tetrodotoxin and Anaphylaxis

1. In sensitized guinea-pigs, the effects of gamma-aminobutyric acid (GABA) and GABAmimetic drugs have been investigated on tracheal segments contracted by cumulative application of an allergen (ovoalbumin, OA) and on serosal mast cells. The same drugs have also been tested on activation of alveolar macrophages isolated from unsensitized guinea-pigs. 2. Superfusion with GABA (1-1000 microM) reduced the contraction intensity of tracheal strips. The effect of GABA (100 microM) was not affected by the carrier blockers, nipecotic acid and beta-alanine (300 microM each). It was mimicked by the GABAB agonist (-)-baclofen (100 microM) but not 3-aminopropanephosphinic acid (100 microM, 3-APA). The GABAA agonist, isoguvacine (100 microM) did not exert any effect. GABA (10 microM)-induced inhibition of tracheal contractions was reduced by the GABAB antagonist, 2-hydroxysaclofen (100 microM, 2-HS), but not by the GABAA antagonist, bicuculline (30 microM). 3. The reduction in contraction intensity induced by GABA (100 microM) was prevented by a 40 min preincubation of tracheal strips with capsaicin (10 microM), but not tetrodotoxin (TTX, 0.3 microM). The effect of GABA (1000 microM) was absent after preincubation with indomethacin (2.8 microM) but unmodified when nordihydroguaiaretic acid (NDGA, 3.3 microM) was used. Finally, removal of the epithelium prevented the GABA effect. 4. Anaphylactic histamine release from serosal mast cells isolated from sensitized animals was not affected either by GABA (10-1000 microM) or the selective receptor agonists (-)-baclofen (0.1-1000 microM) and isoguvacine (10-1000 microM). The release of platelet-activating factor (PAF) from alveolar macrophages stimulated by formyl-Met-Leu-Phe (FMLP; 1 microM) was modified neither by GABA (100 microM)nor by (-)-baclofen (100microM).5. In conclusion, these data show that GABA can inhibit allergic phenomena in the guinea-pig airways through activation of GABAB receptors. An involvement of neuropeptidergic sensory structures is suggested but a role for epithelial cells and arachidonate metabolites is not definitely proved.

Topics: Anaphylaxis; Animals; beta-Alanine; Bronchial Hyperreactivity; Capsaicin; Drug Interactions; Epithelial Cells; GABA Agonists; GABA Antagonists; gamma-Aminobutyric Acid; Guinea Pigs; Histamine Release; Macrophage Activation; Macrophages, Alveolar; Male; Mast Cells; Muscle Contraction; Muscle, Smooth; N-Formylmethionine Leucyl-Phenylalanine; Nipecotic Acids; Ovalbumin; Platelet Activating Factor; Proline; Receptors, GABA; Tetrodotoxin; Trachea

Antigen challenge of sensitized rats leads to delayed gastric emptying, but the mechanism (gastroparesis or prolonged trituration) and mediators are unknown.. Hooded Lister rats were sensitized by intraperitoneal injection of egg albumin as antigen, and control rats were sham-sensitized. On day 14, antral manometric and antroduodenal myoelectric activities in sensitized and sham-sensitized rats were recorded in response to antigen challenge, and the contractility of gastric antral circular muscle strips (mucosa intact) in standard tissue baths was measured in response to antigen or other agents.. In vivo, the intragastric antigen challenge of sensitized (but not sham-sensitized) rats provoked diarrhea, reduction in the antral motility index, and disruption of the duodenal migrating motor complex. In vitro, antigen induced a tonic contraction of antral circular muscle segments from sensitized animals. Doxantrazole, but not disodium cromoglycate, inhibited antigen-induced contraction. Whereas histamine, 5-hydroxytryptamine, prostaglandins, leukotrienes, and platelet-activating factor contracted gastric muscle strips, neither specific antagonists, prostaglandin synthase, or 5-lipoxygenase inhibitors inhibited antigen-induced contraction. Tetrodotoxin increased antigen-induced contraction; however, the antigen-induced contraction was unaffected by atropine, guanethidine, or NG-nitro-L-arginine methyl ester.. Food protein-induced, immunoglobulin E-mediated delayed gastric emptying in sensitized rats is associated with a transient reduction in gastric antral contractions. Antigen-induced contraction appears to be under nonadrenergic, noncholinergic, nonnitroxinergic inhibitory neural control.

