The new radioligand [3H]-L 748,337 differentially labels human and rat β3-adrenoceptors

As no suitable radioligand exists for the detection of beta(3)-adrenoceptors, we have explored the radioligand binding properties of a tritiated version of the selective beta(3)-adrenoceptor antagonist L 748,337 Kinetic and equilibrium saturation and competition binding experiments were performed with [H-3]-L 748,337 on membrane fractions of HEK and CHO cells stably transfected with human and rat beta-adrenoceptor subtypes. Based on both association/dissociation kinetic and equilibrium saturation binding studies in transfected HEK cells, [H-3]-L 748,337 exhibited an affinity of approximately 2 nM for human beta(3)-adrenoceptors. Competition studies with agonists and subtype selective antagonists validated its binding to beta(3)-adrenoceptors. In CHO cells transfected with human beta(3)-adrenoceptors similar saturable high affinity of [H-3]-L 748,337 was observed. While some isoprenaline-sensitive [H-3]-L 748,337 binding was also observed in CHO cells transfected with human beta(1)- or beta(2)-adrenoceptors, this was not saturable in a similar concentration range and/or not sensitive to the antagonists propranolol and SR 59,230, indicating that it did not primarily involve beta-adrenoceptors. In CHO cells transfectecl with rat beta(3)-adrenoceptors [H-3]-L 748,337 exhibited a considerably lower affinity than with the human subtype (12-95 nM). Low affinity for the rat beta(3)-adrenoceptor was also found with unlabelled L 748,337 in rat bladder strip relaxation experiments. We conclude that L 748,337 apparently has lower affinity for the rat than the human beta(3)-adrenoceptors and that [H-3]-L 748,337 can bind to a low-affinity site distinct horn the orthosteric pocket of beta-adrenoceptors. Nevertheless, [H-3]-L 748,337 appears to be the most promising radioligand for the selective labelling of human beta(3)-adrenoceptors reported to date. (C) 2013 Elsevier EN. All rights reserved.