HEPARIN-INSENSITIVE CALCIUM-RELEASE FROM INTRACELLULAR STORES TRIGGERED BY THE RECOMBINANT HUMAN PARATHYROID-HORMONE RECEPTOR

1 In the present study we have characterized the parathyroid hormone (PTH)-induced calcium signalling in 293 cells stably transfected with the human PTH receptor cDNA. In these cells, human PTH-I(1-38) strongly stimulates adenosine 3':5'-cyclic monophosphate (cyclic AMP) formation (EC(50) = 0.39 nM) but fails to activate phosphoinositide (PI) turnover. The latter pathway is strongly activated, however, by carbachol (CCh) acting through endogenous M(3)-muscarinic receptors. 2 Despite the lack of detectable inositol phosphate (IF) formation, hPTH-(1-38) elicited calcium transients (EC(50) = 11.2 nM) which were comparable to the signals evoked by CCh. These signals are independent of cyclic AMP generation as cyclic AMP elevating agents did not mimic or modify the PTH response. 3 The PTH-stimulated calcium signal still occurred in calcium-free medium but was absent in cells pretreated with thapsigargin, an inhibitor of the calcium pump of the endoplasmic reticulum (ER). hPTH-(1-38) did not accelerate Mn2+-influx through the plasma membrane. These data indicate that PTH releases calcium from intracellular stores. 4 Using heparin, an inhibitor of the IP3-activated calcium release channel of the ER, we tested whether the formation of a low amount of IP3, escaping detection by our biochemical assay, might be the origin of the PTH-induced calcium response. However, intracellular infusion of heparin through patch pipettes in voltage clamp experiments failed to block hPTH-(1-38)-induced calcium signals, whereas it abolished the CCh response. 5 The PTH response, like the CCh response, was insensitive to micromolar concentrations of ryanodine and ruthenium red, eliminating the possibility that hPTH-(1-38) stimulates calcium-induced calcium release through ryanodine receptors. 6 We conclude that the recombinant human PTH receptor stimulates calcium release from intracellular stores through a novel pathway not involving IP3- or ryanodine receptors.