title: Activation of a voltage-dependent chloride current in human neutrophils by phorbol 12-myristate 13-acetate and formyl-methionyl-leucyl-phenylalanine. The role of protein kinase C.
abstract: Calcium-activated, voltage-independent Cl- currents have been demonstrated in human neutrophils (Krause, K.-H., and Welsh, M.J. (1990) J. Clin. Invest. 85, 491-498). The activation is mediated by calcium/calmodulin-dependent protein kinase and not by protein kinase C (PKC) (Schumann, M., Gardner, A.P., and Raffin, T.A. (1993) J. Biol. Chem. 268, 2134-2140). It is not known whether there are Ca(2+)-independent, voltage-dependent Cl- currents in these cells. Using K(+)-free solutions and patch-clamp recordings from human neutrophils, we separated the whole cell Cl- current. Base-line Cl- currents of unstimulated cells were small and displayed time and voltage independence; some showed voltage dependence. With a Ca(2+)-free pipette solution, bath-administered 1 microM phorbol 12-myristate 13-acetate (PMA) or 0.1 microM N-formyl-methionyl-leucyl-phenylalanine (fMLP) for 10 s induced augmented Cl- currents with voltage- and time-dependent outwardly rectifying properties. The threshold voltage of tail Cl- current activation was -69 mV. With a pipette solution containing 0.1 mM Ca2+, bath-administered 0.1 microM fMLP, 1 microM PMA, or 1 microM Ca2+ ionophore A23187 for 30 s induced augmented Cl- currents with voltage-independent properties. With intracellular application of 5 microM PKC inhibitor PKC(19-36), voltage-dependent Cl- currents were no longer activated by PMA or fMLP. Similar application of 5 microM PKC noninhibitory analog [Glu27]PKC(19-36) did not block PMA (or fMLP)-induced Cl- currents. These results indicate that, in addition to Ca(2+)-activated Cl- currents, human neutrophils have voltage-dependent Cl- currents which are regulated by PKC.
authors.affiliation: Division of Pulmonary and Critical Care Medicine, Stanford University School of Medicine, California 94305-5236.
authors.forename: M A
authors.initials: MA
authors.lastname: Schumann
authors.forename: T A
authors.initials: TA
authors.lastname: Raffin
mesh_terms: D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000097:blood; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D018118:Chloride Channels / Q000097:blood* / Q000187:drug effects / Q000502:physiology; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D006801:Humans; D066298:In Vitro Techniques; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology*; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D009842:Oligopeptides / Q000494:pharmacology; D010446:Peptide Fragments / Q000494:pharmacology; D011493:Protein Kinase C / Q000097:blood*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
chemical_list: D018118:Chloride Channels; D009842:Oligopeptides; D010446:Peptide Fragments; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; D000001:Calcimycin; D004533:Egtazic Acid; D009240:N-Formylmethionine Leucyl-Phenylalanine; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium