title: Nongenomic activation of the calcium message system by vitamin D metabolites in osteoblast-like cells.
abstract: 1,25-dihydroxycholecalciferol (1,25(OH)2D3) rapidly affects calcium (Ca2+) transport in several cell systems, suggesting physiological actions independent of genomic activation. To test this hypothesis, we studied immediate to early effects (0.5-300 sec) of 1,25(OH)2D3 on cytosolic Ca2+ [Ca2+]i in single osteogenic sarcoma ROS 17/2.8 cells loaded with fura-2. An acute rise in [Ca2+]i was observed in 40% of the cells following addition of 1,25(OH)2D3, with a threshold concentration of 10(-11) M. In most cases, the [Ca2+]i rise was transient, with return to baseline within 1 min; less frequently a more prolonged effect was observed, with variable recovery times. 25-hydroxycholecalciferol (25(OH)D3) reproduced the effect of 1,25(OH)2D3 on [Ca2+]i, with equal potency and similar responses, whereas 24,25-dihydroxycholecalciferol, 1 alpha-hydroxycholecalciferol, and 22 oxa-1,25(OH)2D3 were not effective. 1,25(OH)2D3 also increased [Ca2+]i in ROS 24/1 cells, which are defective of receptors for the vitamin D metabolites. At high doses (10(-8)-10(-7) M) of 1,25(OH)2D3 the [Ca2+]i rise in ROS 17/2.8 cells was due to both influx of extracellular Ca2+ and release of Ca2+ from intracellular stores, as the effect was only partially inhibited by Ca2(+)-channel blockade by nifedipine. At low doses (10(-9)-10(-10) M), the effect was entirely dependent on extracellular Ca2+. 1,25(OH)2D3 also increased the production of inositol 1,4,5 trisphosphate (Ins(1, 4, 5)P3) and diacylglycerol, at a threshold dose of 10(-9) M, indicating activation of phospholipase C (PLC). In two thirds of the cells studied, a second addition of 1,25(OH)2D3 within 5 min to cells prestimulated with equimolar doses of the vitamin D metabolite resulted in a [Ca2+]i transient of higher amplitude than the first, a phenomenon occurring at all doses of the hormone, and associated with production of Ins(1, 4, 5)P3. This response amplification was not produced by 25(OH)D3, and pretreatment with 1 alpha(OH)D3 did not significantly enhance 1,25(OH)2D3-induced production of Ins(1, 4, 5)P3. In conclusion, activation of the Ca2+ message system by vitamin D metabolites is a rapid, nongenomic effect; 1,25(OH)2D3 specifically activates both PLC and dihydropyridine-sensitive Ca2+ channels, and "primes" the cells to respond with an enhanced [Ca2+]i rise to a subsequent homologous stimulation; the presence of both the 1 alpha and 25 hydroxyl groups is necessary to express the full hormonal action of vitamin D on [Ca2+]i.
authors.affiliation: Division of Bone and Mineral Metabolism, Jewish Hospital, Washington University Medical Center, St. Louis, Missouri 63110.
authors.forename: R
authors.initials: R
authors.lastname: Civitelli
authors.forename: Y S
authors.initials: YS
authors.lastname: Kim
authors.forename: S L
authors.initials: SL
authors.lastname: Gunsten
authors.forename: A
authors.initials: A
authors.lastname: Fujimori
authors.forename: M
authors.initials: M
authors.lastname: Huskey
authors.forename: L V
authors.initials: LV
authors.lastname: Avioli
authors.forename: K A
authors.initials: KA
authors.lastname: Hruska
mesh_terms: D000818:Animals; D002112:Calcifediol / Q000494:pharmacology; D002117:Calcitriol / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism / Q000502:physiology*; D006887:Hydroxycholecalciferols / Q000494:pharmacology; D009994:Osmolar Concentration; D010006:Osteoblasts / Q000378:metabolism / Q000502:physiology*; D015398:Signal Transduction; D014407:Tumor Cells, Cultured; D014807:Vitamin D / Q000378:metabolism*
chemical_list: D006887:Hydroxycholecalciferols; D014807:Vitamin D; D002117:Calcitriol; D002112:Calcifediol; D002118:Calcium; C008088:alfacalcidol