Subclassification of beta-adrenergic receptors in cultured rat cardiac myoblasts and fibroblasts.

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Abstract
.beta.-Adrenergic receptors in primary cultures of neonatal rat cardiac cells were identified with the radioligand [125I]iodohydroxybenzylpindolol ([125I]IHYP). At the time of cell plating, a differential attachment procedure was employed to separate myocardial (M) cells from fibroblast-like (F) cells. After 3-4 days, the cultures enriched in M cells were still > 80% pure and the cultures enriched in F cells were > 95% pure. For binding studies, confluent cell layers were nonenzymatically detached as single cell suspensions. M and F cells contained a limited number of .beta.-adrenergic receptors (M, 7600 .+-. 2100 sites/cell; F, 9000 .+-. 2400 sites/cell) which had very high affinity (M, Kd = 88 .+-. 33 pM; F, Kd = 71 .+-. 23 pM) for [125I]IHYP. For each cell type, the binding sites were stereoselective for the l-isomers of agonists and antagonists. Further binding studies on the relative potency of .beta.-agonists showed that the .beta.-receptors in M cells could be subclassified as .beta.1 (isoproterenol > epinephrine .simeq. norepinephrine); the .beta.-receptors in F cells were more typical of .beta.2 (isoproterenol > epinephrine > norepinephrine). The order of potency of these catecholamines in stimulating the adenylate cyclase activity in M and F cells was consistent with the order of potency observed in binding studies. Practolol, a .beta.1 inhibitor, was 30 times more effective as a competitor of [125I]IHYP binding in M cells than in F cells. Whereas M and F cells share a similar number of receptors per cell and similar affinities for [125I]IHYP, the receptors in the 2 cell types can be distinguished on the basis of their subclassification. The importance of obtaining a homogeneous cell population for studies of the .beta.-adrenergic receptor in cultured cardiac tissue is emphasized.