Platelet activating factor-induced increase in cytosolic calcium and transmembrane current in human macrophages.
Platelet-activating factor (PAF) is synthesized and secreted by macrophages in responding to inflammatory stimuli. When exogenously applied to human monocyte derived macrophages (HMDMs), PAF induces a rapid rise in cytosolic free calcium (Cai) believed to be an early triggering event in macrophage activation. We investigated PAF-induced Ca2+ signaling in HMDMs using the calcium indicator Fura-2, combining single cell ratio fluorimetry and digital video imaging with whole-cell recording techniques. Application of PAF (20 ng/ml) to adherent macrophages induced transient increases in Cai that were biphasic, consisting of an initial phase that could be observed in Ca(2+)-free solutions and a second phase that was critically dependent upon Ca2+ entry. When Mn2+ was applied to cells in the presence and absence of Ca2+, PAF increased the rate of Mn2+ entry rate only when Ca2+ was absent. PAF increased the rate of Ba2+ entry even when measured in the presence of external Ca2+. Ca2+ entry was reversibly inhibited in the presence of external La3+ (1 mM). Data obtained from simultaneous voltage-clamp/microfluorimetry experiments demonstrated the activation of a nonselective cation current which closely paralleled the rising phase of the Cai transient. We investigated whether the non-selective cation conductance provided for the bulk of the agonist-induced Ca2+ influx. Changes in Cai following removal of extracellular Ca2+ (Cao) during the agonist-induced Cai response were not associated with changes in whole-cell current. The inability to detect whole-cell current changes correlated with a decrease in Cao suggests that the bulk of the Ca2+ influx was not through the nonselective conductance and either does not occur through a conductance pathway or occurs via a parallel pathway consisting of channels which are both low conductance and highly Ca2+ selective.
Katnik
C
C
University of Chicago, Dept. of Neurology, Illinois 60637.
Nelson
D J
DJ
D001464:Barium / Q000493:pharmacokinetics / Q000494:pharmacology; D002118:Calcium / Q000032:analysis* / Q000378:metabolism / Q000493:pharmacokinetics*; D015220:Calcium Channels / Q000502:physiology*; D002352:Carrier Proteins / Q000502:physiology; D002462:Cell Membrane / Q000502:physiology; D002463:Cell Membrane Permeability; D002478:Cells, Cultured; D003600:Cytosol / Q000737:chemistry* / Q000378:metabolism; D016257:Fura-2; D006801:Humans; D007811:Lanthanum / Q000493:pharmacokinetics; D008264:Macrophages / Q000187:drug effects* / Q000378:metabolism; D008345:Manganese / Q000493:pharmacokinetics / Q000494:pharmacology; D008536:Meglumine / Q000494:pharmacology; D008564:Membrane Potentials; D010972:Platelet Activating Factor / Q000494:pharmacology*; D015222:Sodium Channels / Q000502:physiology; D019831:Sodium-Calcium Exchanger
D015220:Calcium Channels; D002352:Carrier Proteins; D010972:Platelet Activating Factor; D015222:Sodium Channels; D019831:Sodium-Calcium Exchanger; D001464:Barium; D008345:Manganese; D008536:Meglumine; D007811:Lanthanum; D002118:Calcium; D016257:Fura-2