S-PHASE INDEPENDENCE OF PARATHYROID HORMONE-INDUCED CALCIUM SIGNALINGIN PRIMARY OSTEOBLAST-LIKE CELLS

Cell cycle-dependence of parathyroid hormone (PTH)-induced signal tran sduction was studied at the single cell level in fetal rat osteoblast- like cells (ROB) in primary culture. Responsiveness to 10(-7) M rPTH1- 34 was measured as changes in the intracellular calcium concentration ([Ca2+](i)) of cells loaded with Fura-2 using a video imaging setup. C ells in S-phase were identified by staining for incorporated 5-bromo-2 '-deoxyuridine (BrdU). ROB were cultured on glass coverslips which had alphanumerically marked gratings so that cells could be located. Dire ctly after measurement of the PTH-induced [Ca2+](i) response of the in dividual cells, cells were fixed and stained for incorporated BrdU. Ph ase-contrast images taken with the video imaging setup were compared w ith phase-contrast micrographs and fluoro-micrographs taken after stai ning for BrdU incorporation. We found that 43% of the cells responding to PTH with a rise in [Ca2+](i) had also incorporated BrdU. This perc entage was not different from the percentage BrdU positive cells in th e whole culture, showing that calcium responsiveness was randomly dist ributed between ROB in S-phase and those not in S-phase. We therefore conclude that PTH does not induce calcium responses preferentially dur ing the S-phase of the cell cycle in ROB in primary culture.