Arachidonic Acid (AA) and Tetradecanoylphorbol Acetate (TPA) Exert Systemic Effects When Applied Topically in the Mouse

We studied the systemic actions of topically applied arachidonic acid (AA) and tetradecanoylphorbol acetate (TPA) in the mouse. AA or TPA-induced ear edema, increases in vascular permeability, eicosanoid levels, neutrophil and mononuclear influx were determined in both phlogogen-treated and contralateral vehicle-treated ear of each mouse and were compared with vehicle-treated ears from control mice. Edema and vascular permeability increases appeared only in AA- or TPA-applied ears. Moreover, in contralateral ears from AA-treated mice an increase in 6-keto-PGF₁ and LTB₄ was found. Only LTB₄ increased in the contralateral ear after TPA. Contralateral ears from AA- or TPA-treated mice also showed a significant increase in MPO levels. The increased levels of 6-keto-PGF₁ but not those of LTB₄ in contralateral ears were reduced by indomethacin applied simultaneously with the phlogogen. AA also increased plasma and serum levels of LTB₄, but not those of 6-keto-PGF₁. In contrast, TPA increased plasma and serum levels of 6-keto-PGF₁ and LTB₄. The results show that both AA and TPA applied topically exert systemic effects.     

Effect of Topically Applied Cyclosporin A on Arachidonic Acid (AA)- and Tetradecanoylphorbol Acetate (TPA)-Induced Dermal Inflammation in Mouse Ear

Topical cyclosporin A (CsA) was compared with dexamethasone, indomethacin and phenidone in edema, increases in vascular permeability, eicosanoids and cell-influx induced by arachidonic acid (AA) and tetradecanoylphorbol acetate (TPA) in mouse ears. CsA ED₅₀ on AA-edema (7.7 g/ear) was similar to dexamethasone and lower than indomethacin and phenidone. CsA ED₅₀ in TPA edema (21 g/ear) was higher than dexamethasone and lower than indomethacin or phenidone. All drugs equally reduce the AA-induced increase in vascular permeability, but CsA and dexamethasone had more activity on TPA. AA-increase in 6-keto-PGF₁ was reduced by dexamethasone, indomethacin and phenidone but not by CsA; only phenidone reduced LTB₄. TPA-increase in 6-keto-PGF₁ was reduced by CsA and indomethacin while CsA, dexamethasone and phenidone decreased LTB₄. CsA, indomethacin and phenidone, but not dexamethasone, suppressed AA-neutrophil influx. In TPA-ears all drugs produced similar reduction in neutrophil influx. CsA was shown to be a good topical anti-inflammatory drug.     

Increase in vascular permeability induced by leukotriene b4 and the role of polymorphonuclear leukocytes

Leukotriene B₄ (LTB₄), a metabolite of arachidonic acid, is known to be a potent chemotactic and chemokinetic substance. We have used the hamster cheek pouch microcirculation model to study the effect of LTB₄ on vascular permeability and the involvement of neutrophil granulocytes in this response. Intravascular fluorescein-labeled dextran (mol wt 150,000) was used as a tracer of macromolecular permeability. Topical application of LTB₄ (150 nM-5 M) to the hamster cheek pouch resulted in an immediate increase in adhering leukocytes in postcapillary venules and later larger venules. Leukocyte accumulation was reversible, but continued longer the higher the dose of LTB₄ used. Subsequently, a dose-dependent increase in vascular permeability was seen at postcapillary and larger venules, with a maximum 10–20 min after application; the maximum occurred later the higher the dose of LTB₄. Depletion of neutrophil granulocytes by pretreatment of the animals with antineutrophil serum obtained from immunized rabbits significantly decreased the permeability response to LTB₄, whereas the response to histamine was unaffected. These results suggest that neutrophil granulocytes play a role in LTB₄-mediated permeability increase. LTB₄ may be of importance both for the leukocyte accumulation and for the edema formation seen in inflammatory reactions.     

Leukotriene B4-induced permeability increase in postcapillary venules and its inhibition by three different antiinflammatory drugs

Leukotriene B₄ (LTB₄) is a potent chemotactic and chemokinetic substance that causes leukocyte accumulation and increases vascular permeability. We have used the hamster cheek pouch model to study the effect of some different antiinflammatory drugs on the LTB₄-induced microvascular permeability. Intravascular fluorescein-labeled dextran (mol wt 150,000) was used as a tracer of macromolecular leakage, which was measured as the number of venular leaky sites and as efflux of the vascular tracer. Topical application of LTB₄ (10^–8 M, 5 min) resulted in a reversible increase of the vascular permeability and could be repeated with 40-min intervals without any significant reduction in permeability response. The LTB₄-induced response could be inhibited by budesonide (a glucocorticoid) given locally, by iloprost (a prostacyclin analog) given intravenously and by a combined local and intravenous treatment with terbutaline (a ₂-receptor agonist) but not by local terbutaline treatment only.     

