AJP - Renal Physiology, Vol 245, Issue 2 263-F271, Copyright © 1983 by American Physiological Society
Nucleotide inhibition of phosphate transport in the renal proximal tubule
R. P. Lang, N. Yanagawa, E. P. Nord, L. Sakhrani, S. H. Lee and L. G. Fine
The observation that NAD inhibits sodium-dependent phosphate (P) uptake by
the luminal brush border membrane (BBM) of the proximal tubule prompted us
to examine the specificity and mechanism of this process. Addition of
10(-5) M NAD to the perfusate of isolated perfused rabbit proximal straight
tubules inhibited lumen-to-bath P flux by approximately 50%. ADP-ribose had
an identical effect, whereas nicotinamide had no effect. ADP and 5'-AMP
(10(-5) M) also inhibited P flux. Na-dependent uptake of 32P by rabbit BBM
vesicles was inhibited by 0.1-0.3 mM NAD, ADP-ribose, ADP, ATP, 5'-AMP, and
GDP, which were preincubated with the vesicles for 30 min. The kinetics of
inhibition showed an apparent increase in the Km for P but no change in
Vmax. These findings are consistent with "competitive inhibition." The
nucleotides inhibited P uptake even when BBM alkaline phosphatase was
inhibited by 96% with 10 mM theophylline. Evidence of nonspecific
phosphatase activity was present, since incubation of BBM with 0.1 mM
solution of nucleotides for 30 min resulted in an elevation of free P in
the medium of approximately 0.15-0.22 mM. Correction of 32P specific
activity for this change resulted in values for Km and Vmax that were not
significantly different from control. The "competitive inhibition" could
thus be ascribed to an isotope-dilution effect. There was no evidence to
suggest that NAD caused ADP-ribosylation of the luminal membrane. These
studies indicate that adenine and guanine nucleotides do not inhibit P
transport by a direct action on the luminal membrane of the proximal tubule
but do inhibit lumen-to-bath P flux in isolated perfused proximal tubules
at concentrations of 10(-5) M. Since there is no direct inhibitory effect
of these compounds at the level of the BBM, it is possible that they
inhibit P transport by altering some event subsequent to the transfer of P
across the luminal membrane.