Topics: Albumins; Anaphylaxis; Animals; Dietary Proteins; Gastrointestinal Motility; Histamine Antagonists; Indomethacin; Lipoxygenase Inhibitors; Mast Cells; Muscle Contraction; Ovum; Platelet Activating Factor; Rats; Rats, Inbred Strains; Serotonin Antagonists; Stomach; Tetrodotoxin

In vivo uptake of the probe 51Cr-labeled EDTA from the jejunum of egg albumin (EA)-sensitized rats was compared with controls at baseline and after intraluminal antigen challenge. Probe recovery in blood was 60-80% greater in sensitized animals during the baseline period, suggesting that sensitization resulted in increased intestinal permeability. Sensitized, but not control, rats demonstrated a 15-fold increase in 51Cr-EDTA uptake after intraluminal antigen; no change occurred with an unrelated protein. Macromolecular recovery was also enhanced in sensitized animals, since serum levels of immunoreactive EA were elevated 14-fold compared with controls. Antigen challenge was accompanied by biochemical (protease release) and morphological (reduced numbers) evidence of mast cell degranulation in sensitized rats. The neurotoxin tetrodotoxin (applied directly to ligated jejunal segments) inhibited EA-induced uptake of 51Cr-EDTA and antigen. In isolated jejunum from sensitized rats, tetrodotoxin reduced secretory responses to luminal, but not serosal, antigen. These results indicate that neural factors may influence the uptake of molecules from the gut lumen during intestinal anaphylaxis.

Topics: Anaphylaxis; Animals; Cell Membrane Permeability; Electric Conductivity; Electrophysiology; Epithelium; Hypersensitivity; Immunoglobulin E; Jejunum; Male; Membrane Potentials; Muscle, Smooth; Ovalbumin; Rats; Rats, Sprague-Dawley; Tetrodotoxin

1. Exogenous vasoactive intestinal polypeptide (VIP) infused into the pulmonary artery of isolated and ventilated lungs of guinea-pigs decreased, in a dose-dependent fashion (1.0-10.0 nmol), airway resistance and thromboxane B2 (TXB2, the stable hydrolysis product of TXA2) release in the perfusion medium. Prostacyclin (PGI2) synthesis, as reflected by the release of its stable hydrolysis product 6-oxo-PGF1 alpha, was unaffected. Pretreatment with the 5-lipoxygenase inhibitor BWA4c (3.5 x 10(-5) M) did not modify the bronchodilatory effect of VIP or its inhibitory action on TXB2 release. 2. Basal release of immunoreactive VIP from perfused lungs decreased from an initial value of 0.96 +/- 0.10 ng min-1 (mean +/- s.e.mean) in the first 2 min to an average of 0.58 +/- 0.10 ng min-1 in the following 15-20 min. 3. Antigen challenge with ovalbumin (0.1%) in sensitized lungs caused an anaphylactic reaction in 45% of tested lungs, concomitant with a 5 fold increase in both VIP and TXB2 release. Tetrodotoxin pretreatment (10(-6) M) reduced basal VIP release by > 80% and abolished the VIP increase observed during anaphylaxis, without modifying TXB2 release or the bronchoconstrictor response. 4. Indomethacin (10(-6) M) inhibited TXB2 synthesis and release by > 90%, delayed the bronchoconstrictor response and blunted the increased VIP release during lung anaphylaxis, without influencing basal VIP release. 5. The 5-lipoxygenase inhibitor BWA4c (3.5 x 10(-5) M) blunted the increase of TXB2 and VIP release from guinea-pig lung and attenuated the bronchoconstrictor response following ovalbumin challenge. 6. The administration of exogenous VIP as a continuous infusion (10-8 M) attenuated the bronchoconstriction and the release of cyclo-oxygenase metabolites following antigen challenge.7. Acetylcholine (10-6-l0-5 M) infused into the pulmonary artery induced a dose-dependent bronchoconstriction not associated with enhanced VIP or TXB2 release.8. The TXA2 mimetic U-46619 (0.1-1.0 nmol) caused dose-dependent increases in airway resistance,concomitant with an up to 10 fold increase in VIP release. VIP inhibited arachidonate-induced in vitro aggregation of washed rabbit platelets in a dose-dependent manner over a dose range 10-8 10-6 M.Despite the antiaggregatory effect of VIP, TXB2 and PGE2 synthesis was reduced only to a minor extent,and there was no redirection of arachidonate metabolism from TXA2 to PGE2, indicating that VIP does not act as a TX synthase inhibitor in vitro.

