Activation of the Ca²⁺ Message System by Parathyroid Hormone Is Dependent on the Cell Cycle

To verify whether the heterogeneous intracellular calcium ([Ca²⁺]_i) responses to PTH observed in the UMR 106-01 osteogenic sarcoma cells are secondary to cell cycle asynchrony or to genotypic differences within the population, we synchronized cell monolayers at the G1/S boundary using a sequential thymidine-aphidicolin block. Video image analysis of fura-2-loaded cells revealed that PTH (10^-7 M) induced transient increases of [Ca²⁺]_i preferentially in cells in S phase (82% response frequency, n = 63; 286 ± 33% of baseline, n = 29), whereas cells in G1 phase responded poorly to PTH (10% response frequency, n = 51; 140 ± 8% of baseline, n = 5). In contrast, cell exposure to 2% fetal calf serum was followed by [Ca²⁺]_i transients in 83% (n = 42) of cells in G1 phase, but in only 25% (n = 63) of cells in S phase, with similar response amplitude. Hormonal responsiveness was heterogeneous in small clones obtained from single UMR 106-01 cells, with response frequency similar to that observed in nonsynchronized cultures. Pretreatment with either La³⁺, nifedipine, or pertussis toxin reduced both frequency and amplitude of PTH response in S phase to levels close to G1 phase, whereas there was no significant difference in inositol trisphosphate generated by PTH stimulation in either phase. Therefore, the heterogeneous [Ca²⁺]_i responses of UMR 106-01 cells to hormonal stimulation is dependent on the phase of the cell cycle, rather than on genotypic heterogeneity. The switch from the G1 to the S phase mode of response is driven by active coupling between the PTH receptor and a Ca²⁺ channel through a pertussis toxin-sensitive G protein.