Effect of Topically Applied Cyclooxygenase-2-Selective Inhibitors on Arachidonic Acid- and Tetradecanoylphorbol Acetate-Induced Dermal Inflammation in the Mouse

The topical effects of cyclooxygenase-2 (COX-2)-selective inhibitors, flosulide (CGP 28238), L-745,337 and SC-57,666 were examined in AA- and TPA-induced ear dermal inflammation in the mouse. The doses that caused 50% inhibition in AA edema (ED₅₀) were 2.4, 0.45 and 0.35 mg/ear for flosulide, L-745,337 and SC-57,666, respectively. The respective ED_50s in TPA-edema were 1, 0.45 and 0.14. Indomethacin and zileuton showed higher activity than the COX-2-selective inhibitors in both models. Flosulide and L-745,337 inhibited the AA-induced increase in 6-keto-PGF₁, while SC-57,666 was inactive. 80% inhibition was seen with indomethacin while zileuton had no effect. COX-2 selective inhibitors and indomethacin had no effect on LTB₄ levels, while zileuton produced a 50% inhibition. The TPA-induced increase in 6-keto-PGF₁ was greatly inhibited by all COX-2 inhibitors while LTB₄was potentiated by both flosulide and L-745,337. Indomethacin inhibited 6-keto-PGF₁ and zileuton reduced 6-keto-PGF and strongly reduced LTB₄. The neutrophil influx induced by AA was lower than that of TPA. Myeloperoxidase (MPO) levels were lowered by flosulide and L-745,337 but not by SC-57,666. TPA-induced MPO increase was decreased by all COX-2 inhibitors. Indomethacin and zileuton had similar effect on AA and TPA-induced increase in MPO. The results indicate that COX-2-selective inhibitors showed lower topical anti-inflammatory activity than indomethacin or zileuton.     

The modulation of enhanced vascular permeability by prostaglandins through alterations in blood flow (hyperemia)

The enhanced vascular permeability induced by histamine or bradykinin in the skin of the guinea-pig and rabbit was significantly augmented by small amounts of prostaglandins of the E type. When injected alone these prostaglandins had little effect on vascular permeability. Furthermore, E type prostaglandins were found to be more potent at inducing hyperemia than either histamine or bradykinin. Prostaglandin F₂ did not enhance the vascular permeability induced by histamine or bradykinin nor did it produce hyperemia in the skin. In the rat, prostaglandins alone enhanced vascular permeability but they also increased the effect of histamine, serotonin and bradykinin. Using⁸⁵Sr-microspheres to measure blood flow a correlation was found between the degree of hyperemia produced by prostaglandins and the degree to which they augmented enhanced vascular permeability due to histamine, serotonin or bradykinin. Prostaglandins therefore can directly mimic the hyperemia of the inflammatory process and can also modulate the changes in vascular permeability caused by other mediators of inflammation.Supported by grants MA-5046 and MT-1251 of the Medical Research Council of Canada and the Ontario Heart Foundation.     

Role of corticosterone in modulation of eicosanoid biosynthesis and antiinflammatory activity by 5-lipoxygenase (5-LO) and cyclooxygenase (CO) inhibitors

The effectiveness of 5-lipoxygenase (LO) and dual LO/cyclooxygenase (CO) inhibitors when administered by the topical or oral routes was significantly decreased in corticosterone depleted (adrenalectomized, Adx) mice as compared to sham mice in the mouse arachidonic acid (AA) induced ear edema model. In contrast, rat carrageenan paw edema was inhibited similarly in sham and Adx animals by 5-LO and dual 5-LO/CO inhibitors. Supplementation of cortisol levels (100 g/dl) in human whole blood for 2 hr increased the observed inhibition of LTB₄ biosynthesis by A-64077, WY-50,295 tromethamine and naproxen while having no effect on thromboxane B₂ (TXB₂) biosynthesis. Thus, corticosteroids may have a permissive effect, by modulating 5-LO inhibitor, effects on mouse AA induced ear edema and human blood leukocytes.     

