Intracellular Ca2+ concentration and H2O2 production in mouse peritoneal macrophages are stimulated by platelet activating factor.
The intracellular calcium concentration ([Ca2+]i) in mouse peritoneal macrophages cultured on a coverglass was measured in the superfusion system using fura-2 as a fluorescent calcium probe and platelet activating factor (PAF) as a stimulant. In the presence of extracellular Ca2+, 10(-7) M PAF sharply increased [Ca2+]i from a basal level of 90 nM to 340 nM. Thereafter, the [Ca2+]i level gradually decreased in two phases, in an initial rapid phase and a subsequent slow phase of decrease. The calcium response was dependent on the PAF concentration. In the absence of extracellular Ca2+, a single sharp peak was observed, suggesting two different modes of Ca2+ movement, one from intracellular stores and the other from the extracellular medium. A simple, sensitive fluorometric assay system was developed for measuring H2O2 in the superfusate of macrophages after stimulation by use of immobilized peroxidase and 3-(p-hydroxyphenyl)propionic acid as a fluorogenic substrate. With this system, as little as 2 pmol of H2O2 could be measured. PAF (1 microM) increased H2O2 production in peritoneal macrophages in the presence of extracellular Ca2+, but H2O2 production was not observed in the absence of extracellular Ca2+.
Sasaki
M
M
Department of Pharmacology I, Tohoku University School of Medicine, Miyagi, Japan.
Maeyama
K
K
Watanabe
T
T
D000818:Animals; D002118:Calcium / Q000378:metabolism* / Q000494:pharmacology; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004533:Egtazic Acid / Q000494:pharmacology; D006861:Hydrogen Peroxide / Q000378:metabolism*; D007700:Kinetics; D008264:Macrophages / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D051379:Mice; D008815:Mice, Inbred Strains; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D010972:Platelet Activating Factor / Q000494:pharmacology*; D013050:Spectrometry, Fluorescence / Q000295:instrumentation / Q000379:methods
D010972:Platelet Activating Factor; D004533:Egtazic Acid; D009240:N-Formylmethionine Leucyl-Phenylalanine; D006861:Hydrogen Peroxide; D002118:Calcium