Histamine-releasing effect of a corticotrophin derivative. I. Histamine-releasing effect of a nonadecapeptide in comparison with that of other histamine liberators.

The histamine-liberating properties of a synthetic polypeptide (an a synthetic polypeptide (an alkylprolyl derivative of beta1-19-corticotrophin) were investigated under a variety of experimental conditions. It was found to be more potent in this respect than Compound 48/80, Melittin and Triton X-100. The degree of cell destruction observed in conjunction with the release of histamine induced by C 44 680-Ba suggests that the mode of action of the substance more closely resembles that of Compound 48/80 than that of Triton.     

Influence of histamine releasing agents on gastric acid secretion of isolated bullfrog gastric mucosa.

The influence of histamine releasing agents on gastric acid secretion was studied in isolated bullfrog gastric mucosa preparations. Maximum acid secretory responses in our preparations were obtained by stimulation with tetragastrin (5 X 10(-7) g/ml), histamine (1 X 10(-5) g/ml) and bethanechol (1 X 10(-6) g/ml). Compound 48/80 (1 X 10(-4) g/ml) showed a transient stimulatory action which was followed by a gradual depression of basal acid secretion. The stimulatory phase of compound 48/80 was completely antagonized by burimamide (1 X 10(-5) g/ml), a histamine H2-receptor antagonist. In gastric mucosa preincubated with compound 48/80, the secretagogue action of tetragastrin or bethanechol was not exerted, although this preparation continued to respond to histamine. The effects of Triton X-100, decylamine and polymixin B were quite similar to those of compound 48/80. After pretreatment with compound 48/80, the gastric mucosa preparation became refractory to the stimulatory action of compound 48/80 or Triton X-100. It is thus suggested that endogenous histamine may play an important role in the secretagogue action of tetragastrin and bethanechol.     

Dimethyl sulfoxide inhibits histamine release induced by various chemicals

Dimethyl sulfoxide was found to inhibit histamine release induced by compound 48/80 at concentrations ranging from 0.6 to 10%. At higher concentrations (20 and 40%) the substance caused histamine release by itself. Heat inactivation at 50 degrees C of the mast cells did not prevent this release, suggesting a lytic mechanism for this action and this was further proved by the trypan blue dye exclusion test. Dimethyl sulfoxide was also active in inhibiting histamine release induced by other polyamines such as bradykinin, polymyxin B and protamine, by concanavalin A and dextran, by ionophore A23187 and by three anthracyclines, doxorubicin, daunomycin and epirubicin. In contrast, the substance did not inhibit histamine release induced by a monoamine, chlorpromazine and by the detergent Triton X-100.     

Modulation of histamine release from rat peritoneal mast cells by non-cytotoxic concentrations of the detergents Cremophor El (oxethylated castor oil) and Triton X100. A possible explanation for unexpected adverse drug reactions?

Clinically relevant histamine release caused by drugs and/or their solvents is a well known phenomenon. The mechanisms whereby these reactions occur are largely unknown. It was thought that the solubilizing agents potentiate the histamine release elicited by the drugs. Therefore the ability of the two detergents, Cremophor El and Triton X100, to modulate histamine release from rat peritoneal mast cells was examined. Both detergents were used in concentrations that did not themselves induce histamine release. The addition of the detergents to incubation media containing compound 48/80 (0.1 microgram/ml) elevated the release considerably (48/80 alone = 16.2 +/- 2.1% (n = 3); plus Cremophor El (5%) = 41.1 +/- 3.3% (n = 4); plus Triton X100 (0.02 microliter/ml) = 39.7 +/- 3.9% (n = 3); plus Triton X100 (0.01 microliter/ml) = 33.4 +/- 5.0% (n = 3)). In contrast, histamine release induced by Concanavalin A or the calcium ionophore A 23187 was inhibited by both detergents. Thus low concentrations of detergents appear to have a dual role, with both potentiation and inhibition of histamine release being observed. Surgical patients receive many drugs, some soluble in aqueous solutions, others only with the aid of solubilizing agents. 'Hangover effects' due to different plasma half lives, may therefore cause a seemingly harmless drug to act as a histamine liberator. It is therefore important to examine the action of clinically used solvents on histamine liberation caused by therapeutic agents, in order to gain a further understanding of the reaction mechanisms of adverse reactions to drugs.     

