Effect of parathyroid hormone on Na +-dependent phosphate transport and cAMP-dependent 32P phosphorylation in brush border vesicles from isolated perfused canine kidneys

Abstract

Concentrative uptake of 32 P i induced by the dissipation of a Na + gradient (overshoot) was demonstrated in brush border membrane vesicles obtained from isolated perfused canine kidneys. Na +-dependent 32 P i transport was decreased in brush border vesicles from isolated kidneys perfused with parathyroid hormone (PTH) for 2 h compared to uptake measured in vesicles from kidneys perfused without PTH. Cyclic AMP-dependent 32P phosphorylation of a 62,000 M r protein band was demonstrable on autoradiograms of sodium dodecyl sulfate-polyacrylamide gels of membrane suspensions from kidneys perfused ± PTH. Evidence that perfusion with PTH resulted in cAMP-dependent phosphorylation in isolated kidneys from parathyroidectomized dogs (decreased cAMP-dependent 32P phosphorylation of the 62,000- M r band in brush border vesicles) was obtained after 2-h perfusion with PTH. Decreased 32P phosphorylation was not observed if membranes were allowed to dephosphorylate prior to 32P phosphorylation in vitro. We conclude that brush border vesicles from isolated perfused canine kidneys can be used to study the action of PTH on Na +- P i cotransport in brush border membranes and on cAMP-dependent phosphorylation of the membrane. It is strongly suggested that PTH effects changes in Na +-dependent 32 P i transport in isolated brush border vesicles and changes in 32P phosphorylation of vesicles via a direct action on the renal cortical cell rather than as a consequence of extrarenal actions of the hormone.