Abstract
Cells were isolated from the thick ascending limb of Henle's loop of rabbit kidney outer medulla and a plasma membrane fraction was prepared by differential centrifugation. Sodium and rubidium uptake into the plasma membrane vesicles were determined by a rapid filtration technique. In the presence of a 100 mM KCl gradient and 0.5 mM sodium the vesicles took up 252±82 pmoles sodium/mg protein x 15s; 52% of the uptake was dependent on the presence of chloride or inhibited by 10−3 M bumetanide. If KCl was stepwise replaced by choline chloride, sodium uptake decreased in the absence of bumetanide but was only insignificantly altered in the presence of bumetanide. Potassium exerted a halfmaximum stimulation at 22.3±9.7 mM. In tracer exchange experiments under zero salt gradient conditions, sodium uptake was also strongly reduced in the absence of potassium. Rubidium uptake into the same membrane fraction was highest in the presence of a NaCl gradient, and decreased 41% when sodium was replaced by choline or when 10−3 M bumetanide was present. Replacement of sodium chloride by sodium nitrate also inhibited rubidium uptake. When the sodium, chloride and potassium dependence of the bumetanide sensitive sodium uptake was investigated in more detail, Hill coefficients for sodium of 1.0±0.03, for chloride of 1.8±0.2 and for potassium of 0.98±0.03 were obtained. These results are consistent with the presence of a Na−Cl−K cotransport system in the medullary thick ascending limb of Henle's loop which may operate with a stoichiometry of 1/2/1.
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Aull F (1981) Potassium chloride cotransport in steady-state ascites tumor cells. Does bumetanide inhibit? Biochim Biophys Acta 643:339–345
Bode F, Pockrandt-Hemstedt H, Baumann K, Kinne R (1974) Analysis of pinocytic process in rat kidney. J Cell Biol 63:998–1008
Brazy PC, Gunn RB (1976) Furosemide inhibition of chloride transport in human red blood cells. J Gen Physiol 68:583–599
Brown CDA, Simmons NL (1981) K+ fluxes in cultured renal epithelial cells of canine origin. J Physiol 319:99
Burg MB, Green N (1973) Function of the thick ascending limb of Henle's loop. Am J Physiol 224:659–668
Burg M, Stoner L, Cardinal J, Green N (1973) Furosemide effect on isolated perfused tubules. Am J Physiol 225:119–124
Chipperfeld AR (1980) An effect of chloride on (Na+K) cotransport in human red blood cells. Nature 286:281–282
Dunham PB, Stewart GW, Ellory JC (1980) Chloride-activated passive potassium transport in human erythrocytes. Proc Natl Acad Sci (USA) 77:1711–1715
Dunn MJ (1970) The effect of transport inhibitors on sodium outflux and influx in red blood cells: evidence for exchange diffusion. J Clin Invest 49:1804–1814
Eveloff J, Kinne R, Kinne-Saffran E, Murer H, Silva P, Epstein PH, Stoff J, Kinter WB (1978) Coupled sodium and chloride transport into plasma membrane vesicles prepared from dogfish rectal gland. Pflügers Arch 378:87–92
Eveloff J, Haase W, Kinne R (1980) Separation of renal medullary cells: isolation of cells from the thick ascending limb of Henle's loop. J Cell Biol 87:672–680
Eveloff J, Bayerdoerffer E, Haase W, Kinne R (1980) Biochemical and physiological studies on cells isolated from the medullary thick ascending limb of Henle's loop. Int J Biochem 12:55–59
Eveloff J, Bayerdoerffer E, Silva P, Kinne R (1981) Sodiumchloride transport in the thick ascending limb of Henle's loop: Oxygen consumption studies in isolated cells. Pflügers Arch 389:263–270
Eveloff J, Kinne R (1982) Use of isolated thick ascending limb of Henle's loop cells to study renal development. In: Spitzer A (ed) The kidney during development: morphology and function. Masson Publ, New York, pp 85–88
Eveloff J, Kinne R (1982) Sodium-chloride transport in the medullary thick ascending limb of Henle's loop: evidence for a sodium-chloride cotransport system in plasma membrane vesicles. J Membr Biol 72:173–181
Field M, Kimberg LS, Orellana S, Frizzell R (1981) Potassium dependence of chloride transport in the intestine of the flounder, Pseudopleuronectes americanus. Mt Desert Island Bull 21:93–95
Fleischer S, Fleischer B (1967) Removal and binding of polar lipids in mitochondria and other membrane systems. In: Estabrook RW, Pullman ME (eds) Methods of Enzymology X. Academic Press, New York, pp 406–433
Forbush III B, Palfrey HC (1982) Bumetanide and benzmetanide binding to membranes from shark rectal glands and canine kidney. Biophys J 37:161
Geck P, Heinz E, Pietrzyk C, Pfeiffer B (1978) Effect of furosemide on ouabain-insensitive K+ and Cl− movements in Ehrlich cells. In: Straub R, Bolis L (eds) Cell membrane receptors of drug and hormones. Raven Press, New York, pp 301–307
