Activation of adenylyl cyclase by β-adrenergic receptors (βARs) plays a major role in adipose tissue homeostasis. The increase in cAMP promotes lipolysis in white adipose tissue, activates both thermogenesis and lipolysis in brown adipose tissue (BAT), and induces BAT hypertrophy. Previous studies indicated that among the three βAR subtypes present in adipose tissue, β3AR could be a potential target for antiobesity treatments in humans. We studied immortalized human brown adipocytes (PAZ6 adipocytes) as a model ofβ -adrenergic response in human BAT. PAZ6 adipocytes and freshly isolated mature human brown adipocytes display the same proportions ofβ AR subtypes, with β3AR being the most abundant (∼80% of the total). However, β3AR was poorly coupled to the adenylyl cyclase pathway in PAZ6 cells, contributing to only 10% of the isoproterenol-induced accumulation of cAMP, whereas 20% and 70% of the signal depended on β1- andβ 2-subtypes, respectively. Upon isoproterenol stimulation, β1- and β2AR down-regulated with a half-life of about 3 h and the β3AR with a half-life of 30–40 h. Long term stimulation with both saturating (micromolar) and nonsaturating (nanomolar) concentrations ofβ -adrenergic agonists caused a complete desensitization of theβ -adrenergic response at the adenylyl cyclase level and loss of stimulated protein kinase A activity and CREB phosphorylation. These results suggest that cAMP-dependent processes will be desensitized upon permanent treatment with β3AR agonists. Further studies should establish whether the β3AR is coupled to other signaling pathways in human brown adipocytes and whether these may contribute to BAT hypertrophy and/or thermogenesis.