Hormonal regulation of (Ca2+ + Mg2+)ATPase activity in canine renal basolateral membrane.

ABSTRACT

High affinity (Ca2+ + Mg2+)ATPase activity was demonstrated in proximal tubule basolateral membranes (BLM) obtained from canine kidney. The Km of the enzyme for free Ca2+ was 0.12 +/- 0.02 microM, and at 3 microM free Ca2+, the enzyme reached its maximal velocity. To evaluate hormonal regulation of this enzyme, we studied the in vitro effects of several polypeptide hormones on enzyme activity. We measured the effects of insulin and human (h) PTH-(1-34) and their inactive analogs desoctapeptide insulin, bovine (b) PTH-(3-34), and oxidized hPTH-(1-34); insulin-like growth factors (IGFs) I and II; calcitonin; and the common cellular mediator for PTH and calcitonin, cAMP. At 0.1 microM free Ca2+, insulin (25-100 microU/ml) increased (Ca2+ + Mg2+)ATPase activity in a dose-dependent manner by 35-52% (P less than 0.01) and by 8-13% (P less than 0.05 to P less than 0.01) at a 3-microM free Ca2+ concentration; hPTH-(1-34) PTH (10(-9)-10(-7) M) increased the enzyme activity at a free Ca2+ concentration of 0.1 microM by 13-25% (P less than 0.05 to P less than 0.01). IGF-I increased (Ca2+ + Mg2+)ATPase activity by 40% (P less than 0.05) at 0.1 microM free Ca2+ at high peptide concentrations (10 ng/ml). No effect was obtained at 2 ng/ml IGF-I. cAMP (10(-7)-10(-4) M) stimulated enzyme activity by 18-27% (P less than 0.05 to P less than 0.02) at 0.1 microM Ca2+ and by 8-12% (P less than 0.05 to P less than 0.01) at 3 microM Ca2+. The effects of insulin and cAMP on (Ca2+ + Mg2+)ATPase activity were additive. No effect on the enzyme activity was obtained by the inactive analogs desoctapeptide insulin, bPTH-(3-34), and oxidized hPTH-(1-34), or by calcitonin or IGF-II. The data suggest that insulin and PTH have a specific stimulatory effect on (Ca2+ + Mg2+)ATPase activity in canine kidney proximal tubular BLM. While the insulin action is independent of cAMP, a role of cAMP in the regulatory effect of PTH on this enzyme cannot be ruled out. The direct stimulatory effect of insulin and PTH on (Ca2+ + Mg2+)ATPase in canine kidney proximal tubular BLM suggests that these hormones mediate their cellular effects in part by changes in cellular calcium homeostasis.