Reconstituted amiloride-inhibited sodium transporter from rabbit kidney medulla is responsible for Na +-H + exchange

Abstract

Microsomes formed from rabbit kidney medulla and reconstituted proteoliposomes formed from these microsomes were capable of amiloride-inhibited Na + transport that was insensitive to valinomycin either with or without K +. This indicated that the Na + transport process was electroneutral. This Na + transport process was insensitive to extravesicular Cl − or HCO 3 − and not stimulated by high intravesicular gradients of K +, Ca 2+ or Mg 2+, which indicated that the process did not require NaCl or NaHCO 3 co-transport or Na +/K +, Na +/Ca 2+ or Na +/Mg 2+ counter-transport. Na + uptake into microsomes or proteoliposomes was inhibited by extravesicular K +, Ca 2+, Mg 2+ or La 3+, which indicated that these ions interacted with the Na +-binding site on the transport protein. Na + uptake into microsomes was stimulated by intravesicular protons and inhibited by extravesicular protons. This suggested that microsomes were capable of Na +-H + exchange and this was confirmed when Na + was shown to stimulate H + efflux from microsomes. The amiloride-inhibited Na + transporter from medulla microsomes which has been reconstituted into proteoliposomes is most likely a Na +-H + exchanger. This Na + transporter was totally insensitive to the uncoupler 1799, either in the presence or absence of valinomycin plus K + and less sensitive to NH 3 than to amiloride. This indicated that amiloride inhibited Na + transport not merely by acting as a weak-base uncoupler but by directly interacting with the protein responsible for Na +-H + exchange.