Desensitization to Parathyroid Hormone in the Isolated Perfused Canine Kidney: Reversal of Altered Receptor-Adenylate Cyclase System by Guanosine Triphosphate in Vitro*

Abstract

The mechanism of desensitization to PTH was examined in the isolated perfused canine kidney. Paired kidneys from six mongrel dogs were perfused for 20–22 h in vitro. Synthetic bovine PTH-(1–34) (1.5 μg/h) was administered to one of each pair of kidneys. Adenylate cyclase activity and PTH receptor binding were studied in purified basolateral cortical membranes. The specific binding of synthetic bovine PTH-(1– 34) was reduced in the PTH-treated kidneys (maximum binding, 1.47 ± 0.43 fmol eq/μg protein vs. 3.28 ± 0.66 in the control). These data correlated with a decrease in hormone-stimulated adenylate cyclase activity (maximum velocity, 2055 ± 193 pmol cAMP produced/mg membrane protein in 30 min in the PTHtreated and 3440 ± 322 in the control kidneys). In the presence of 10^-3 M GTP, PTH-stimulated adenylate cyclase activities in control and PTH-treated kidneys were similar. In further studies, membranes from control and PTH-treated kidneys were preincubated with GTP to dissociate receptor-bound PTH. This incubation medium was assayed for immunoreactive PTH (iPTH), and the membranes were washed extensively to remove GTP. After this procedure, PTH binding and adenylate cyclase activity in the membranes from the PTH-treated kidneys became very similar to those in control kidneys. iPTH was detected in the medium of the PTH-treated membranes, but not in that from control membranes. iPTH was not detected if GTP was omitted from the preincubation. These data show that exposure to PTH results in decreased PTH binding and decreased PTHstimulated adenylate cyclase activity. Resensitization was achieved in vitro by GTP, which dissociates bound PTH. Thus, in this experimental model, desensitization is due to persistent receptor occupancy as a consequence of altered guanine nucleotide regulation of the hormone-receptor complex.