[3H]phenamil, a radiolabelled diuretic for the analysis of the amiloride-sensitive Na+ channels in kidney membranes.
The interaction of amiloride and amiloride derivatives with the Na+ channels of pig kidney membranes was studied from 22Na+ uptake experiments. The order of potency of the different molecules tested is: phenamil greater than benzamil greater than amiloride, ethylisopropylamiloride. [3H]labelled phenamil was prepared and used to titrate Na+ channels in pig kidney membranes. Kinetics experiments, equilibrium binding studies and competition experiments between [3H]phenamil and unlabelled phenamil indicate that phenamil recognizes a single family of binding sites with a Kd value of 20 nM and a maximum binding capacity of 11.5 pmol/mg of protein. The order of potency of different amiloride analogs tested in [3H]phenamil competition experiments is identical to that found for the inhibition of 22Na+ uptake by apical Na+ channels.
Barbry
P
P
Frelin
C
C
Vigne
P
P
Cragoe
E J
EJ
Lazdunski
M
M
D000584:Amiloride / Q000031:analogs & derivatives* / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D004305:Dose-Response Relationship, Drug; D004848:Epithelium / Q000378:metabolism; D066298:In Vitro Techniques; D007473:Ion Channels / Q000502:physiology*; D007668:Kidney / Q000187:drug effects / Q000378:metabolism*; D012964:Sodium / Q000378:metabolism*; D013552:Swine; D014316:Tritium
D007473:Ion Channels; C013407:benzamil; D014316:Tritium; C047088:phenylamil; C039465:5-dimethylamiloride; D000584:Amiloride; D012964:Sodium