Spectroscopic studies of rat mast cells,mouse mastocytoma cells,and compound 48/80—III: Evidence for a protein binding site for compound 48/80

The interaction of compound 48/80 with mast cells, mastocytoma cells, isolated membranes and liposomes has been studied using electron spin resonance (e.s.r.) and light-scattering techniques. Unlike spin-labeled stearic acid, spin-labeled 48/80 (SL-48/80) bound to mastocytoma cells in a manner that was unaffected by the non-ionic detergent, Triton X-100. In contrast, sodium dodecylsulfate (SDS), a protein denaturant, altered the SL-48/80 binding site(s), causing an apparent increase in the motion of the spin label. Treatment of SL-48/80-labeled mast cells with trypsin also increased the apparent motion of the bound label. Binding of 48/80 to mast cells, mastocytoma cells, erythrocyte ghosts, and isolated mastocytoma cell membranes increased their respective light-scattering properties. In contrast, the Rayleigh scatter of protein-free liposomes was unaffected by 48/80 treatment. Cationic, anionic, and non-ionic detergents also increased the light-scattering properties of mastocytoma cells, in contrast to the effect of several other cationic drugs, including histamine liberators. Increased light-scattering occurred after 48/80 treatment of inhibited mast cells (treated at 5°), indicating that the effect was not a consequence of degranulation. Untreated cells heated above 40° showed a similar increase in light scattering that became irreversible after reaching 45°. This indicates that cells treated with 48/80 resemble those in which the proteins have been denatured by heat. The light-scattering and the e.s.r. data, therefore, are consistent with the hypothesis that 48/80 binds to proteins in biological membranes.     

Erythrocyte membrane stabilization by calcium channel blockers, calmodulin antagonists and scavengers of oxygen free radicals

Calcium channel blockers (nifedipine, verapamil, diltiazem), calmodulin antagonists (trifluoperazine, calmidazolium, compound 48/80) and anti-free radical agents (allopurinol, desferrioxamine, mannitol, L-methionine) were tested for their potency to stabilize human erythrocytes against hypotonic hemolysis. The anti-free radical agents and compound 48/80 did not confer the membrane stabilization. Nifedipine, verapamil, diltiazem, calmidazolium and trifluoperazine at low concentrations, protected the cells from the hypotonic hemolysis while at higher concentrations they caused lysis. Similar biphasic changes were produced by the detergents sodium dodecyl sulphate (SDS) and Triton X-100. The drug concentration-dependency of the biphasic changes in the erythrocytes osmotic fragility produced by calcium channel blockers and calmodulin antagonists was not affected by low concentrations of SDS and Triton X-100. On the other hand, these drugs did not prevent the hemolysis produced by high concentrations of the detergents. The above as well as the observation that the membrane stabilization is conferred only by relatively high concentrations of calcium channel blockers and calmodulin antagonists suggest that membrane stabilization is not responsible for anti-ischemic effects of these agents reported in the literature.     

Characteristics of histamine release from rat mast cells induced by a bracken toxin, braxin A1.

The effect of braxin A1, a new bracken glucoside, on histamine release from isolated rat peritoneal mast cells was studied. Braxin A1 caused the release of histamine in a dose-dependent manner; the release was slow and increased gradually with time, finally reaching a maximum release of 100%. The action of braxin A1 depended on the incubation temperature in the range from 4 degrees C to 49 degrees C, while it was almost abolished at 0 degree C. The action of braxin A1 was unaffected by removing calcium or any inorganic ions from the incubation medium and by the addition of 2,4-dinitrophenol or theophylline. The mast cells exposed to braxin A1 were vitally stained with trypan blue and swelled greatly. The cell swelling was characterized by the protrusion of swollen cytoplasmic granules. The present results for braxin A1 were similar to those for the ionophore X537A except for the extracellular inorganic ion dependency, but they were different from those observed with compound 48/80. These results suggest that braxin A1 releases histamine from mast cells without both exocytosis and membrane lysis, but with a cytotoxic action on cytoplasmic membranes by a different mode of action from that of X537A.     

On the actions of substance P,somatostatin, and vasoactive intestinal polypeptide on rat peritoneal mast cells and in human skin

Substance P (SP), somatostatin (Som), and vasoactive intestinal polypeptide (VIP) induced a concentration-dependent release of histamine from isolated rat peritoneal mast cells. The release of histamine induced by these neuropeptides was inhibited by preincubation of the cells with the SP analogue [D-Pro4,D-Trp7,9,10]-SP4-11 (SP-A) (10 microM), and also by benzalkonium chloride (10 microM). In addition, SP-A inhibited histamine release induced by compound 48/80, whilst that induced by goat anti-(rat-IgE) was unaffected. In human skin, intradermal injection of SP, Som, or VIP produced flare and wheal responses. The flares to all three peptides were inhibited by preinjection of the skin with SP-A (25 pmol), whilst the wheal responses were unaffected. It is concluded that the receptors mediating histamine release and the flare response are similar, and that SP, Som, and VIP are acting at a similar receptor to produce these effects. It is probable that this receptor is also the site of action of compound 48/80.     

