An epithelial high-affinity amiloride-binding site, different from the Na+ channel.
Specific binding of the radioactive amiloride analogues [3H]phenamil and [3H]benzamil was studied in plasma membrane from chicken lower intestine. A single population of sites whose affinities and specificities towards pyrazinecarboxamides roughly resemble those of the epithelial Na+ channel, was identified. However, a matched comparison of pyrazinecarboxamide binding and Na+ transport inhibition revealed substantial differences between the high-affinity [3H]phenamil-binding site detected, and the site whose occupancy by phenamil blocks Na+ transport. First, 5-(N-ethyl-N-isopropyl)-amiloride was found to displace bound [3H]phenamil at concentrations that are at least 10-fold lower than those needed to block the channel. Second, the rates at which [3H]phenamil associates and dissociates from this site are lower than the rates at which Na+ channels are inhibited and reactivated, under similar conditions. A site with high affinity to both amiloride and 5-(N-ethyl-N-isopropyl)-amiloride was detected also in membranes from other epithelia. We conclude that tight epithelia contain a major high-affinity amiloride receptor other than the Na(+)-conducting channel, the Na+/H+ antiport or the Na+/Ca2+ exchanger. This site could be associated with a pool of nonconducting channels, another (but structurally related) channel, or a totally unrelated protein.
Department of Membrane Research and Biophysics, Weizmann Institute of Science, Rehovot, Israel.
O
O
Goldstein
C
C
Asher
P
P
Barbry
E
E
Cragoe
W
W
Clauss
H
H
Garty
D000450:Aldosterone / Q000494:pharmacology; D000584:Amiloride / Q000031:analogs & derivatives* / Q000378:metabolism* / Q000494:pharmacology; D000818:Animals; D001665:Binding Sites; D001667:Binding, Competitive; D002024:Bufo marinus; D002462:Cell Membrane / Q000378:metabolism; D002645:Chickens; D003106:Colon / Q000378:metabolism; D004848:Epithelium / Q000378:metabolism; D007420:Intestine, Large / Q000378:metabolism*; D007700:Kinetics; D012964:Sodium / Q000378:metabolism*; D015222:Sodium Channels / Q000378:metabolism*; D012982:Sodium, Dietary / Q000494:pharmacology; D001743:Urinary Bladder / Q000187:drug effects / Q000378:metabolism*
D015222:Sodium Channels; D012982:Sodium, Dietary; C013407:benzamil; C047088:phenylamil; D000450:Aldosterone; D000584:Amiloride; D012964:Sodium; C039614:ethylisopropylamiloride