title: Intracellular acidification induced by passive and active transport of ammonium ions in astrocytes.
abstract: We describe an unconventional response of intracellular pH to NH4Cl in mouse cerebral astrocytes. Rapid alkalinization reversed abruptly to be replaced by an intense sustained acidification in the continued presence of NH4Cl. We hypothesize that high-velocity NH4+ influx persisted after the distribution of ammonia attained steady state. From the initial rate of acidification elicited by 1 mM NH4Cl in bicarbonate-buffered solution, we estimate that NH4+ entered at a velocity of at least 31.5 nmol.min-1.mg protein-1. This rate increased with NH4Cl concentration, not saturating at up to 20 mM NH4Cl. Acidification was attenuated by raising or lowering extracellular K+ concentration. Ba2+ (50 microM) inhibited the acidification rate by 80.6%, suggesting inwardly rectifying K+ channels as the primary NH4+ entry pathway. Acidification was 10-fold slower in rat hippocampal astrocytes, consistent with the difference reported for K+ flux in vitro. The combination of Ba2+ and bumetanide prevented net acidification by 1 mM NH4Cl, identifying the Na(+)-K(+)-2Cl- cotransporter as a second NH4+ entry route. NH4+ entry via K+ transport pathways could impact "buffering" of ammonia by astrocytes and could initiate the elevation of extracellular K+ concentration and astrocyte swelling observed in acute hyperammonemia.
authors.affiliation: Department of Pharmacology and Experimental Therapeutics, University of Maryland School of Medicine, Baltimore 21202, USA.
authors.forename: T N
authors.initials: TN
authors.lastname: Nagaraja
authors.forename: N
authors.initials: N
authors.lastname: Brookes
mesh_terms: D000143:Acids / Q000378:metabolism*; D000643:Ammonium Chloride / Q000037:antagonists & inhibitors / Q000494:pharmacology; D000818:Animals; D001253:Astrocytes / Q000378:metabolism*; D001464:Barium / Q000494:pharmacology; D001692:Biological Transport / Q000502:physiology; D001693:Biological Transport, Active / Q000502:physiology; D002021:Buffers; D002034:Bumetanide / Q000494:pharmacology; D002478:Cells, Cultured; D005110:Extracellular Space / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007425:Intracellular Membranes / Q000378:metabolism*; D007477:Ions; D051379:Mice; D009994:Osmolar Concentration; D011188:Potassium / Q000037:antagonists & inhibitors / Q000378:metabolism; D000644:Quaternary Ammonium Compounds / Q000378:metabolism*; D051381:Rats
chemical_list: D000143:Acids; D002021:Buffers; D007477:Ions; D000644:Quaternary Ammonium Compounds; D000643:Ammonium Chloride; D002034:Bumetanide; D001464:Barium; D011188:Potassium