Effect of disodium cromoglycate and cyclic AMP-active drugs on cytotoxic histamine release from rat mast cells

Disodium cromoglycate and compounds which elevated levels of cyclic AMP in the mast cell variously inhibited cytotoxic histamine release induced by the surface active agents melittin, Tween 20 and Triton X-100. These results are inconsistent with the postulated effects of the drugs on receptor mediated calcium channels and alternative explanations of their action are considered.     

Involvement of enteric neurones in the response of guinea-pig ileum preparations to metoclopramide.

The role of myenteric neurones in mediating the stimulant effects of metoclopramide in vitro in the guinea-pig ileum has been investigated using the non-ionic surfactant Triton X-100. Histological examination of the ileum 30 days after application of Triton X-100 to the serosal surface demonstrated a marked reduction in the number of ganglion cells and nerve elements in the myenteric plexus. Longitudinal muscle-myenteric plexus (LM-MP) preparations from Triton X-100-treated animals were unresponsive to dimethylphenylpiperazinium and responded poorly or not at all to electrical field stimulation. Metoclopramide (30 microM) elicited small contractions in LM-MP preparations from control and sham-operated animals but failed to contract Triton X-100-treated tissues. However, tissues responded in a similar manner to exogenous acetylcholine (ACh). These results demonstrate the importance of a prejunctional site of action for metoclopramide in this tissue and suggest that contractile responses to the drug are mediated indirectly, probably by increased release of ACh from myenteric neurones.     

The effect of adrenocorticotropin on histamine and 5-hydroxytryptamine secretion from rat mast cells

Characteristics of histamine (Hi) and 5-hydroxytryptamine (5-HT) release from rat peritoneal mast cells in response to the polypeptide adrenocorticotropin (ACTH) were studied. During a 15 min incubation at 37 degrees C, ACTH evoked Hi as well as 5-HT release from rat mast cells at concentrations of 1 X 10(-4) M-1 X 10(-3) M. The release was dose-dependent and very rapid. After 15 sec the amount of the amines released was the same as after 4.5 min. In most experiments, the percentage of Hi release was slightly but significantly higher than the percentage of 5-HT release. Hi and 5-HT release induced by ACTH also took place in a calcium-free medium. However, the release of the amines was decreased when calcium was omitted. Comparison of the effects of ACTH, compound 48/80 and substance P on mast cell secretion showed that ACTH is about 100 times less active then substance P which was in turn about 100 times less active than compound 48/80. When both ACTH and compound 48/80 were used together as liberators , the release was significantly higher than with either liberator alone. Our results indicate that there are receptor sites for the endogenous polypeptide ACTH on the mast cell membrane which mediate Hi and 5-HT release. This release was found to resemble that evoked by the basic secretogogue compound 48/80 but in some aspects to be different from that evoked by substance P.     

Mast cell heterogeneity in man: unique functional properties of skin mast cells in response to a range of polycationic stimuli.

Human mast cell heterogeneity was assessed by histochemical and detailed functional criteria using mast cells isolated from foreskin, uterine myometrium and lung parenchyma. The skin mast cells were histochemically distinct from their counterparts in the other two tissues by being predominantly insensitive to blockage of dye-binding following formalin fixation (ca. 80%). Functionally, a wide range of structurally diverse polycationic compounds induced selective histamine release from the skin mast cells (ca. 10% at top concentrations) although these cells were less responsive to immunological ligands and calcium ionophores when compared with the uterine and lung cells. The basic compounds, polyarginine and histone, proved to be more generalised histamine liberators as compared with their structural analogues, polylysine and protamine sulphate, probably by virtue of their high content of arginine residues and hydrophobic nature (histone). Studies with the anaphylatoxin, C3a, and its analogues 21R and C3ades Arg on skin mast cells emphasized the importance of basic amino acids for histamine-liberating peptides. Skin mast cells also proved more susceptible than their uterine counterparts to lysis by the detergents, Triton X-100 and Tween 20, suggesting that fundamental differences in membrane structure and/or fluidity might account for functional heterogeneity within the human mast cell population.     

