title: Characterization of the human fMLP receptor in neutrophils and in Xenopus oocytes.
abstract: 1. N-formyl peptides (e.g. fMLP; N-formyl-L-methionyl-L-leucyl-phenylalanine) are potent mediators for inflammatory reactions. We report functional expression in Xenopus oocytes of human fMLP-R98 cDNA, without co-expression of the promiscuous G-protein subunit, Galpha-16. 2. Stimulation of voltage-clamped oocytes (-70 mV) with fMLP produced a dose-dependent biphasic inward current with fast and slow components. Analysis using GTP-gamma-S and cholera and pertussis toxins suggested these currents are mediated by an endogenous G-protein of the Gq family. 3. The fast current reversed at -25 mV and was blocked by SITS (4-acetamido-4'-isothiocyanatostilbene-2,2'-disulphonic acid), suggesting the current is carried by Cl(-). The slow current showed weak inward rectification, was Ca(2+)-dependent and blocked by Cd(2+), 4-AP (4-aminopyridine) and haloperidol, suggesting activation of a mixed population of cation channels. 4. Comparative experiments with human neutrophils using flow cytometric analysis showed that the proportion of neutrophils activated by fMLP was reduced in the presence of SITS, in the absence of external calcium and in the presence of Cd(2+), TEA (tetraethylammonium) and haloperidol but not 4-AP. In addition, the oxidative burst from activated neutrophils was reduced by SITS and by the absence of external calcium but not by Cd(2+), TEA, 4-AP or haloperidol. 5. We suggest that in human neutrophils activation by fMLP is dependent on store-operated calcium influx that appears to be regulated by Cl(-) channels and linked, in part, to non-selective cation channels.
authors.affiliation: Department of Anaesthesia, University of Regensburg, 93042 Regensburg, Germany. Welsh School of Pharmacy, Cardiff University, Cardiff CF10 3XF. sigrid.wittmann@klinik.uni-regensburg.de
authors.forename: Sigrid
authors.initials: S
authors.lastname: Wittmann
authors.forename: Dieter
authors.initials: D
authors.lastname: Fröhlich
authors.forename: Stephen
authors.initials: S
authors.lastname: Daniels
mesh_terms: D000328:Adult; D000818:Animals; D004305:Dose-Response Relationship, Drug; D005260:Female; D006801:Humans; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism*; D009865:Oocytes / Q000187:drug effects / Q000378:metabolism*; D044042:Receptors, Formyl Peptide; D011971:Receptors, Immunologic / Q000819:agonists / Q000096:biosynthesis / Q000737:chemistry*; D018000:Receptors, Peptide / Q000819:agonists / Q000096:biosynthesis / Q000737:chemistry*; D014982:Xenopus laevis
chemical_list: D044042:Receptors, Formyl Peptide; D011971:Receptors, Immunologic; D018000:Receptors, Peptide; D009240:N-Formylmethionine Leucyl-Phenylalanine