Aggregation of polymorphonuclear leucocytes (PMNs) by leukotriene B4: Effects of cyclooxygenase products and metabolic inhibitors

Leukotriene B₄ isomer III (LTB₄) stimulates the aggregation of rat PMNsin vitro. The effects of deoxyglucose, iodoacetate, dinitrophenol, cyclohexamide, colchicine, prostaglandins and thromboxane B₂ (TXB₂) on aggregation were examined. The results demonstrate that, first, aggregation is an active process, the energy being supplied by glycolysis and not mitochondrial respiration. The response can be elicited in the absence of extracellular glucose. Synthesis of protein does not occur during aggregation. An intact microtubular system is required for the expression of a full aggregation response. Prostaglandins E₁, E₂, F₂ and TXB₂, products of the cyclooxygenase pathway of arachidonic acid metabolism, partially inhibit LTB₄-induced aggregation by a mechanism which has not yet been elucidated. The prostaglandins and TXB₂ themselves do not promote aggregation.     

Priming effects of leukotriene B4 on endothelial cell injury induced by tpa-activated leukocytes

Among arachidonic acid metabolites, leukotriene B₄ (LTB₄) plays an important role in inflammation, such as in the activation, adhesion, chemotaxis, and invasion of leukocytes. In this paper, we examined the effect of LTB₄ on endothelial cell injury induced by polymorphonuclear leukocytes (PMNLs).⁵¹Crrelease, a marker of cellular injury, was elicited from prelabeled endothelial cells when the cells were cocultured with PMNLs activated by phorbol ester (TPA, 12-0-tetradecanoyl-phorbol-13-acetate). Under this condition, pretreatment of PMNLs with LTB₄ enhanced their injury in a dose-dependent manner (0.2–2 M). However, LTB₄ alone at any dose could not induce any cellular injury. We also determined the amount of active oxygen species produced by PMNLs in response to TPA. The intensity of luminoldependent chemiluminescence, a marker of active oxygen production, in PMNLs was also increased by pretreatment with 1 M LTB₄. These data suggest that LTB₄ enhances endothelial cell injury by the priming effect on active oxygen production in activated PMNLs.     

Differential effects of monohetes (monohydroxyeicosatetraenoic acids) on arachidonic acid metabolism in glycogen-elicited rat polymorphonuclear leukocytes

We investigated the effects of monohydroxyeicosatetraenoic acids (monoHETEs) on lipoxygenase- and cyclooxygenase-catalyzed reactions in glycogen-elicited rat PMNs challenged with A23187 and exogenous [¹⁴C]arachidonic acid. A23187 (10M) stimulated a 10-, 4-, 1.7- and 1.8-fold increase in the synthesis of radiolabeled 5-HETE, LTB₄, TxB₂, and PGE₂ by rat PMNs. Addition of 5-HETE, 5 lactone-HETE, 12-HETE, and 15-HETE led to a dose-related reduction in [¹⁴C]5-HETE and [¹⁴C]LTB₄ synthesis by these cells. These monoHETEs also inhibited [¹⁴C]TxB₂ synthesis, but only 5-HETE and 5 lactone-HETE inhibited the synthesis of [¹⁴C]PGE₂. Both 12-HETE and 15-HETE failed to reduce the formation of [¹⁴C]PGE₂. These results suggest that monoHETEs differ significantly in their effects on arachidonic acid metabolism in rat PMNs and may play a role in modulating the synthesis of both lipoxygenase and cyclooxygenase products.     

Comparison of in vitro effects of flunixin and tolfenamic acid on human leukocyte and platelet functions

A study was made to compare the effects of two nonsteroidal antiinflammatory drugs (NSAIDs), flunixin and tolfenamic acid, on the leukotriene B₄ (LTB₄) production and migration of human polymorphonuclear leukocytes (PMNs) as well as on platelet aggregation and thromboxane B₂ (TxB₂) production during blood clotting. Tolfenamic acid inhibited LTB₄ production in PMNs as well as FMLP- and LTB₄-induced PMN migration (IC₅₀ values 23 ± 3, 39 ± 11, and 68 ± 13 M, respectively), whereas flunixin inhibited these cell functions only with the highest concentration tested (100 M). On the other hand, flunixin was clearly a more potent inhibitor of TxB₂ production and adrenaline-induced platelet aggregation than tolfenamic acid, the IC₅₀ values in TxB₂ production being 0.28 ± 0.02 M and 2.6 ±0.3 M for flunixin and tolfenamic acid, respectively. We suggest that inhibition of PMN functions may be an additional mechanism in the antiinflammatory action of tolfenamic acid. At least in human PMNs and platelets, flunixin seems to be only an inhibitor of cyclooxygenase.     

