Direct labelling of β2-adrenoceptors
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A radioligand that selectively labels β2-adrenoceptors, 3H-ICI 118,551 (3H-ICI), is introduced. Experiments were performed on guinea-pig tissues. The binding characteristics of 3H-ICI on lung membrane particles are compared with the blocking characteristics of ICI 118,551 against the tracheo-relaxant effects of (−)-noradrenaline, (−)-adrenaline and (±)-fenoterol. Binding to both β1- and β2-adrenoceptors were also performed with 3H-(−)-bupranolol on lung and ventricular myocardium. The binding inhibition characteristics of unlabelled ICI 118,551 on ventricle were compared with its characteristics as antagonist of the positive chronotropic effects of (−)-noradrenaline, (−)-adrenaline and (±)-fenoterol in spontaneously beating right atria.
ICI 118,551 blocked more the relaxant effects of (±)-fenoterol and (−)-adrenaline than those of (−)-noradrenaline on trachea. The positive chronotropic effects of (±)-fenoterol on sinoatrial node were blocked more than those of both (−)-adrenaline and (−)-noradrenaline. A non-linear regression analysis of blocking data with ICI 118,551 according to the model of Lemoine and Kaumann (1983) revelas that both β1- and β2-adrenoceptors contribute to the tracheo-relaxant and positive chronotropic effects of agonists. The estimated equilibrium dissociation constants pKB (-log KB=pKB; mol/l) were 7.1 and 9.6 for β1- and β2-adrenoceptors, respectively. Tracheal β2-adrenoceptors contribute 99%, 97% and 7%, sinoatrial β2-adrenoceptors contribute 76%, 3% and 0% to the fractional stimuli induced by (±)-fenoterol, (−)-adrenaline and (−)-noradrenaline, respectively.
3H-ICI associated to β2-adrenoceptors of lung membranes with a kon of 0.52 l·nmol−1·min−1 and dissociated with a koff of 0.19 min−1. 3H-ICI bound to lung β2-adrenoceptors with an equilibrium dissociation constant pKL* of 9.2. Unlabelled ICI 118,551, (−)-bupranolol, (+)-bupranolol, (−)-adrenaline, (−)-noradrenaline and (±)-fenoterol competed with 3H-ICI for lung β2-adrenoceptors with pKL-values of 9.0, 9.4, 8.1, 5.9, 4.9 and 6.4, respectively.
3H-(−)-bupranolol associated to β-adrenoceptors of lung membranes with a kon 1.2 l·nmol−1·min−1 and dissociated with a koff of 0.26 min−1. 3H-(−)-bupranolol bound to lung β2-adrenoceptors and to heart β1-adrenoceptors with a pKL of 9.6 and a pKL of 8.8, respectively. Lung β2- and β1-adrenoceptors comprised 3/4 and 1/4 of the β-adrenoceptor population, as estimated independently with 3H-ICI and 3H-(−)-bupranolol; 1/5 of ventricular β-adrenoceptors was β2, 4/5 β1.
The binding characteristics including stereoselectivity show that 3H-(−)-ICI 118,551 is useful to label nearly exclusively β2-adrenoceptors in a system containing both β1- and β2-adrenoceptors. The affinity for β2-adrenoceptors of competing ligands can be determined straightforwardly without interference of β1-adrenoceptors. The low affinity for lung β2-adrenoceptors but high tracheorelaxant potency of agonists suggest the existence of a large β1-adrenoceptor reserve.
Key wordsLung and heart β1- and β2-adrenoceptors 3H-ICI 118,551 and 3H-(−)-bupranolol (−)-Adrenaline and (−)-noradrenaline Binding and blockade
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