Interaction of non-ionic surfactants with hepatic CYP in Prochilodus scrofa

Cytochromes P450 (CYP) constitute a superfamily of hemeproteins that play a vital role in the metabolism of a wide variety of endogenous and xenobiotic compounds. Xenobiotic metabolism and the role of CYP are of particular interest in studies regarding the prevention of the damage caused by chemical pollutants. We investigated, in this study, the interaction of Triton X-100 and Tween 80 with CYP and antioxidant defenses in Curimbatá, a Brazilian fish. Aiming to clarify the effects of non-ionic surfactants in the monooxigenase system of fish through in vitro study, the effects of Triton X-100 and Tween 80 were analyzed using monooxygenases and antioxidant system as experimental model. Total CYP and EROD were strongly inhibited by Triton X-100 and Tween 80 in a concentration-dependent way; the content of CYP was reduced until zero while EROD activity was completely inhibited in the presence of Triton X-100 and more than 40% inhibited in the presence of Tween 80. Each surfactant causes a different effect on each antioxidant enzyme. No effect was detected in SOD activity in the presence of even Triton X-100 or Tween 80. Triton X-100 increase catalase activity, while Tween 80 decreases this enzyme activity. The molecular structure of the surfactants causes the alteration of this system, since they are able to interact with the microsomal protein, especially with monooxigenase's components, altering their conformation and, consequently destroying their function. Our results suggest that surfactants can interact with components of the microsomal system leading to inhibition of CYP. Therefore, CYP activity, which has been used as a biomarker of xenobiotic exposure, should be used as a marker in association with other enzymes.     

Morphological and physiological changes in Tetrahymena pyriformis for the in vitro cytotoxicity assessment of Triton X-100.

Non-ionic surfactants such as Triton X-100 have been widely used in industrial processing and in cleaning products for almost 50 years, being effective and economic emulsifying, wetting agents, dispersants and solubilizers. Cleaning products containing these surfactants are disposed of mainly by discharge into wastewater, which receives biological treatment in wastewater treatment systems. However, surface-active agents interact with eukaryotic cell membranes leading to biological damage at high concentrations. Tetrahymena pyriformis was used here as model organism to assess the effects of Triton X-100 through a series of in vitro cytotoxicity tests. Growth rates and morphological changes were, by their simplicity and reproducibility, the simplest toxicological assays. Cytoskeleton analysis seemed to be related with phagocytosis rate. Viability was evaluated by two different tests. Calcein AM/EthD-1 was used to assess T. pyriformis membrane damage during the 48-h experiment. The colorimetric MTT assay proved to be highly sensitive even at very short periods of Triton X-100 exposure. Tests performed in this study included simple and fast bioassays that provide overall information on the morphological and physiological state of cells exposed to different non-lytic and lytic concentrations of Triton X-100.     

The action of Triton X-100 and sodium dodecyl sulphate on lipid layers. Effect on monolayers and liposomes

The action of two detergents, Triton X-100 and sodium dodecyl sulphate (SDS), on large, unilamellar liposomes was determined as liposome size variation, polydispersity and ability to release a soluble marker from liposomes. Triton X-100 produced stronger effects than SDS. Nevertheless, these differences in behaviour of such detergents could not be deduced from the interaction of the detergents with monolayers of the same composition as liposomes.     

The effect of a new antihistamine drug, Loderix (EGIS-2062), on stimulus-evoked histamine release from rat peritoneal mast cells

The ability of a new, non-sedative antihistamine drug, Loderix (EGIS-2062), to inhibit stimulus-evoked histamine release from rat peritoneal mast cells has been investigated and compared with that of ketotifen (Zaditen). At low concentrations Loderix preincubated with the cells for 10 min prior to the addition of various stimulants (immune aggregates, ionophore A23187 and compound 48/80) produced a concentration-dependent inhibition of histamine release, while at high concentrations it induced the release of histamine. The IC50 values were calculated as 0.2, 15 and 50 microM by using immune aggregate, (rat IgG2 alpha + anti rat IgG) calcium ionophore and 48/80 as stimulants, respectively. At the IC50 level Loderix was more effective than Zaditen (100 times more effective when immune aggregate and 4 times when 48/80 were used for stimulation). Both drugs had a dual effect on mast cells. The morphological observations confirmed the pharmacological action of the drugs, showing also that the histamine release at high concentrations is due to their cytotoxic effect.     

