Electrode sandwich technique: a method for studying K+ transport in renal cells.
Distal tubules were harvested from frog kidney and placed on the membrane of a K+-selective macroelectrode. Then the renal tissue was covered with a dialysis membrane to produce a closed extracellular compartment with a constant volume (40 microliter). K+ fluxes in and out of the cells could be determined, since the steady-state K+ activity during constant perfusion changed to a new steady state when perfusion was stopped. Inhibition of passive K+ permeability by the addition of Ba2+ resulted in K+ uptake by the cells because of the function of the Na+-K+ pump. Inhibition of the pump by the addition of ouabain led to K+ efflux from cells reflecting the passive K+ permeability. Because K+ net movement under control conditions (no Ba2+ or ouabain) results from both uptake and efflux, subtraction of K+ uptake (in the presence of Ba2+) from control K+ net flux reveals the passive K+ efflux. This value agrees well with that obtained with ouabain. Furosemide led to a significant K+ shift from the extracellular compartment into the intracellular compartment. Reduction of extracellular pH from 7.8 to 6.0 decreased the rate of K+ uptake by 39 +/- 7% and the K+ leak by 51 +/- 11%. We conclude that K+ uptake and K+ release can be functionally separated. This so-called "electrode sandwich technique" permits evaluation of pump and leak independently in the same cell population.
Westphale
H J
HJ
Department of Physiology, University of Würzburg, Federal Republic of Germany.
Oberleithner
H
H
D000818:Animals; D001464:Barium / Q000494:pharmacology; D004566:Electrodes; D004594:Electrophysiology / Q000295:instrumentation / Q000379:methods; D005665:Furosemide / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007684:Kidney Tubules / Q000378:metabolism*; D007686:Kidney Tubules, Distal / Q000187:drug effects / Q000378:metabolism*; D007700:Kinetics; D010042:Ouabain / Q000494:pharmacology; D010477:Perfusion; D011188:Potassium / Q000378:metabolism*; D011893:Rana esculenta
D001464:Barium; D010042:Ouabain; D005665:Furosemide; D011188:Potassium