The present study was undertaken to determine the effects of catecholamines, agonists, and antagonists of β-adrenergic receptors (AR) in the LNCaP cell line. Changes in cellular cyclic adenosine-3′,5′-monophosphate (cAMP) levels were quantified by the use of a 6 cAMP response element (CRE)-luciferase reporter gene assay. LNCaP cells were transiently transfected with this gene construct, incubated in 96-well microtiter plates for 24hr, and then treated with β-AR agonists and/or antagonists for 4hr. The rank order of potency for catecholamines and known β-AR agonists was terbutaline (3.31 nM )> isoproterenol (8.31 nM )≥ fenoterol (15 nM )= epinephrine (16.2 nM )> norepinephrine (77.5 nM )> BRL -37344 [(R^∗,R^∗)-(±)4-[2-[(2-(3-chlorophenyl)-2-hydroxyethyl)amino]propyl]phenoxy acetic acid, sodium salt] (1000 nM )> dobutamine (1770 nM )> CGP 12177 (4-[3-[(1,1-dimethylethyl)amino]-2-hydroxypropoxy]-1,3-dihydro-2H-benzimidazole-2-one hydrochloride) (inactive). The non-selective β₁-/-β₂-AR antagonists; propranolol and CGP 12177, at 10⁻⁷M, inhibited luciferase activity induced by these agonists by 80–96%. Propranolol blocked isoproterenol-induced luciferase responses in a competitive manner (K_B=1.4nM). In addition, isoproterenol-activated luciferase expression was blocked more potently by ICI 118,551 [(±)-1-[2,3-(dihydro-7-methyl-1H-inden-4-yl)oxy]-3-[(1-methylethy) amino]-2-butanol], a β₂-AR antagonist than by ICI 89,406 [(±)-N-[2-[3-(2-cyanophenoxy-)]-2-hydroxypropylamino]ethyl-N-phenylurea], a β₁-AR antagonist, giving K_B values of 1.07 and 161nM, respectively. These results suggest that the β₂-AR is the major subtype mediating catecholamine-induced cAMP changes in LNCaP cells.