(-)-[125I]-iodopindolol, a new highly selective radioiodinated beta-adrenergic receptor antagonist: measurement of beta-receptors on intact rat astrocytoma cells

Abstract

(-)-Pindolol, one of the most potent beta-adrenergic receptor antagonists, was radioiodinated using chloramine-T oxidation of carrier-free Na 125I and separated from unreacted pindolol to yield 2200 Ci/mmole (-)-[125I]-iodopindolol ((-)-[125I]-IPin). Mass and ultraviolet spectra confirmed that the iodination occurred on the indole ring, presumably at the 3 position. The binding of radiolabeled (-)-[125I]-IPin to beta-adrenergic receptors has been studied using intact C6 rat astrocytoma cells (2B subclone) grown in monolayer cultures. Binding of (-)-[125I]-IPin was saturable with time and concentration. Using 13 pM (-)-[125I]-IPin, binding equilibrium was reached in 90 min at 21-22 degrees C. The reverse rate constant was 0.026 min-1 at 21 degrees C. Specific binding (expressed as 1 microM (-)-propranolol displaceable counts) of (-)-[125I]-IPin was 95% of total binding. Scatchard analysis of (-)-[125I]-IPin binding revealed approximately 4300 receptors/cell and a dissociation constant of 30 pM. This was in excellent agreement with the kinetically determined dissociation constant of 35 pM. Displacement by propranolol and isoproterenol showed that (-)-[125I]-IPin binding sites were pharmacologically and stereospecifically selective. These results indicate that (-)-[125I]-IPin, a pure (-)-stereoisomer, high specific activity radioligand, selectively binds to beta-adrenergic receptors in whole cells with a high percentage of specific binding and should therefore be useful in the study and measurement of cellular beta-adrenergic receptors.