The 25‐hydroxyvitamin D₃‐24‐hydroxylase mRNA is tightly and reciprocally regulated by 1,25‐dihydroxyvitamin D₃ (1,25(OH)₂D₃) and parathyroid hormone (PTH). The upregulation of the 24‐hydroxylase by 1,25(OH)₂D₃ is well established and occurs at the transcriptional level through two vitamin D response elements in the promoter of the gene. However, this induction is blocked by the protein synthesis inhibitor cycloheximide (CHX) indicating a protein component in the regulation pathway. CHX treatment reduced total vitamin D receptor (VDR) protein levels in cells, but reintroduction of VDR and/or retinoid X receptor protein into cells by transfection did not reduce the inhibition by CHX. This indicates that production of another transcription factor or mRNA‐stabilizing protein synthesized in response to 1,25(OH)₂D₃ is required for optimal accumulation of 24‐hydroxylase mRNA. PTH downregulates the 24‐hydroxylase mRNA by affecting its stability. The half‐life of 24‐hydroxylase mRNA is reduced 4.2‐fold in AOK‐B50 cells by PTH. Untranslated regions of the 24‐hydroxylase mRNA in reporter gene assays did not confer PTH responsiveness. Further analysis of the coding region of the rat 24‐hydroxylase may reveal sites of action of PTH. J. Cell. Biochem. 88: 234–237, 2003. © 2002 Wiley‐Liss, Inc.