Type II 5'-iodothyronine deiodinase (D2), produces triiodothyronine (T(3)) and is stimulated by cold exposure via norepinephrine (NE) release in brown adipose tissue. Cultured rat brown adipocytes require T(3) for the adrenergic stimulation of D2 activity. D2 mRNA expression in cultured brown adipocytes is undetectable with the use of basal conditions or NE without T(3). Full D2 expression is achieved using NE + T(3), especially after prolonged T(3) exposure. beta(3)-Adrenergic agonists mimic the NE action, whereas cAMP analogs do not. Prolonged exposure to T(3) alone increases D2 mRNA. High T(3) doses (500 nM) inhibit the adrenergic stimulation of D2 activity while increasing D2 mRNA. The effects obtained with NE + T(3) or T(3) alone are suppressed by actinomycin, but not by cycloheximide, which leads to accumulation of short D2 mRNA transcripts. Prolonged or short exposure to T(3) did not change D2 mRNA half-life, but T(3) seemed to elongate it. In conclusion, T(3) is an absolute requirement for the adrenergic stimulation of D2 mRNA in brown adipocytes. T(3) upregulates D2 mRNA, an effect that might involve stimulation of factors required for transcription or for stabilization of D2 mRNA.