Cl- channels in basolateral renal medullary membranes: VII. Characterization of the intracellular anion binding sites.
A unique property of basolateral membrane Cl- channels from the mTAL is that the Cl- concentration facing the intracellular aspects of these channels is a determinant of channel open time probability (Po). The K1/2 for maximal activation of Po by Cl- facing intracellular domains of these channels is 10 mM Cl-. The present experiments evaluated the nature of these Cl(-)-interactive sites. First, we found that the impermeant anion isethionate, when exposed to intracellular Cl- channel faces, could augment Po with a K1/2 in the range of 10 mM isethionate without affecting conductance (gCl, pS). Second, pretreatment of the solutions facing the intracellular aspects of the channels with either 1 mM phenylglyoxal (PGO), an arginine-specific reagent, or the lysine/terminal amine reagent trinitrobenzene sulfonic acid (TNBS, 1 mM), prevented the activation of Po usually seen when the Cl- concentration of solutions facing intracellular channel domains was raised from 2 to 50 mM. However, when the Cl- channel activity was increased by first raising the Cl- concentration bathing intracellular channel faces from 2 to 50 mM, subsequent addition of either PGO or TNBS to solutions bathing intracellular Cl- channel faces had no effect on Po. We conclude that the intracellular aspects of these Cl- channels contain Cl(-)-interactive loci (termed [Cl]i) which are accessible to impermeant anions in intracellular fluids and which contain arginine- and lysine-rich domains which can be inactivated, at low ambient Cl- or isethionate concentrations, by interactions with PGO or TNBS.
Winters
C J
CJ
Department of Internal Medicine, University of Arkansas College of Medicine, Little Rock 72205.
Reeves
W B
WB
Andreoli
T E
TE
D000818:Animals; D000838:Anions / Q000378:metabolism*; D001120:Arginine / Q000032:analysis; D001665:Binding Sites; D002462:Cell Membrane / Q000378:metabolism / Q000502:physiology / Q000648:ultrastructure; D018118:Chloride Channels / Q000032:analysis* / Q000502:physiology; D007513:Isethionic Acid / Q000494:pharmacology; D007679:Kidney Medulla / Q000737:chemistry* / Q000166:cytology / Q000648:ultrastructure; D008239:Lysine / Q000032:analysis; D008564:Membrane Potentials / Q000502:physiology; D010658:Phenylglyoxal / Q000494:pharmacology; D011817:Rabbits; D014302:Trinitrobenzenesulfonic Acid / Q000494:pharmacology
D000838:Anions; D018118:Chloride Channels; D014302:Trinitrobenzenesulfonic Acid; D001120:Arginine; D007513:Isethionic Acid; D008239:Lysine; D010658:Phenylglyoxal