Phorbol ester-stimulated human neutrophil membrane depolarization is dependent on Ca2(+)-regulated Cl- efflux.
The ionic basis of phorbol 12-myristate 13-acetate (PMA)-stimulated membrane depolarization in the human neutrophil has not previously been established. Alterations in cation permeability are probably not directly responsible for the depolarization response, since the rate or Rb+ influx or efflux is unchanged upon PMA stimulation, and although Na+ fluxes are increased, depolarization is not changed by either the addition of ouabain or reduction of extracellular Na+ from 140 to 0 meq. Furthermore, the enhanced Na+ influx in stimulated cells is blocked by amiloride at 10(-3) M, but not by 10(-5) M, suggesting Na+ influx proceeds through the electroneutral Na(+)-H+ antiporter and is therefore not responsible for depolarization. Upon stimulation, Cl- content of PMA-stimulated neutrophils decreases without change in Na+ or K+ content, as determined by electron probe analysis. In addition, acute reduction in extracellular Cl- enhances the rate and extent of depolarization induced by PMA. This change in intracellular Cl- and effect of reduction in extracellular Cl- concentration on depolarization can best be accounted for by an enhanced efflux via an electrogenic mechanism. Thus enhanced conductive Cl- efflux can account for the observed depolarization. That Ca2+ regulates depolarization is evidenced by the dependence of depolarization on external Ca2+ (Cao2+). Depolarization is absent in Ca2(+)-depleted cells [internal Ca2+ (Cai2+) less than 15 nM] and is restored with titration of extracellular Ca2+, exhibiting a 50% effective dose (ED50) of 100 mM. Thus PMA-initiated depolarization is regulated by Ca2+, either from intra- or extracellular sources, but the Ca2(+)-dependent activity responsible for control of Cl- efflux is as yet uncharacterized.
Myers
J B
JB
William B. Castle Hematology Research Laboratory, Boston City Hospital, Massachusetts.
Cantiello
H F
HF
Schwartz
J H
JH
Tauber
A I
AI
D002118:Calcium / Q000097:blood* / Q000494:pharmacology; D002462:Cell Membrane / Q000187:drug effects / Q000502:physiology; D002712:Chlorides / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008564:Membrane Potentials / Q000187:drug effects; D009504:Neutrophils / Q000187:drug effects / Q000502:physiology*; D011188:Potassium / Q000494:pharmacology; D012413:Rubidium / Q000097:blood; D012964:Sodium / Q000097:blood; D013481:Superoxides / Q000097:blood; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014634:Valinomycin / Q000494:pharmacology
D002712:Chlorides; D013481:Superoxides; D014634:Valinomycin; D012964:Sodium; D012413:Rubidium; D013755:Tetradecanoylphorbol Acetate; D011188:Potassium; D002118:Calcium