NH4+ metabolism and the intracellular pH in isolated perfused rat liver.
The effects of NH4+ on the intracellular pH (pHi) and on the ATP content in isolated perfused rat liver were studied by 31P n.m.r. spectroscopy. In the initial phase of perfusion an average pHi of 7.29 +/- 0.04 was estimated. The presence of low (0.5 mmol/l) and high (10 mmol/l) doses of NH4Cl induced significant intracellular acidification by -0.06 +/- 0.03 and -0.11 +/- 0.03 pH unit respectively. This effect was in contrast with the transient intracellular alkalinization observed in preliminary studies on isolated hepatocytes, which was caused by a passive entry of NH3 by non-ionic diffusion and subsequent conversion into NH4+. During application of 0.5 mmol/l NH4Cl the liver released 0.54 +/- 0.06 mumol of urea/min per g into the perfusate. When the intracellular availability of HCO3- was decreased by acetazolamide (0.5 mmol/l) or by removal of HCO3- from the perfusion medium, the decrease in pHi by NH4Cl application was significantly lower than under control conditions. Furthermore, synthesis of urea was significantly inhibited by the decrease in intracellular HCO3-. Under these conditions, 10 mmol/l NH4Cl caused the transient alkalinization that was expected because of the passive uptake of uncharged NH3. Therefore, it is concluded that the intracellular acidification induced by NH4Cl is caused by the continuous utilization of intracellular HCO3- via the synthesis of urea. This metabolic effect on pHi dominates the effects of passive NH3 entry. The rate of urea formation depends on continuous efflux of H+, which is strictly limiting the degree of intracellular acidification within a small range. If the extrusion of H+ by the Na+/H+ exchanger was inhibited by amiloride (0.5 mmol/l) during the NH4Cl application, the decrease in pHi was amplified and the formation of urea was significantly inhibited. The application of NH4Cl at 0.5 or 10 mmol/l decreased the ATP content by 11% or 22% respectively.
Max-Planck-Institut für Systemphysiologie, Dortmund, Germany.
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Zange
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Gronczewski
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Jans
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D000086:Acetazolamide / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000096:biosynthesis; D000584:Amiloride / Q000494:pharmacology; D000643:Ammonium Chloride / Q000378:metabolism*; D000818:Animals; D002255:Carbonic Acid / Q000378:metabolism; D002352:Carrier Proteins / Q000187:drug effects / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D008297:Male; D010477:Perfusion; D051381:Rats; D017208:Rats, Wistar; D017923:Sodium-Hydrogen Exchangers; D014508:Urea / Q000378:metabolism
D002352:Carrier Proteins; D017923:Sodium-Hydrogen Exchangers; D000643:Ammonium Chloride; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D002255:Carbonic Acid; D000584:Amiloride; D000255:Adenosine Triphosphate; D014508:Urea; D000086:Acetazolamide; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid