AJP - Renal Physiology, Vol 252, Issue 3 517-F524, Copyright © 1987 by American Physiological Society
N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline, amiloride analogues, and renal Na+/H+ antiporter
V. K. Rocco, E. J. Cragoe Jr and D. G. Warnock
N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline (EEDQ) is a
carboxyl-activating agent and has been shown to inhibit the renal Na+/H+
antiporter. The purposes of the present studies were to characterize the
kinetics of inhibition of the Na+/H+ antiporter by EEDQ and to determine
whether amiloride analogues affect the ability of EEDQ to inhibit the rate
of Na+/H+ exchange. Brush-border membrane vesicles (BBMV) were prepared
from rabbit kidneys; Na+/H+ exchange rate was assessed by the fluorescence
quenching of acridine orange. EEDQ produced a concentration-dependent
inhibition of Na+/H+ exchange; the effect was to decrease the maximum
activity (Vmax) from 5.51 to 2.03 fluorescence units X mg protein-1 X S-1)
and Km (from 14.1 to 8.7 mM) compared with control BBMV. Pretreatment of
BBMV with amiloride before the addition of EEDQ maintained both Vmax and Km
at values that were not significantly different from those for control
BBMV. Compared with a series of analogues, amiloride was only the third
most potent inhibitor of the rabbit renal Na+/H+ antiporter; amiloride,
however, provided the greatest protection against inhibition of the
antiporter by the subsequent addition of EEDQ. These findings suggest that
the 2-carbonylguanidininum moiety and 6-chloro atom are important for
binding of amiloride to sites at or near the antiporter; the group at
position 5 is important in determining the ability of amiloride to protect
against inhibition of the Na+/H+ antiporter by EEDQ. Finally, the ability
of amiloride to protect against inactivation of the renal Na+/H+ antiporter
by EEDQ is reversible.
