Kinetic analysis of electrogenic 2 Na+-1 H+ antiport in crustacean hepatopancreas.
Na+ uptake by short-circuited brush-border membrane vesicles of the hepatopancreatic epithelium from the freshwater prawn Macrobrachium rosenbergii was Cl- independent, amiloride sensitive, and stimulated by a transmembrane proton gradient ([H+]i greater than [H+]o). Na+ influx (3-s uptake) was a sigmoidal function of [Na]o (2.5-150 mM), when pHi = 6.0, pHo = 8.0, and followed the Hill equation for binding cooperativity [maximal Na+ influx (Jm) = 140.6 nmol mg-1s-1; affinity constant (K') = 82.2 mM Na+; Hill coefficient (n) = 2.07]. Influx kinetic analyses at physiological conditions suggested two external cation-binding sites shared by Na+ and H+ (proton dissociation constant Pk1 = 5.7; Pk2 = 4.0) and a single internal cation site used only by H+ (Pk = 6.5). Amiloride was a competitive inhibitor of Na+ transport at both external binding sites (Ki1 = 50 microM; Ki2 = 1,520 microM). Electrogenic Na+-H+ exchange by these vesicles was demonstrated using an equilibrium-shift method of analysis and a transmembrane electrical potential difference as the only driving force for transport. In addition, electrogenic net Na+ influx (3-s uptake) was observed in vesicles loaded with 5 mM 22Na at pH 7.0 and exposed to media containing several 22Na or proton concentrations. Results suggest the following exchange model: low [Na]o, (1 Na+ and 1 H+)-1 H+; high [Na+]o, 2 Na+-1 H+. This antiport mechanism may account for two major functional operations of the gastrointestinal tract in these animals: 1) proton secretion against considerable concentration gradients leading to stomach luminal acidification, and 2) Na+ absorption from lumen to cytoplasm potentially making a significant contribution to organismic ion balance.
Ahearn
G A
GA
Department of Zoology, University of Hawaii at Manoa, Honolulu 96822.
Clay
L P
LP
D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D033364:Decapoda / Q000378:metabolism*; D004064:Digestive System / Q000378:metabolism; D006859:Hydrogen / Q000378:metabolism; D007477:Ions; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008871:Microvilli / Q000378:metabolism; D009994:Osmolar Concentration; D010179:Pancreas / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism; D017923:Sodium-Hydrogen Exchangers
D002352:Carrier Proteins; D007477:Ions; D017923:Sodium-Hydrogen Exchangers; D000584:Amiloride; D006859:Hydrogen; D012964:Sodium