Topics: 15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5,13-dienoic Acid; Acetylcholine; Anaphylaxis; Animals; Antigens; Benzeneacetamides; Bronchoconstriction; Guinea Pigs; Hydroxamic Acids; In Vitro Techniques; Indomethacin; Lipoxygenase Inhibitors; Lung; Male; Muscle, Smooth; Ovalbumin; Platelet Aggregation; Prostaglandin Endoperoxides, Synthetic; Prostaglandin-Endoperoxide Synthases; Rabbits; Radioimmunoassay; Tetrodotoxin; Thromboxane B2; Thromboxanes; Vasoactive Intestinal Peptide; Vasoconstrictor Agents

The role of cholinergic neurons in mediating chloride secretion in anaphylaxis was assessed in muscle-stripped segments of distal colon from guinea pigs immunized to bovine milk. beta-Lactoglobulin evoked a concentration-dependent increase in short-circuit current (Isc) in immune, but not nonimmune, tissues. The Isc response to beta-lactoglobulin was reduced by piroxicam, pyrilamine, and cimetidine. Tetrodotoxin and atropine reduced the Isc response to beta-lactoglobulin in immune animals, whereas mecamylamine and ICS 205-930 were ineffective. beta-Lactoglobulin evoked a concentration-dependent increase in acetylcholine (ACh) release in immune, but not nonimmune, animals. In immune tissues after challenge with beta-lactoglobulin, ACh release paralleled the change in Isc. Piroxicam, cimetidine plus pyrilamine, or a combination of piroxicam, cimetidine, and pyrilamine significantly reduced the release of ACh after beta-lactoglobulin challenge. Histamine, dimaprit, and prostaglandins E2 evoked an increase in ACh release. These results suggest that beta-lactoglobulin releases prostaglandins and histamine probably from mast cells. Secretory responses that occur when immune animals are challenged with beta-lactoglobulin result, in part, from activation of cholinergic neurons that utilize muscarinic synapses for transfer of signals to the epithelium.

Topics: Acetylcholine; Anaphylaxis; Animals; Cholinergic Antagonists; Colon; Dinoprostone; Electrophysiology; Guinea Pigs; Histamine; Histamine Antagonists; Intestinal Diseases; Lactoglobulins; Male; Neuroimmunomodulation; Neurons; Parasympathetic Nervous System; Piroxicam; Serotonin Antagonists; Tetrodotoxin

To investigate the role of mast cells in transport abnormalities during intestinal anaphylaxis, we examined responses to antigen in isolated intestinal preparations from ovalbumin-sensitized genetically mast cell-deficient WBB6F1-W/Wv (W/Wv) mice and congenic normal WBBGF1(-)+/+ (+/+) mice. Changes in ion transport (primarily secretion of chloride ions) were indicated by increases in short-circuit current (Isc). In tissues from +/+ mice, antigen caused increases in Isc which were significantly inhibited by antagonists to histamine (diphenhydramine) and serotonin (ketanserin), by a cyclooxygenase inhibitor (piroxicam) and by a neurotoxin (tetrodotoxin). In preparations from W/Wv mice, antigen-stimulated responses were approximately 30% of that in +/+ mice and were inhibited only by piroxicam. Responses to electrical transmural stimulation of nerves were approximately 50% in W/Wv versus +/+ mice, and were inhibited by antagonists of mast cell mediators in +/+ but not W/Wv mice. Reconstitution of mast cells in W/Wv mice by intravenous injection of +/+ bone marrow cells restored the normal responses to both antigen and nerve stimulation. Our results indicate that mast cell-dependent mechanisms are primarily responsible for the ion secretion associated with intestinal anaphylaxis, but that other cells are also involved. In addition, our data provide evidence for the functional importance of bidirectional communication between nerves and mast cells in the regulation of ion transport in the gastrointestinal tract.

Topics: Anaphylaxis; Animals; Biological Transport; Bone Marrow Transplantation; Diphenhydramine; In Vitro Techniques; Intestinal Diseases; Intestinal Mucosa; Mast Cells; Mice; Mutation; Piroxicam; Serotonin; Tetrodotoxin