Inhibition of leukotriene B4(LTB4) by recombinant interleukin-1 receptor antagonist (IL-1RA) on human monocytes

Macrophages are a primary source of interleukin-1 (IL-1), a glycoprotein which plays an important and essential role in the immune response and inflammation. Cytokines stimulate many different cells to produce increasing amounts of arachidonic acid metabolites such as prostaglandins and leukotrienes. Recently, interleukin-1 receptor antagonist (IL-1ra), a natural inhibitor of IL-1 released by macrophages, has been reported to inhibit PGE₂. In accordance with these data our results show that the pretreatment, for 60 min, of purified human peripheral monocytes with IL-1ra at different concentrations (0.25–250 ng/ml) inhibits, in a dose-dependent manner, the generation of LTB₄ released after 10 min treatment with calcium ionophore A23187 (5 M). The inhibition of LTB₄ synthesis by hrIL-1ra suggests the possibility that this new glycoprotein plays a modulatory role in immunity and inflammation.     

Effect of 5-lipoxygenase inhibitors onin situ LTB4 biosynthesis following calcium ionophore stimulation in the rat pleural cavity

The intrapleural injection of carrageenan in the rat induces exudate formation, cellular influx and leukotriene generation in the pleural cavity. We have demonstrated that the inflammatory response (exudate volume, and LTB₄ levels) is increased,in situ by the intrapleural administration of calcium ionophore A 23187 (100 nmol/rat) at 4, 16, 24, 48, and 72 h after the injection of carrageenan and that the A 23187-induced increase is dose-dependent. The oral administration of A-64077 and MK-886, two 5-lipoxygenase inhibitors (5-LOIs), at 10 mg/kg causes marked decreases in LTB₄ release at the above-mentioned time intervals. However, A 23187-induced augmented exudate formation is not affected by the treatment with 5-LOIs. The results suggest that the use of 5-LOIs to inhibit LTB₄ biosynthesis may be beneficial in various LTB₄-dependent pathological conditions.     

Ebselen is a specific inhibitor of LTB4-mediated migration of human neutrophils

Ebselen is a seleno-organic anti-inflammatory compound with glutathione peroxidase-like activity that has the unique characteristic of mediating the isomerization of 5-HETE and LTB₄ to their biologically inactive trans isomers, both directly in fluid phase and indirectly through metabolic pathways in stimulated peripheral blood leukocytes. LTB₄ is an inflammatory mediator with potent chemotactic activity for neutrophilic leukocytes. We studied the effects of ebselen on the chemotactic and chemokinetic responses with human-blood-derived neutrophils. With the use of 120-m-thick 5-m-pore durapore filters and low BSA concentrations (0.05%) in the chemotaxis buffers, ebselen was evaluated for its effect on both chemotactic and chemokinetic responses to LTB₄, C5a, and fMLP. Ebselen at 3–20 M concentrations inhibited both chemotactic and chemokinetic responses to optimal concentrations of LTB₄ without altering chemotactic responses to C5a or fMLP. Likewise, ebselen at 20 M specifically inhibited LTB₄-stimulated transendothelial migration of neutrophils, while not altering responses to C5a nor fMLP.     

A comparison of the anti-inflammatory activity of selective 5-lipoxygenase inhibitors with dexamethasone and colchicine in a model of zymosan induced inflammation in the rat knee joint and peritoneal cavity

