The Journal of Membrane Biology

, Volume 52, Issue 3, pp 257–272

Interaction of chemotactic factors with human polymorphonuclear leukocytes: Studies using a membrane potential-sensitive cyanine dye

Authors

  • Bruce E. Seligmann
    • Neurobiology and Experimental Hematology DepartmentsArmed Forces Radiobiology Research Institute
    • Department of ChemistryUniversity of Maryland
  • Elaine K. Gallin
    • Neurobiology and Experimental Hematology DepartmentsArmed Forces Radiobiology Research Institute
    • Department of ChemistryUniversity of Maryland
  • David L. Martin
    • Neurobiology and Experimental Hematology DepartmentsArmed Forces Radiobiology Research Institute
    • Department of ChemistryUniversity of Maryland
  • William Shain
    • Neurobiology and Experimental Hematology DepartmentsArmed Forces Radiobiology Research Institute
    • Department of ChemistryUniversity of Maryland
  • John I. Gallin
    • Neurobiology and Experimental Hematology DepartmentsArmed Forces Radiobiology Research Institute
    • Department of ChemistryUniversity of Maryland
Articles

DOI: 10.1007/BF01869194

Cite this article as:
Seligmann, B.E., Gallin, E.K., Martin, D.L. et al. J. Membrain Biol. (1980) 52: 257. doi:10.1007/BF01869194

Summary

Changes in the fluorescence intensity of the dye 3-3′ dipentyloxacarbocyanine were measured in suspensions of purified human peripheral blood polymorphonuclear leukocytes (PMNs) during exposure to the chemotactic factors N-formyl-methionylleucyl-phenylalanine (f-met-leu-phe) and partially purified C5a. Incubation of PMNs with dye resulted in a stable fluorescence reflecting the resting membrane potential of the cell. Exposure of PMNs to dye did not affect stimulated chemotaxis or secretion. The mechanism of cell-associated dye fluorescence involved solvent effects from partitioning of the dye between the aqueous incubation medium and the cell and not dye aggregation, Chemotactically active concentrations of f-met-leu-phe (5×10−9m or greater) produced a biphasic response characterized as a decrease followed by an increase in fluorescence. No fluorescence response was seen in lysed PMNs, and no response was elicited by an inhibitor of f-met-leu-phe binding (carbobenzoxy-phenylalanyl-methionine). The ability of several other synthetic peptides to elicit a fluorescence response corresponded to their effectiveness as chemotactic agents. Although the first component of the response suggested a depolarization, it was not influenced by variation in the external concentration of sodium, potassium, chloride, or calcium, and could not be characterized as a membrane potential change. The second component of the response, which was inhibited by both Mg2+ (10mm)-EGTA (10mm) and high external potassium, was compatible with a membrane hyperpolarization. The data indicate that chemotactic factors produce changes in dye fluorescence which can, at least in part, be attributed to a hyperpolarizing membrane potential change occurring across the plasma membrane.

Copyright information

© Springer-Verlag New York Inc 1980