Osteopontin (OP) or bone sialoprotein is a recently characterized extracellular matrix protein which is abundant in bone and is produced by osteoblasts. Parathyroid hormone (PTH) is a potent calcitropic hormone which regulates osteoblastic function including the synthesis of extracellular matrix proteins. This study examines the effect of human PTH (hPTH-[1-34]) on the expression of this novel protein in rat osteoblast-like cells. hPTH(1-34) significantly decreased the amount of OP in culture media of the rat osteoblastic osteosarcoma cell line, ROS 17/2.8, detected by Western immunoblot analysis. hPTH(1-34) also suppressed the steady-state level of OP mRNA two- to threefold with an ED50 of approximately 3 X 10(-10) M. This inhibition was detectable at 24 h, reached its nadir at 48 h, and lasted at least up to 96 h. The hPTH(1-34) effects were mimicked by isobutylmethylxanthine, cholera toxin, 8-bromo-cAMP, forskolin, and isoproterenol. hPTH(1-34) suppressed by two- to threefold the rate of OP gene transcription, estimated by nuclear run-on assays. The suppression of OP mRNA levels by hPTH(1-34) was also seen when basal levels were increased by transforming growth factor type beta, or 1,25-dihydroxyvitamin D3, or were decreased by dexamethasone. A similar decrease in the steady-state level of OP mRNA by hPTH(1-34) was also observed in primary cultures of osteoblast-enriched cells from fetal rat calvaria. These findings indicate that hPTH(1-34) suppresses the production of the novel extracellular matrix protein, OP, in osteoblasts at least in part through transcriptional control.