Intraperitoneal and intra-articular (knee joint) injection of zymosan in the rat caused two phases of increased vascular permeability, a rapid increase (0.25–0.5 h) and a secondary increase (2–3 h) which was temporally associated with the onset of leukocyte infiltration. Intraperitoneal injection of zymosan led to a single peak of eicosanoid production (LTB₄, C₄, D₄, E₄ and 6-oxo-PGF₁ ) which was maximal at 0.125–0.25 h. Intra-articular injection led to an initial peak of LTB₄ production (maximal at 0.25 h) and a secondary peak of LTB₄ and PGE₂ production (maximal at 3 h). Oral administration of the 5-lipoxygenase (5-LO_ inhibitors phenidone, BW A4C (N-hydroxy-N-[3-(3-phenoxyphenyl)-2-propenyl] acetamide), A63162 (N-hydroxy-N-[1-(4-(phenylmethoxy) phenyl)ethyl] acetamide and ICI 207 968 (2-[3-pyridylmethyl]-indazolinone inhibited LTB₄ production in A23187 stimulation bloodex vivo. The glucocorticosteroid dexamethasone had no effect in this model. The initial phase of increased vascular permeability in the peritoneal cavity and LTB₄ production was dose dependently inhibited by the 5-LO inhibitors phenidone, BW A4C, A63162, and ICI 207 968 but not by dexamethasone or colchicine. The initial phase of increased permeability in the joint was unaffected by phenidone, BW A4C, dexamethasone or colchicine. However the latter two drugs inhibited the later phase of increased permeability and leukocyte infiltration in the joint and peritoneal cavity. These results demonstrate that zymosan induces eicosanoid productionin vivo but the relative importance of these mediators varies depending on the inflammatory site. Dexamethasone had a similar profile of anti-inflammatory activity to colchicine suggesting that inhibition of cell infiltration is the major mechanism of its therapeutic effect. No evidence for dexamethasone induced inhibition of eicosanoid production could be found suggesting that inhibition of the production or activity of other mediators is more important.     

Interactions between synthesis of platelet-activating factor and leukotriene B4 in isolated human polymorphonuclear leukocytes

The aim of the present work was to study interactions between the synthesis of platelet-activating factor (PAF) and leukotriene B₄ (LTB₄) in human polymorphonuclear leukocytes (PMNs) in vitro. PAF, at nanomolar concentrations, stimulated calcium ionophore A23187-activated PMNs to release LTB₄ and 5-hydroxyeicosatetraenoic acid (5-HETE). This seems to be a receptor-mediated process as it was blocked by a PAF receptor antagonist WEB 2086 (IC₅₀ 6.6±3.9M). Moreover, LTB₄ stimulated the formation of PAF in activated PMNs. WEB 2086 did not, however, alter PMN migration towards either LTB₄ or the chemotactic peptide FMLP. This suggests that the enhancement of PAF synthesis in response to LTB₄ is a concomitant event rather than a mediating process in LTB₄-induced chemotactic movement of PMNs. These effects are implicated in the complex network of interactions between inflammatory mediators that results accumulation and activation of PMNs in the exacerbation of inflammatory processes.     

Pharmacological profile of a novel,orally active leukotriene B4 antagonist,SM-15178

SM-15178, a new hydroxyacetophenone derivative, was evaluated to determine its antiinflammatory activity and antagonistic activity against leukotriene B₄ (LTB₄). SM-15178 inhibited [³H]LTB₄ binding to its receptors on human neutrophils (IC₅₀=0.30M). It inhibited LTB₄-induced chemotaxis of human neutrophils (IC₅₀ =0.72M) with little inhibitory effect against C5a or FMLP-induced chemotaxis at concentrations up to 30M. The compound alone did not cause human neutrophil chemotaxis at concentrations up to 10M. LTB₄-induced chemotaxis of mouse and rat neutrophils and guinea pig eosinophils was also inhibited by the compound, with IC₅₀ values of 0.55, 0.52, and 0.58 M, respectively. In an in vivo study, SM-15178, given orally, significantly prevented LTB₄-induced transient leukopenia. It also suppressed LTB₄-induced bronchoconstriction in the guinea pig almost completely when given orally at a dose of 40 mg/kg. Furthermore, orally given SM-15178 suppressed arachidonic acid-induced neutrophil infiltration in mouse ears and Arthus reaction-induced paw edema in the mouse in a dose-dependent manner. These results suggest that SM-15178 is a selective and orally active LTB₄ antagonist and that it might be effective for the treatment of some types of inflammatory diseases.     

Antiinflammatory effects of second-generation leukotriene B4 receptor antagonist,SC-53228: Impact upon Leukotriene B4- and 12(R)-HETE- mediated events

