AJP - Cell Physiology, Vol 271, Issue 5 C1457-C1462, Copyright © 1996 by American Physiological Society
Inhibition of amiloride-sensitive Na+ channel by isothiouronium derivatives
A. Avigdor, C. Asher, D. M. Tal, S. J. Karlish and H. Garty
Department of Membrane Research, Weizmann Institute of Science, Rehovot, Israel.
The effects on the amiloride-blockable Na+ channel of a family of recently
synthesized isothiouronium derivatives were measured in plasma membrane
vesicles from rat distal colon. Some of these derivatives act as
high-affinity Na(+)-like antagonists on the
Na(+)-K(+)-adenosinetriphosphatase. One of the reagents tested,
1-bromo-2,4,6-tris(isothiouronium methyl)-benzene tribromide (Br-TITU), was
found to be a potent blocker of the Na+ channel. At neutral pH, Br-TITU
rapidly inhibits the channel mediated 22Na+ uptake, with an inhibition
constant of 94 +/- 39 nM. The inhibition observed is specific and
reversible. 1,3-Dibromo-2,4,6-tris(isothiouronium methyl)benzene tribromide
and Br-TITU derivatives with methyl and phenyl substitutions on the
isothiouronium moiety were much less effective blockers. Incubation of
cells with Br-TITU at alkaline (but not neutral) pH produces irreversible
inactivation of channels, possibly due ot covalent modification of a lysine
residue. This inactivation can be attenuated by amiloride but not by Na+.
Thus Br-TITU may be a useful reagent in identifying essential residues of
the channel protein.