Geck P, Pietrzyk C, Burckhardt BC, Pfeiffer B, Heinz E (1980) Electrically silent cotransport of Na+, K+ and Cl− in Ehrlich cells. Biochim Biophys Acta 600:432–447
Geck P, Heinz E, Pfeiffer B (1981) Influence of high ceiling diuretics on ion fluxes and cell volume of Ehrlich ascites tumor cells. Scand Audiol Suppl 141:25–37
Greger R (1981) Cation selectivity of the isolated perfused cortical thick ascending limb of Henle's loop of rabbit kidney. Pflügers Arch 390:30–37
Greger R (1981) Chloride reabsorption in the rabbit cortical thick ascending limb of the loop of Henle. A sodium dependent process. Pflügers Arch 390:39–43
Greger R, Schlatter E (1981) Presence of luminal K+, a prerequisite for active NaCl transport in the cortical thick ascending limb of Henle's loop of rabbit kidney. Pflügers Arch 392:92–94
Greger R, Schlatter E, Lang F (1983) Evidence for electroneutral sodium chloride cotransport in the cortical thick ascending limb of Henle's loop of rabbit kidney. Pflügers Arch 396:308–314
Greger R, Schlatter E (1983) Properties of the lumen membrane of the cortical thick ascending limb of Henle's loop of rabbit kidney. Pflügers Arch 396:315–324
Greger R, Schlatter E (1983) Properties of the basolateral membrane of the cortical thick ascending limb of Henle's loop of rabbit kidney. A model for secondary active chloride transport. Pflügers Arch 396:325–334
Haas M, McManus TJ (1982) Bumetanide inhibition of (Na+K+2Cl) cotransport and K/Rb exchange at a chloride site in duck red cells. Biophys J 37:214a
Hannafin J, Kinne-Saffran E, Friedman D, Kinne R (1983) Presence of a sodium-potassion chloride cotransport system in the rectal gland of Squalus acanthias. J Membr Biol 75:73–83
Hoffmann N, Thees M, Kinne R (1976) Phosphate transport by isolated renal brush border vesicles. Pflügers Arch 362:147–156
Imai M (1977) Effect of bumetanide and furosemide on the thick ascending limb of Henle's loop of rabbits and rats perfused in vitro. Eur J Pharm 41:409–416
Kinne R, Kinne-Saffran E, Koenig B, Bayerdoerffer E, Eveloff J (1981) Control mechanism in active chloride transport. Scand Audiol Suppl 14:17–24
Krasny EJ, Halm DR, Frizzell R (1982) Apical membrane potassium conductance in flounder intestine: relation to chloride absorption. Fed Proc 41:1261
Lineweaver H, Burk D (1934) The determination of enzyme dissociation constants. J Amer Chem Soc 56:658
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurements with the folin reagent. J Biol Chem 193: 265–275
McManus TJ, Schmidt III WF (1978) Ion and co-ion transport in avian red cells. In: Hoffmann JF (ed) Membrane transport process. Raven Press, New York, pp 79–106
Erlinger S, McRoberts A, Rindler MJ, Saier Jr MH (1982) Furosemide-sensitive salt transport in the Madin-Darby canine kidney cell line. J Biol Chem 257(5):2260–2266
Musch M, Kimberg L, Orellana S, Field M, Frizzell R (1982) Active K transport in flounder intestine: cotransport with Na and Cl. Fed Proc 41:1495
Oberleithner H, Guggino W, Giebisch G (1982) Mechanism of distal tubular chloride transport in Amphiuma kidney. Am J Physiol 242:F332-F339
Oberleithner H, Guggino W, Giebisch G (1983) The effect of furosemide on luminal sodium, chloride and potassium transport in the early distal tubule of Amphiuma kidney. Pflügers Arch 396:27–33
Palfrey HC, Feit PW, Greengard P (1980) cAMP-stimulated cation cotransport in avian erythrocytes: inhibition by “loop” diuretics. Am J Physiol 238:C139-C148
Rocha A, Kokko JP (1973) Sodium chloride and water transport in the medullary thick ascending limb of Henle: Evidence for active chloride transport. J Clin Invest 52:612–623
Wallach DFH, Kamat VB (1966) Preparation of plasma membrane fragments from mouse ascites tumor cells. In: Neufeld EF, Ginsburg V (eds) Methods in Enzymology, vol 8. Academic Press, New York, pp 164–166
Wiley JS, Cooper RA (1974) A furosemide-sensitive cotransport of sodium plus potassium in the human red cell. J Clin Invest 53:745–755
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In partial fulfillment of the Ph.D. degree requirements for Barbara Koenig
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Koenig, B., Ricapito, S. & Kinne, R. Chloride transport in the thick ascending limb of Henle's loop: potassium dependence and stoichiometry of the NaCl cotransport system in plasma membrane vesicles. Pflugers Arch. 399, 173–179 (1983). https://doi.org/10.1007/BF00656711
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DOI: https://doi.org/10.1007/BF00656711