Multiple types of neurotransmitter binding sites in the rat neuroblastoma B 50 cell line. I. Characterization and...

In this study, it is shown that the neuroblastoma B 50 cell line of rat central nervous system origin possesses both adrenergic and cholinergic binding sites. The adrenergic binding site population comprises a major alpha 1-adrenergic component (using [3H]prazosin as ligand; KD = 0.55 nM; 19.3 fmol/mg protein), and a minor beta-adrenergic species (KD = 0.45 nM; 3.4 fmol/mg protein using [125I]cyanopindolol as ligand). The cholinergic binding site is of a muscarinic type. Binding studies with quinuclidinyl benzylate (QNB) indicate the presence of two binding sites with different affinities. This is supported by competition studies with agonists. However, in view of this unusual binding behavior of the antagonist, the possibility of a one-site model for QNB is also considered and affords the following parameters: KD = 0.247 nM, Bmax = 29.6 fmoles/mg protein. When the cells are induced to differentiate with dibutyryl cyclic AMP (db-cAMP) or high extracellular calcium, all of these binding sites undergo a variety of changes in their affinities for their respective ligands. Db-cAMP increases the affinity of the alpha 1-site for prazosin while high extracellular calcium lowers it. In both cases the number of binding sites remains unchanged. The affinity of the beta-adrenergic component is markedly enhanced upon induction of differentiation with either extracellular calcium or/and db-cAMP. Finally, the muscarinic cholinergic binding sites exhibit a decrease in receptor number upon db-cAMP treatment and an apparent loss of one of the binding sites upon calcium treatment. The significance of these affinity changes is discussed for each type of binding site.