Thapsigargin stimulates intracellular calcium mobilization and inhibits parathyroid hormone release

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Abstract
Ca2+ and other divalent cations like Sr2+, Ba2+, and Mg2+ stimulate rapid and sustained increases in intracellular Ca2+ ([Ca2+]i) and 1,4,5-inositol trisphosphate (1,4,5-InsP3) presumably by interacting with recently identified parathyroid cell membrane Ca2+ receptors. We used thapsigargin (THAPS), an inhibitor of the microsomal Ca2+-ATPase, to deplete InsP3-sensitive intracellular Ca2+ stores to determine whether sustained increases in [Ca2+]i due to divalent cations require intact cytosolic Ca2+ pools. In Fura 2-loaded parathyroid cells, THAPS produced a gradual increase in [Ca2+]i which reached a steady-state level by 2–3 minutes. The effect of THAPS (3 × 10−6 M) was substantial with [Ca2+]i, rising from 281 ± 27 nM at 0.5 mM Ca2+ to a peak value of 684 ± 30 nM (p < 0.0001). The addition of Sr2+ to cells at 0.5 mM extracellular Ca2+ induced an immediate 2-to 3-fold increase in [Ca2+]i which stabilized at a [Ca2+]i above baseline for ≥10 minutes. THAPS (3 × 10−6 M) pretreatment for ≥5 minutes blocked this sustained-phase increment in [Ca2+]i due to Sr2+. In the absence of extracellular Ca2+, there was a slight but nonsignificant effect of THAPS on [Ca2+]i. Incubation of cells with THAPS did not change the levels of 3H-inositol phosphates (InsP3, InsP2, and InsP1) or alter Sr2+-induced accumulation of InsP3, InsP2, and InsP1. THAPS substantially reduced parathyroid hormone secretion at 1.0 mM Ca2+ by 20 ± 16, 57 ± 8, 75 ± 10, and 83 ± 9% at 10−7, 3 × 10−7, 10−6, and 3 × 10−6 M THAPS, respectively. We conclude that depletion of intracellular Ca2+ stores by THAPS stimulates Ca2+ mobilization, presumably from extracellular sources, and that this agent and divalent cations such as Sr2+ activate the same pathway for sustained Ca2+ mobilization. The inhibition of secretion by THAPS supports the idea that increases in [Ca2+]i play a suppressive role in the control of hormone release in the parathyroid.
Funding Information
  • VA Merit Review (R29 DK39594)
  • NIH (DK43400)