Interleukin-18 (IL-18) can regulate osteoblast and osteoclast function. We have identified, using cDNA microarray technology, that IL-18 expression is increased in UMR 106-01 rat osteoblastic cells in response to parathyroid hormone (PTH) treatment. Confirmation of these data using real-time reverse transcription-PCR showed that steady-state levels of IL-18 mRNA increased by 2 h (3-fold), peaked by 4 h (10-fold), and had diminished after 12 h (4.4-fold) and that this regulation was via the protein kinase A signaling pathway and did not involve activation of the PKC signal cascade. PTH regulation of IL-18 was confirmed at the protein level, and analysis of differentiating primary rat calvarial osteoblasts verified that both IL-18 mRNA and protein are regulated by PTH in primary rat osteoblasts. Promoter reporter assays revealed that PTH regulated the upstream IL-18 promoter and induced the exon 1 containing 1.1-kb IL-18 mRNA transcript in primary osteoblast cells. The in vivo physiological role of IL-18 in the anabolic actions of PTH on bone was then assessed using IL-18 knock-out mice. Female IL-18 null mice and wild-type littermate controls were injected with vehicle or 8 microg/100 g of human 1-38 PTH for 4 weeks. In IL-18 knock-out animals the anabolic effect of PTH (determined by bone mineral density changes in the proximal tibia) was abolished in trabecular bone but not in the cortical component. These data characterize the PTH regulation of IL-18 expression in osteoblastic cells and suggest that this cytokine is involved in the anabolic actions of PTH.