Radioligand Binding Detection of Receptors in Brain Membranes

Abstract

Transmitters, hormones, and most of the therapeutic drugs exert their biological functions by generating signals in target cells through interaction with a receptor molecule in the cellular membrane, cytosol, or even at the nucleus of the cell. Because receptors play such a crucial role in cell function, especially within the central nervous system (CNS), we need lab techniques to measure their expression and functionality. There are techniques to determine regulatory changes in receptor number (receptor density), subcellular distribution, anatomical distribution, the physiological function of the receptors as well as the ligand affinity.

Here, we describe a classic method in which cells or tissue membranes are exposed to radiolabeled molecules (radioligands) that bind selectively to the receptors in the sample (receptor binding assay). Similarly, the functional consequence of the receptor occupancy or level of G protein activation can be measured by [³⁵S]GTPγS binding assay, also known as functional binding. This assay measures the binding of the nonhydrolyzable analog [³⁵S]GTPγS ([³⁵S]guanosine-5′-O-(3-thio) triphosphate) to Gα subunits.

The above radiometric assays are useful to determine the affinity and receptor density of a drug, and to differentiate among agonist, antagonist, and inverse agonist activities, the traditional pharmacological parameters of potency and efficacy, as well as receptor constitutive activity. Moreover, radioligand binding assays provide a powerful tool for screening drug candidates for many receptors in drug discovery research.