Parathyroid hormone (PTH) stimulates osteoclast formation by binding to its receptor on stromal/osteoblastic cells and stimulating the production of receptor activator of NFkappaB ligand (RANKL) and inhibiting the expression of osteoprotegerin (OPG). However, the mechanisms through which PTH regulates these genes remain unknown. Here we report that PTH stimulated RANKL gene transcription and increased RANKL mRNA stability in murine stromal/osteoblastic cells stably expressing human PTH/PTH-related protein receptor 1. PTH also potently suppressed OPG mRNA in these cells. Cycloheximide did not block the effects of PTH on RANKL but did inhibit the suppression of OPG mRNA. Activation of protein kinase A (PKA) was necessary and sufficient for the effect of PTH on both genes. Conditional expression of a dominant-negative form of the transcription factor CREB, but not c-fos or Runx2, significantly reduced PTH stimulation of RANKL. CREB activity was also required for full stimulation of RANKL by oncostatin M or 1,25-dihydroxyvitamin D(3). Dominant-negative forms of CREB and c-fos reduced the suppression of OPG by PTH. These results demonstrate that PTH directly stimulates RANKL expression via a PKA-CREB pathway and that CREB may be a central regulator of RANKL expression. Furthermore, they suggest that PTH suppression of OPG involves CREB and c-fos.