Leukotriene B₄ (LTB₄) and 12(R)-hydroxyeicosatetraenoic acid [12(R)-HETE] are proinflammatory products of arachidonic acid metabolism that have been implicated as mediators in a number of inflammatory diseases. When injected intradermally into the guinea pig, LTB₄ and 12(R)-HETE elicit a dose-dependent migration (chemotaxis) of neutrophils (PMNs) into the injection sites as assessed by the presence of a neutrophil marker enzyme myeloperoxidase. SC-41930 {7-[3-(4-acetyl-3-methoxy-2-propylphenoxy)propoxyl]-3,4-dihydro-8-propyl-2H-1-benzopyran-2-carboxylic acid}, a first-generation LTB₄ receptor antagonist, inhibited the chemotactic actions of LTB₄ when given orally with an ED₅₀ value of 1.7 mg/kg. The second-generation LTB₄ receptor antagonist, SC-53228 [(+)-(S)-7-(3-{2-(cyclopropylmethyl)-3-methoxy-4-[(methylamino)carbonyl]phenoxy} propoxy)-3,4-dihydro-8-propyl-2H-1-benzopyran-2-propanoic acid], inhibited LTB₄-induced chemotaxis when given intragastrically with an ED₅₀ value of 0.07 mg/kg. Furthermore, SC-53228 inhibited 12(R)-HETE-induced granulocyte chemotaxis with an oral ED₅₀ value of 5.8 mg/kg. When dosed orally over a range of 0.03–100 mg/kg, SC-53228 gaveC _max plasma concentrations of 0.015–41.1g/ml. SC-53228 inhibited LTB₄-primed membrane depolarization of human neutrophils with an IC₅₀ value of 34 nM. As a potent LTB₄ receptor antagonist, SC-53228 may well have application in the medical management of disease states such as asthma, rheumatoid arthritis, inflammatory bowel disease, contact dermatitis, and psoriasis, in which LTB₄ and/or 12(R)-HETE are implicated as inflammatory mediators.     

The Proinflammatory Cytokines and Arachidonic Acid Metabolites in Human Overnight Tears: Homeostatic Mechanisms

The tear film plays an important role in the defense of the external ocular surface. During sleep a number of changes take place, including increased production and release of various inflammatory mediators. We have studied the hypothesis that closed-eye tears contain proinflammatory cytokines and lipid inflammatory mediators, which serve to recruit polymorphonuclear leukocytes (PMNs) and regulate the function of PMNs and IgA during sleep. We investigated interleukin-1, interleukin-6, interleukin-8, monocyte chemotactic protein 1, granulocyte–macrophage colony stimulating factor (GM-CSF), leukotriene B₄ (LTB₄), and platelet activating factor (PAF) in open and closed-eye tears of normal healthy subjects. Results showed that IL-6, IL-8, GM-CSF, LTB₄, and PAF were present in high levels in closed-eye tears compared to open-eye tears. Closed-eye tears were able to recruit neutrophils, with maximal recruitment after 8 hr of sleep, suggesting that chemokine IL-8 and the lipid chemoattractant LTB₄ were active. Flow cytometric analysis revealed that incubation of neutrophils with closed-eye tears up-regulated the surface expression of IgA receptor, indicating that the GM-CSF in tears was functionally active. Up-regulation of cytokines and the lipid inflammatory mediator LTB₄ during eye closure are noteworthy, as each of these cytokines has an established role in initiation and amplification of the inflammatory response. IL-8 and LTB₄ may act as potent chemoattractants and activators for PMNs, whereas IL-6 and GM-CSF potentiate the secretion and function of IgA and enhance neutrophil responsiveness to proinflammatory agonists.     

In vitro effects of leukotriene B4 (LTB4) on canine PMN effector function(s)

The canine has become an accepted research model for the examination of a number of human clinical conditions. Despite it's status as a research model, little is known regarding the peripheral effects of inflammatory mediator substances. Products of arachidonic acid metabolism (leukotrienes) are reported capable of altering leukocyte functions. Because of the emerging importance of the canine research model and leukotrienes we examined the effects of leukotriene B₄ (LTB₄) on severalin vitro functions of isolated canine peripheral polymorphonuclear leukocytes (PMN). Changes in forward angle light scatter properties of the cells were used as one measure of PMN activation. Other functional changes examined following LTB₄ pretreatment included chemotactic capability, the electrophysiological state of the cell plasma membrane, and the metabolic oxidative response (i. e. H₂O₂ production). Random cellular movement of PMNs increased by 120% and 72% following preincubation with 10^?7 and 10^?9 M LTB₄, respectively. LTB₄ between 10^?7 and 10^?13 M did not significantly alter cellular resting membrane potential. Between 10^?7 and 10^?9 M LTB₄ elicited significant levels of cellular H₂O₂ production. Although significant, H₂O₂ production was <40% that induced by phorbol myristate acetate (PMA). In numerous respects, caninein vitro PMN responses parallel previous reports of human cell function(s) in the presence of inflammatory mediators and may represent an attractive alternative for investigation of PMN dysfunctions.