to elevation of infiltrated cells in CNS and neuroinflammation. In contrast, there are evidence that show the protective role of histamine in MS and its animal model, EAE. In this review, we try to clarify the role of histamine in pathogenesis of MS, as well as we evaluate the efficacy of histamine receptors agonists and antagonists in treatment of this disease.