Pharmacological characterization of [3H]idazoxan, [3H]RX821002 and p-[125I]iodoclonidine binding to alpha 2-adrenoceptors in rat cerebral cortical membranes

Binding characteristics of alpha 2-adrenoceptors in rat cerebral cortical membranes were compared using the antagonist radioligands 3H]idazoxan, 3H]2-(2-methoxy-1,4-benzodioxan-2-yl)-2-imidazoline (3H]RX821002), and the partial agonist radioligand 125I]2-2,6-(dichloro-4-iodophenyl)imino]imidazoline (125I]iodoclonidine). With 3H]RX821002 and alpha 2-adrenoceptor subtype-selective competitors, both alpha 2A/D- and alpha 2C-adrenoceptor subtypes were detected, suggesting rat cortical membranes contain approximately 90% alpha 2A/D-adrenoceptor subtype and 10% alpha 2C-adrenoceptor subtype. Only alpha 2A/D-adrenoceptors were detected with 3H]idazoxan and 125I]iodoclonidine. All three radioligands bound to a single high affinity site (Kd = 0.3-1.6 nM). However, the densities of sites labeled by 3H]idazoxan and 125I]iodoclonidine were 50% greater than the density labeled by 3H]RX821002, likely representing non-adrenoceptor binding sites. The density of 125I]iodoclonidine binding sites in glycylglycine buffer was similar to that labeled by 3H]RX821002. These results suggest that: (1) alpha 2A/D-adrenoceptors are the predominant subtype in rat cerebral cortex, (2) demonstrate that the small number of alpha 2C-adrenoceptors in this tissue can be detected using prazosin to displace 3H]RX821002 binding, and (3) non-adrenoceptor binding with 125I]iodoclonidine can be minimized with the use of glycylglycine buffer.