Binding characteristics of [3H]guanfacine to rat brain alpha-adrenoceptors. Comparison with [3H]clonidine.

The tritium-labeled alpha-adrenoceptor agonist and antihypertensive drug guanfacine, N-amidino-2-(2,6-dichlorophenyl)-acetamide (sp. act. 24.2 Ci/mmole) was employed for a direct identification and characterization of alpha-adrenoceptors in rat brain membranes. Its usefulness as a radioligand was studied in comparison with [3H]clonidine (sp. act. 26.7 Ci/mmole). The nonspecific binding of [3H]guanfacine to rat cerebral membranes was considerably more pronounced than that observed for [3H]clonidine. The specific binding of [3H]guanfacine (0.1 - 20 nM) and [3H]clonidine (0.1 - 20 nM) as defined as the excess over blanks containing (-)-norepinephrine (10 microM) was saturable. Scatchard analyses of these binding data indicated single populations of binding sites for both ligands. KC values of 3.9 ([3H]guanfacine) and 3.7 nM ([3H]clonidine) were calculated. Maximal number of specific binding sites amounted to 220 and 195 fmole/mg protein for [3H]guanfacine and [3H]guanfacine and [3H]clonidine, respectively. In case unlabeled guanfacine (1 microM) was used to characterize the specific bonding of [3H] guanfacine, KD value and maximal number of binding sites were about twice as high as determined in the presence of excess (-)-norepinephrine. The rate of association of both radioligands was rapid. Binding reached equilibrium by about 10-15 min of incubation. Half-maximal binding was attained at approximately 1-2 min. The rates of dissociation were biphasic. A rapid and a slow component were identified. The specific binding sites of [3H] guanfacine in rat brain possess the general characteristics of alpha 2-adrenoceptors. Selective antagonists of alpha 2-adrenoceptors, like yohimbine and rauwolscine strongly interfered with this binding. However, preferential blocking agents of alpha 1-adrenoceptors, such as prazosin and corynanthine, were weak competitors. The relative potency of agonists and antagonists in displacing [3H]guanfacine was identical to their effectiveness in competing for [3H]clonidine specific binding sites. It is concluded that [3H]guanfacine labels the same alpha 2-adrenoceptor population in rat brain as [3H]clonidine. However, [3H]guanfacine seems not as suitable as [3H]clonidine for routine use in the direct identification of alpha 2-adrenoceptors in view of its relatively high nonspecific binding.