Signal processing times in neutrophil activation: dependence on ligand concentration and the relative phase of metabolic oscillations.
Intracellular NAD(P)H oscillations exhibited by polarized neutrophils display congruent with 20 s periods, which are halved to congruent with 10 s upon stimulation with chemotactic peptides such as FNLPNTL (N-formyl-nle-leu-phe-nle-tyr-lys). By monitoring this frequency change, we have measured accurately the time interval between stimulus and metabolic frequency changes. A microscope flow chamber was designed to allow rapid delivery of FNLPNTL to adherent cells. Using fluorescein as a marker, we found delivery to be complete and stable throughout the chamber within approximately 400 ms. Peptides were injected into the chamber at concentrations ranging from 10(-6) to 10(-9) M. Injections also varied with respect to the relative phase of a cell's NAD(P)H oscillations. The time interval between injection of 10(-6) M FNLPNTL and the acquisition of congruent with 10 s period metabolic oscillations was found to be 12.2+/-3.3 s when injections occurred at the NAD(P)H oscillation peak whereas the lag time was 22.5+/-4.8 s when coinciding with a trough. At 10(-8) M FNLPNTL, lag times were found to be 26.1+/-5.2 and 30.5+/-7.3 s for injections at NAD(P)H peaks and troughs, respectively. FNLPNTL at 10(-9) M had no effect on metabolic oscillations, consistent with previous studies. Our experiments show that the kinetics of transmembrane signal processing, in contrast to a simple transmembrane chemical reaction, can depend upon both ligand dose and its temporal relationship with intracellular metabolic oscillations.
Albrecht
Eric
E
Department of Ophthalmology and Visual Science, The University of Michigan School of Medicine, 1000 Wall Street, Ann Arbor, MI 48105, USA.
Kindzelskii
Andrei L
AL
Petty
Howard R
HR
D000595:Amino Acid Sequence; D006801:Humans; D008024:Ligands; D009249:NADP / Q000378:metabolism*; D018375:Neutrophil Activation / Q000187:drug effects*; D010455:Peptides / Q000737:chemistry / Q000494:pharmacology; D013050:Spectrometry, Fluorescence; D013997:Time Factors
D008024:Ligands; D010455:Peptides; D009249:NADP