Antigen challenge of jejunal epithelium from rats sensitized to egg albumin induces an active Cl- secretory process secondary to release of mucosal mast cell mediators. The present study was designed to define the relative role of these mast cell mediators and the enteric nervous system in the transport abnormalities associated with intestinal anaphylaxis. Net ion transport of stripped jejunal tissue from sensitized and sham-treated animals was studied in Ussing chambers. The Cl- secretory response induced by egg albumin during intestinal anaphylaxis was similar to that after addition of 5-hydroxytryptamine (5-HT), histamine, and prostaglandins D2 and E2 to jejunal tissue. Cinanserin, a 5-HT2-receptor antagonist, virtually abolished the response to 5-HT and totally abolished the response to egg albumin. Methysergide, a 5-HT1-receptor antagonist had no effect on either response. Indomethacin, an inhibitor of prostaglandin synthesis, significantly inhibited the 5-HT and egg albumin response. Diphenhydramine, an H1-receptor antagonist and cimetidine, an H2-receptor antagonist both significantly inhibited the histamine response but neither altered the response to egg albumin. Atropine, an anticholinergic, and tetrodotoxin, a nerve blocker, did not inhibit the antigen induced anaphylactic response. These results indicate that 5-HT, acting through 5-HT2 receptors is largely responsible for the transport abnormalities seen in intestinal anaphylaxis induced by egg albumin while prostaglandins appear to play a partial role. The findings do not support a role for the enteric nervous system for the egg albumin induced changes in Cl- secretion.

Topics: Anaphylaxis; Animals; Biological Transport; Cyclooxygenase Inhibitors; Electric Stimulation; Female; Histamine; Histamine Antagonists; Intestinal Diseases; Intestines; Ions; Mast Cells; Ovalbumin; Prostaglandins; Rats; Serotonin; Serotonin Antagonists; Tetrodotoxin

Net ion transport by jejunum of rats immunized against Trichinella spiralis on challenge with parasite-derived antigen was measured in Ussing chambers as a rapidly expressed, biphasic rise and fall (phase I and II) in short-circuit current (delta Isc). This delta Isc is triggered by mucosal anaphylaxis. Our objective is to identify mast cell-derived substances that mediate the epithelial response. Antigenic challenge of sensitized jejunum caused the release of 5-hydroxytryptamine (5-HT), histamine, and prostaglandin E2 (PGE2). The antigen-induced phase I response was mimicked by exogenous 5-HT or histamine and blocked by pretreatment of tissue with 5-HT and histamine H1-antagonists; the phase II response was mimicked by exogenous PGE2 and blocked by an inhibitor of prostaglandin synthesis. Atropine and tetrodotoxin significantly blunted the phase I response as well as the delta Isc caused by exogenous 5-HT or histamine while only slightly reducing the phase II response and not affecting the delta Isc induced by PGE2. Results support the conclusion that 5-HT, histamine, and PGE2 mediate the antigen-induced change in Isc through direct and neurally mediated stimulation of jejunal epithelium.

Topics: Anaphylaxis; Animals; Atropine; Biological Transport; Chlorides; Dinoprostone; Epithelium; Histamine Release; Hypersensitivity; Immunization; Intestine, Small; Ions; Jejunum; Male; Mast Cells; Prostaglandins E; Rats; Rats, Inbred Strains; Serotonin; Tetrodotoxin; Trichinella

Calcium seems to have two opposing effects on histamine secretion from mast cells. A rise in the cytosol calcium concentration initiates the chain of reactions leading to histamine secretion. On the other hand, calcium appears to have a regulatory role, limiting the secretion. Removal of cell surface calcium enhances histamine secretion. The present work demonstrates an inhibitory effect of calcium in the medium, using low concentrations of compound 48/80 as the secretagogue. Histamine secretion in response to compound 48/80 primarily utilizes intracellular calcium. When low concentrations of compound 48/80 were used (usually 20-50 ng/ml), calcium (1 mM) inhibited the secretion, the inhibition being more pronounced as the pH was increased from 6.5 to 8.5. The higher pH conceivably promotes the binding of calcium to the phospholipids in the cell membrane. Calcium at this site seems to depress the efflux of calcium from the intracellular stores to the cytosol. The possibility that the removal of calcium from the cell surface causes increased sodium permeability was considered. However, the sodium channel blocker tetrodotoxin (10(-5) M) was equally ineffective in influencing histamine release in the presence and absence of calcium, indicating that a change of sodium permeability was not involved. Antigen-induced (anaphylactic) histamine secretion depends mainly on extracellular calcium, although some secretion occurs in a calcium-free medium. Addition of calcium alone to the medium caused only slight increase in the secretion, but when both phosphatidylserine and calcium were added histamine secretion was remarkably stimulated, apparently through the effect of phosphatidylserine on calcium transport across the plasma membrane.

Topics: Anaphylaxis; Animals; Calcium; Histamine Release; Hydrogen-Ion Concentration; In Vitro Techniques; p-Methoxy-N-methylphenethylamine; Phosphatidylserines; Rats; Tetrodotoxin

Inhibitory effect of GABA on anaphylactic histamine release in vitro is not mimicked by 2-aminoethansulphonic acid (taurine), an aminoacid unrelated to GABA neuro-transmission. Tetrodotoxin (TTX) 6 X 10(-7) M, a concentration known to block neuronal mechanism but not to modify muscle membrane and anaphylactic histamine release, strongly prevented the inhibition caused by GABA in the Schultz-Dale reaction and in anaphylactic histamine release. The inhibitory effect of GABA on anaphylactic reaction in vitro thus appears to be specific for this aminoacid and is neurogenic in nature, in that it requires integrity of neuronal mechanisms.

Topics: Anaphylaxis; Animals; gamma-Aminobutyric Acid; Guinea Pigs; Histamine Release; In Vitro Techniques; Male; Tetrodotoxin

Arrhythmogenic effects of anaphylaxis and histamine were studied in guinea pigs by measuring changes of transmembrane potentials from isolated papillary muscles. Antigenic challenge of preparations obtained from passively sensitized animals induced ventricular automaticity in 1/3 of the experiments. The arrhythmogenic effects of cardiac anaphylaxis could be reproduced by exogenous histamine. Abnormal automaticity was associated with a stable diastolic membrane potential in most of the ventricular fibres and only occasionally ectopic pacemaker potentials were observed in fibres near the ventricular septum. The effective refractory period, maximum rate of depolarization of the action potential and electrical threshold were not significantly changed in ventricular anaphylaxis but in the presence of histamine the refractory time was shortened. Ventricular arrhythmias induced by histamine were increased at high extracellular Ca2+ concentrations and inhibited by Mn2+ and D 600 but were only moderately antagonized by tetrodotoxin. Pretreatment with reserpine had no effect on the abnormal automaticity. Spontaneous ventricular activity caused by histamine was markedly inhibited by the histamine H2-receptor antagonist burimamide and also by the antiarrhythmic drug, prajmalium bitartrate. The H1-receptor antagonist brompheniramine, hydrocortisone and propranolol had little or no antiarrhythmic effect.

Topics: Action Potentials; Anaphylaxis; Animals; Anti-Arrhythmia Agents; Arrhythmias, Cardiac; Calcium; Female; Guinea Pigs; Heart; Histamine; In Vitro Techniques; Male; Manganese; Myocardial Contraction; Reserpine; Tetrodotoxin

1. The mast-cell distribution in the various layers of the ileum has been described.2. Histamine-content, anaphylactic histamine-release and the anaphylactic dose-response curve of the full-thickness ileum and of the longitudinal muscle strip have been measured and compared.3. Tetrodotoxin 10(-7) g/ml had no obvious effect on the anaphylactic dose-response curve of either preparation. This suggests that the plexus is not of any great importance in the Schultz-Dale reaction.4. Exposure of the longitudinal muscle strip to octylamine 10(-3) g/ml for 1 min reduced the mast cell content by 95-100%. After this treatment the dose-response curve to antigen was eliminated, although the muscle still responded to small doses of histamine and to anaphylactic mediators. Pretreatment of antibody with octylamine did not impair passive sensitization and subsequent response to antigenic challenge. This suggests that the classical Schultz-Dale reaction in the strip is mediated mainly by mast cells, and possibly other cells, and is probably not due to a direct effect of the antigen-antibody reaction on the smooth muscle.5. The typical three-phase anaphylactic response (quick contraction, quick relaxation, slow contraction) of full-thickness ileum is discussed and compared with the predominantly two-phase response of the longitudinal muscle strip. No evidence was found for the release of a relaxation-factor. It is suggested that the initial fast phase may be due to mediators released from mast cells among the longitudinal muscle fibres, and the sustained contraction to a second wave of mediators reaching the longitudinal muscle from deeper layers of the ileum.

Topics: Amines; Anaphylaxis; Animals; Antigens; Dinitrophenols; Guinea Pigs; Histamine; Histamine Release; Humans; Ileum; In Vitro Techniques; Mast Cells; Muscle, Smooth; Myenteric Plexus; Ovalbumin; Serum Albumin; Tetrodotoxin

Topics: Anaphylaxis; Animals; Antigen-Antibody Reactions; Antigens; Guinea Pigs; Histamine; Histamine Release; Ileum; In Vitro Techniques; Mast Cells; Muscle, Smooth; Myenteric Plexus; Tetrodotoxin