Action of detergents on 3H-dihydroergokriptine binding and localization of alpha-adrenoceptors in synaptosomal membranes

3H-dihydroergokriptine (3H-DHE) binding was carried out in synaptosomal membranes from basal ganglia of the cat. A single type of binding site with Kd 3.7 nM, Hill number = 0.95 and Bmax = 1000 pmol/g protein was found. 3H-DHE bound to alpha-adrenoceptors and not to serotonin or dopamine receptors. At very low concentrations, some detergents enhanced binding, but at higher concentrations of those used (Triton X-100, Nonidet P-40, deoxycholate and digitonin), inhibited the binding of 3H-DHE. After binding to the membrane protein, the 3H-DHE-receptor complex was stable to the action of Triton X-100. At concentrations of Triton X-100. At concentrations of Triton X-100 (0.1--0.2%). in which only the presynaptic membrane disintegrated, the 3H-HDE specific radioactivity was reduced. With a more drastic treatment that disintegrated the postsynaptic membrane, 3H-DHE binding was further reduced. These results suggest that alpha-adrenergic receptors may be localized at both the pre- and postsynaptic membranes of central synapses.     

Adrenergic activity on rat pleural and peritoneal mast cells. Loss of beta-receptors during the purification procedure

Adrenergic agonists inhibit the release of histamine from rat pleural and peritoneal mast cells stimulated with compound 48/80 to a degree dependent on their beta-activity. Isoprenaline takes part in a stereoselective inhibitory action in the range 10(-7)-10(-4) M. Adrenaline induces a similar response pattern, with inhibition at higher concentrations. The response profile, but not the maximum values of inhibition, is clearly dependent on the concentration of the histamine releaser. Noradrenaline by itself is a histamine releaser, no stereoselectivity being observed. In the presence of compound 48/80 it takes part in a non-stereoselective inhibitory reaction at low concentrations. Inhibition of histamine release by isoprenaline was antagonized by 10 or 100 microM propranolol except at the highest isoprenaline concentration (1 mM). Both atenolol and propranolol nullified the inhibitory activity of noradrenaline, but not the increased histamine release it induces at higher concentrations (at least when acting in conjunction with compound 48/80). When rat mast cells are purified through Percoll, a change in their response profiles is observed. Isoprenaline and adrenaline by themselves elicit non-specific release of histamine; with compound 48/80, release is additive in the case of isoprenaline and supra-additive in the case of adrenaline. Results point to the loss of beta-adrenergic inhibitory activity after purification.     

Inhibition of compound 48/80-mediated histamine release from isolated rat mast cells by oosponol-related compounds (4-acyl-isocoumarins).

Oosponol (4-hydroxymethylketone-8-hydroxyisocoumarin) is a metabolic product isolated from Oospora astringens which originated from house dust in a room of an asthmatic patient. The compound and the structurally related isocoumarins were studied to determine the inhibition of histamine release induced by compound 48/80 from isolated rat peritoneal mast cells. The released histamine was assayed by fluorometry. The compounds tested were not observed to release histamine. Some of 4-acyl-isocoumarins inhibited the histamine release at doses less than 10 micrometers, whereas the 3-acyl- and the 4-alkyl-compounds were not effective at doses over 100 microns. The pretreatment of mast cell with the compound for 15 min before the application of compound 48/80 was more effective than the simultaneous administration. The mode of inhibitory action of KIT-302, 4-(4'-carboxy-benzoyl)-isocoumarin, was non-competitive antagonism to compound 48/80 on the mast cells.     

The effect of compound 48/80 on the carbachol-evoked acid secretion of the isolated mouse stomach

The aim of the present work was to determine in the isolated mouse stomach whether the depletion of mast cell histamine by compound 48/80 influences the acid secretion, evoked by carbachol. Each effect of carbachol was compared to the effect of histamine (10(-6) M), applied after each carbachol application. After carbachol and histamine, compound 48/80 was applied twice or three times successively; the second and the third application were not able to evoke any secretion. After 48/80, the applications of carbachol and histamine were repeated and their effects compared to those before the applications of compound 48/80. Three concentrations of the compound were used: 20, 100 and 500 micrograms/ml. The lowest concentration of the histamine liberator did not significantly change the response to carbachol, The highest one reduced it but the effect of histamine was reduced, too. Compound 48/80 in concentration 100 micrograms/ml significantly augmented the secretory effect of carbachol. The possible explanation for this effect is that histamine, liberated from mast cells, is taken up by some other cells in the tissue from where it is liberated by carbachol stimulation.