Evidence for cAMP-dependent protein kinase in mediating the parathyroid hormone-stimulated rise in cytosolic free calcium in rabbit connecting tubules.
We investigated cellular mechanisms mediating the parathyroid hormone (PTH)-induced increase in cytosolic free Ca2+ concentration ([Ca2+]i) in isolated perfused rabbit connecting tubules. Prior and/or concomitant exposure to 0.5 mM of N-[2-(methylamino)ethyl]-5-isoquinolinesulfonamide dihydrochloride (H-8), a cyclic nucleotide-dependent protein kinase inhibitor, abolished the rise in [Ca2+]i produced by 0.1 nM PTH in five connecting tubules and suppressed it by approximately 50% in another five. In the latter, there was a delayed onset in the rise of [Ca2+]i. Such responses contrasted to the prompt increase in [Ca2+]i in PTH-stimulated control tubules. However, when H-8 was withdrawn, [Ca2+]i rose within minutes to reach a plateau value similar to the uninhibited response to PTH in controls, indicating rapidly reversible inhibition by H-8. In an otherwise identical protocol, 0.5 mM H-8 also reversibly suppressed the rise in [Ca2+]i induced by 0.175 mM 8-Br-cAMP. In contrast to the stimulatory effect of 8-Br-cAMP on [Ca2+]i, 1 mM 8-Br-cGMP caused no increase. At a concentration of 0.4 mM, the Rp diastereomer of adenosine cyclic 3',5'-phosphorothioate (Rp-cAMPS), a well-characterized cAMP-dependent protein kinase inhibitor, totally abolished the rise in [Ca2+]i caused by 0.1 nM PTH. We conclude that a cAMP-dependent protein kinase plays an important role in the PTH-stimulated rise in [Ca2+]i in the rabbit connecting tubule. Since the increase in [Ca2+]i was shown previously to depend on extracellular Ca2+, we propose that cAMP-dependent protein phosphorylation is important in mediating PTH-stimulated Ca2+ fluxes across plasma membranes of connecting tubule cells.
Lau
K
K
Department of Medicine, Michael Reese Hospital and Medical Center, Chicago, Illinois 60616.
Bourdeau
J E
JE
D000818:Animals; D002118:Calcium / Q000502:physiology*; D000242:Cyclic AMP / Q000031:analogs & derivatives / Q000502:physiology*; D003600:Cytosol / Q000502:physiology; D066298:In Vitro Techniques; D007546:Isoquinolines / Q000494:pharmacology; D007684:Kidney Tubules / Q000502:physiology*; D010281:Parathyroid Hormone / Q000502:physiology*; D047428:Protein Kinase Inhibitors; D011494:Protein Kinases / Q000502:physiology*; D011817:Rabbits
D007546:Isoquinolines; D010281:Parathyroid Hormone; D047428:Protein Kinase Inhibitors; C053177:N-(2-(methylamino)ethyl)-5-isoquinolinesulfonamide; D000242:Cyclic AMP; D011494:Protein Kinases; D002118:Calcium