Characteristics of the beta-adrenergic adenylate cyclase system of developing rabbit bone-marrow erythroblasts.
After fractionation of rabbit bone marrow into dividing (early) and non-dividing (late) erythroid cells, the adenylate cyclase activity of membrane ghosts was assayed in the presence of guanine nucleotides ((GTP and its analogue p[NH]ppG (guanosine 5'-[beta, gamma-imido]triphosphate))), the beta-adrenergic agonist L-isoprenaline (L-isoproterenol) and the antagonist L-propranolol. Both GTP and p[NH]ppG increased the adenylate cyclase activity of early and late erythroblasts, whereas the stimulating effect of the beta-adrenergic drug L-isoprenaline was limited to the immature dividing bone-marrow cells. The effect of L-isoprenaline was completely inhibited by the antagonist L-propranolol, confirming that the response was due to stimulation of beta-adrenergic receptors on the plasma membrane. The lack of response of non-dividing erythroblasts to beta-adrenergic stimuli is not due to loss of beta-receptors, since both dividing and non-dividing cells bind the selective ligand [125I]iodohydroxybenzylpindolol with almost equal affinities, the apparent dissociation constants, Kd, being 0.91 X 10(-8)M and 1.0 X 10(-8) M respectively. The number of beta-adrenergic receptors per cell was 2-fold higher in the dividing cells. No significant change in binding affinity for GTP and p[NH]ppG during erythroblast development was observed: the dissociation constants of both guanine nucleotides were almost identical with early and late erythroblast membrane preparations [2-3 (X 10(-7) M]. With dividing cells, however, in the presence of L-isoprenaline the dissociation constants of GTP and p[NH]ppG were lower (6 X 10(-8) M). The dose-response curves for isoprenaline competition in binding of [125I]iodohydroxybenzylpindolol by dividing cells showed that the EC50 (effective concentration for half maximum activity) value for isoprenaline was higher in the presence of p[NH]ppG. With non-dividing cells the EC50 value for isoprenaline was equal in the presence and in the absence of p[NH]ppG and similar to that observed with dividing-cell membranes in the presence of the nucleotide. Thus differentiation of rabbit bone-marrow erythroid cells seems to be accompanied by uncoupling of the beta-adrenergic receptors from the adenylate cyclase catalytic protein as well as by a decrease in the number of receptors per cell, but not by changes in the catecholamine and guanine-nucleotide-binding affinities.
D000262:Adenylyl Cyclases / Q000097:blood* / Q000378:metabolism*; D000818:Animals; D001853:Bone Marrow / Q000201:enzymology*; D002454:Cell Differentiation; D004305:Dose-Response Relationship, Drug; D004900:Erythroblasts / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D004912:Erythrocytes / Q000201:enzymology*; D006160:Guanosine Triphosphate / Q000494:pharmacology; D006165:Guanylyl Imidodiphosphate; D066298:In Vitro Techniques; D007545:Isoproterenol / Q000494:pharmacology; D007700:Kinetics; D008297:Male; D010869:Pindolol / Q000031:analogs & derivatives / Q000378:metabolism; D011817:Rabbits
D006165:Guanylyl Imidodiphosphate; C011878:iodohydroxybenzylpindolol; D006160:Guanosine Triphosphate; D010869:Pindolol; D000262:Adenylyl Cyclases; D007545:Isoproterenol