Classification of β-adrenergic subtypes in immature rabbit bone marrow erythroblasts

Abstract The beta-adrenergic receptors of immature rabbit bone marrow erythroid cells (proerythroblasts and basophilic erythroblasts) were identified. [ 125 I]iodocyanopindolol bound to membrane preparations derived from these erythroblasts in a rapid, reversible and saturable manner. Scatchard analysis of binding data revealed a single class of binding sites (Hill coefficient of 0.954) with an apparent equilibrium dissociation constant ( K d ) of 8 pM, and a density of binding sites ( B max ) of 1.53 pM/10 6 cells, corresponding to 920 receptors per cell. The binding of [ 125 I]iodocyanopindolol was inhibited stereospecifically by concentrations of (−)-propranolol 2 orders of magnitude lower than by the (+)-isomer. Only l -isoprenaline and l -adrenaline activated the adenylate cyclase of immature rabbit erythroblasts, while l -noradrenaline, a β 1 -adrenergic agonist, was inactive. The order of potency of different agonists for displacement of bound [ 125 I]iodocyanopindolol was: isoprenaline > adrenaline > noradrenaline with respective EC 50 (concentration required for half maximal inhibition of binding) of 7.9 × 10 −7 M, 1.5 × 10 -5 M and 7.9 × 10 −5 M. This agonist potency series did not change with differentiation of rabbit bone marrow erythroblasts. The inhibition of specific [ 125 I]iodocyanopindolol binding to immature cells by β 1 - and β 2 -selective drugs (noradrenaline, practolol, procaterol and butoxamine) resulted in linear Hofstee plots. The inhibition curves obtained with procaterol and butoxamine, with apparent K d values of 3.1 × 10 −9 M and 4.9 × M, respectively, provide further evidence that the high-affinity binding sites correspond to a homogenous β 2 -receptor subtype.