High concentrations of histamine stimulate equine polymorphonuclear neutrophils to produce reactive oxygen species

Objective and Design: Because high concentrations of histamine are locally released in inflammation, we investigated the effects of supraphysiological doses of histamine on the production of reactive oxygen species (ROS) by n eutrophils. Materials and Methods: Isolated equine neutrophils were activated by 10(-4) to 5 x 10(-3) M histamine. The production of ROS and free radicals was est imated by luminol-enhanced chemiluminescence (CL) and electron spin resonan ce (ESR) with spin trapping technique. In this model of histamine-stimulate d neutrophils, we tested the antagonists of H1 and H2 histamine receptors, the role of Ca2+ and Mg2+, the role of staurosporine and pertussis toxin (i nhibitors of protein kinase C and proteins G) and the effects of superoxide dismutase, catalase, hydroxyl radical scavengers (phenylalanine and mannit ol) and NG-monomethyl-L-arginine (L-NMMA), inhibitor of NO-synthase. Results: Histamine (from 10(-5) to 10(-3) M) stimulated neutrophils to prod uce CL and ESR signals characterized by spin adducts of superoxide anion an d/or hydroxyl radicals. The CL response was inhibited by 10(-4) and 10(-3) M H1 receptor antagonists (promethazine, pyrilamine, and diphenhydramine), by Ca2+ and Mg2+ depletion and by 10 nmoles staurosporine. CL was partially inhibited by pertussis toxin (4 pgi mt). The ESR signals were practically suppressed by pyrilamine (an H1 receptor antagonist) and superoxide dismuta se, and partially inhibited by catalase, hydroxyl radical scavengers and L- NMMA (respectively 59, +/- 30% and 68% inhibition). Conclusions: High concentrations of histamine stimulated the neutrophils to product ROS and free radicals via H1 receptors and the NADPH-oxidase pathw ay.