K+/NH4+ antiporter: a unique ammonium carrying transporter in the kidney inner medulla.
The mechanism of NH4+ transport in inner medulla is not known. The purpose of these experiments was to study the process that is involved in ammonium (NH4+) transport in cultured inner medullary collecting duct (mIMCD-3) cells. Cells grown on coverslips were exposed to NH4+ and monitored for pHi changes by the use of the pH-sensitive dye BCECF. The rate of cell acidification following the initial cell alkalinization was measured as an index of NH4+ transport. The rate of NH4+ transport was the same in the presence or absence of sodium in the media (0.052 +/- 0.003 vs 0.048 +/- 0.004 pH/min. P > 0.05), indicating that NH4+ entry into the cells was independent of sodium. The presence of ouabain, bumetanide, amiloride, barium, or 4,4'-di-isothiocyanostilbene-2-2'-disulfonic acid (DIDS) did not block the NH4(+)-induced cell acidification, indicating lack of involvement of Na+:K(+)-ATPase, Na+:K+:2Cl- transport, Na+:H+ exchange, K+ channel, or Cl-/base exchange, respectively, in NH4+ transport. The NH4(+)-induced cell acidification was significantly inhibited in the presence of high external [K+] as compared to low external [K+] (0.018 +/- 0.001 vs. 0.049 +/- 0.003 pH/min for 140 mM K+ vs. 1.8 mM K+ in the media, respectively, P < 0.001). Inducing K+ efflux by imposing an outward K+ gradient caused intracellular acidification by approximately 0.3 pH unit in the presence but not the absence of NH4+. This K+ efflux-induced NH4+ entry increased by extracellular NH4+ in a saturable manner with a Km of approximately 5 mM, blocked by increasing extracellular K+ and was not inhibited by barium. The K+ efflux-coupled NH4+ entry was electroneutral as monitored by the use of cell membrane potential probe 3,3'-dipropylthiadicarbocyanine. These results are consistent with the exchange of internal K+ with external NH4+ in a 1:1 ratio. The K(+)-NH4+ antiporter was inhibited by verapamil and Schering 28080 in a dose-dependent manner, was able to work in reverse mode, and did not show any affinity for H+ as a substrate, indicating that it is distinct from other NH4(+)-carrying transporters. We conclude that a unique transporter, a potassium-ammonium (K+/NH4+) antiport, is responsible for NH4+ transport in renal inner medullary collecting duct cells. This antiporter is sensitive to verapamil and Schering 28080, is electroneutral, and is selective for NH4+ and K+ as substrates. The K+/NH4+ antiporter may play a significant role in acid-base regulation by excretion of ammonium and elimination of acid.
Amlal
H
H
Department of Medicine, University of Cincinnati School of Medicine, OH, USA.
Soleimani
M
M
D000818:Animals; D017920:Antiporters / Q000037:antagonists & inhibitors / Q000378:metabolism*; D001464:Barium / Q000494:pharmacology; D027682:Cation Transport Proteins; D002460:Cell Line; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D017136:Ion Transport; D007679:Kidney Medulla / Q000166:cytology / Q000378:metabolism*; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000378:metabolism*; D008564:Membrane Potentials; D051379:Mice; D011188:Potassium / Q000378:metabolism; D026902:Potassium Channel Blockers; D000644:Quaternary Ammonium Compounds / Q000378:metabolism* / Q000494:pharmacology; D011802:Quinidine / Q000494:pharmacology
D017920:Antiporters; D027682:Cation Transport Proteins; D004791:Enzyme Inhibitors; D026902:Potassium Channel Blockers; D000644:Quaternary Ammonium Compounds; C043130:ammonium-potassium antiporter; D001464:Barium; D011802:Quinidine; D011188:Potassium