[3H]ethylpropylamiloride, a radio-labelled diuretic for the analysis of the Na+/H+ exchange system. Its use with kidney cell membranes.
The interaction of amiloride and several amiloride derivatives with the Na+/H+ exchange system in Madin-Darby canine kidney cells and in rabbit renal microvillus membrane vesicles was studied from 22Na+ uptake experiments. On both types of preparation, the order of potency of the different molecules tested is: ethylisopropylamiloride greater than ethylpropylamiloride (EPA) greater than amiloride greater than benzamil. 3H-labelled EPA was prepared and used to titrate amiloride binding sites in solubilized microvillus membranes. Kinetics experiments, equilibrium binding studies and competition experiments between [3H]EPA and unlabelled EPA indicate that EPA recognizes a single family of binding sites with a Kd value of 45 nM and a maximum binding capacity of 2 pmol/mg of protein. The order of potency of different amiloride analogs tested in [3H]EPA competition experiments is identical to that found for the inhibition of 22Na+ uptake by the Na+/H+ exchange system, suggesting that [3H]EPA binding sites are associated with the Na+/H+ exchange system. [3H]EPA binding sites are pharmacologically distinct from those of [3H]benzamil and [3H]bumetanide in kidney membranes.
D000584:Amiloride / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D001667:Binding, Competitive; D002352:Carrier Proteins / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D007668:Kidney / Q000378:metabolism*; D007700:Kinetics; D008871:Microvilli / Q000378:metabolism; D011719:Pyrazines / Q000378:metabolism*; D011817:Rabbits; D017923:Sodium-Hydrogen Exchangers; D013329:Structure-Activity Relationship; D014316:Tritium
D002352:Carrier Proteins; D011719:Pyrazines; D017923:Sodium-Hydrogen Exchangers; D014316:Tritium; C043988:5-(ethylpropyl)amiloride; D000584:Amiloride