ABSTRACT
The thiazide-sensitive Na(+)-Cl(-) cotransporter (TSC) is the major pathway for salt reabsorption in the apical membrane of the mammalian distal convoluted tubule. When expressed in Xenopus laevis oocytes, rat TSC exhibits high affinity for both cotransported ions, with the Michaelis-Menten constant (K(m)) for Na(+) of 7.6 +/- 1.6 mM and for Cl(-) of 6.3 +/- 1.1 mM, and Hill coefficients for Na(+) and Cl(-) consistent with electroneutrality. The affinities of both Na(+) and Cl(-) were increased by increasing concentration of the counterion. The IC(50) values for thiazides were affected by both extracellular Na(+) and Cl(-). The higher the Na(+) or Cl(-) concentration, the lower the inhibitory effect of thiazides. Finally, rTSC function is affected by extracellular osmolarity. We propose a transport model featuring a random order of binding in which the binding of each ion facilitates the binding of the counterion. Both ion binding sites alter thiazide-mediated inhibition of transport, indicating that the thiazide-binding site is either shared or modified by both Na(+) and Cl(-).
Subject(s)
Carrier Proteins/metabolism , Receptors, Drug/metabolism , Sodium Chloride Symporter Inhibitors/metabolism , Sodium Chloride Symporter Inhibitors/pharmacology , Sodium/metabolism , Symporters , Animals , Bendroflumethiazide/metabolism , Bendroflumethiazide/pharmacology , Binding Sites/drug effects , Biological Transport/drug effects , Carrier Proteins/genetics , Chlorides/metabolism , Chlorides/pharmacology , Diuretics , Hydrochlorothiazide/metabolism , Hydrochlorothiazide/pharmacology , Hydrogen-Ion Concentration , Inhibitory Concentration 50 , Kinetics , Metolazone/metabolism , Metolazone/pharmacology , Microinjections , Models, Biological , Oocytes/drug effects , Oocytes/metabolism , Osmolar Concentration , Polythiazide/metabolism , Polythiazide/pharmacology , Rats , Receptors, Drug/genetics , Sodium/pharmacology , Sodium Chloride Symporters , Solute Carrier Family 12, Member 3 , XenopusABSTRACT
An assay for polythiazide, sufficiently sensitive to measure plasma concentrations of this high-potency diuretic agent, has been developed. The assay is based on acid hydrolysis to trifluoroethylthioacetaldehyde and electron-capture gas chromatography. Sensitivity down to 0.2 ng/ml was achieved. In a study in normal human subjects receiving single 1-mg oral doses of polythiazide, the mean plasma half-lives for absorption and elimination were 1.2 and 25.7 hr, respectively. The latter is consistent with the extended duration of action of polythiazide. Approximately 25% of the drug was excreted unchanged in the urine.
