Cytoplasmic pH regulation in normal and abnormal neutrophils. Role of superoxide generation and Na+/H+ exchange.
The cytoplasmic pH of human neutrophils was determined fluorometrically using carboxylated fluorescein derivatives. When normal neutrophils were activated by the phorbol ester 12-O-tetradecanoylphorbol 13-acetate (TPA) in Na+-containing medium, the cytoplasmic pH initially decreased but then returned to near normal values. In Na+-free media or in Na+ medium containing amiloride, TPA induced a marked monophasic intracellular acidification. The cytoplasmic acidification is associated with net H+ equivalent efflux, suggesting metabolic acid generation. The metabolic pathways responsible for the acidification were investigated by comparing normal to chronic granulomatous disease neutrophils. These cells are unable to oxidize NADPH and generate superoxide. When treated with TPA in Na+-free or amiloride-containing media, chronic granulomatous disease cells did not display a cytoplasmic acidification. This suggests that in normal cells NADPH oxidation and/or the accompanying activation of the hexose monophosphate shunt are linked to the acidification. Unlike normal neutrophils, chronic granulomatous disease cells treated with TPA in Na+-containing medium displayed a significant cytoplasmic alkalinization. The alkalinization was Na+-dependent and amiloride-sensitive, indicating activation of Na+/H+ exchange. Thus, the Na+/H+ antiport, which can be indirectly stimulated by the metabolic cytoplasmic acidification, is also directly activated by the phorbol ester.
D000584:Amiloride / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism*; D003593:Cytoplasm; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008536:Meglumine / Q000494:pharmacology; D009249:NADP / Q000378:metabolism; D009504:Neutrophils / Q000166:cytology* / Q000187:drug effects; D009550:Nigericin / Q000494:pharmacology; D010427:Pentose Phosphate Pathway; D017923:Sodium-Hydrogen Exchangers; D013481:Superoxides / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology
D002352:Carrier Proteins; D017923:Sodium-Hydrogen Exchangers; D013481:Superoxides; D009249:NADP; D008536:Meglumine; D000584:Amiloride; D013755:Tetradecanoylphorbol Acetate; D009550:Nigericin