[3H]-MK 912 binding delineates two α2-adrenoceptor subtypes in rat CNS one of which is identical with the cloned pA2d α2-adrenoceptor

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Abstract
1 Simultaneous computer modelling of control and guanfacine-masked [3H]-MK 912 saturation curves as well as guanfacine competition curves revealed that the drugs bound to two α2-adrenoceptor subtypes in the rat cerebral cortex with very different selectivities. These α2-adrenoceptor subtypes were designated α2A and α2C. The Kd value of [3H]-MK 912 for the α2A-subtype was 1.77 nm and for the α2C-subtype 0.075 nm; the receptor sites showing capacities 296 and 33 fmol mg−1 protein, respectively. The Kds of guanfacine were 19.9 and 344 nm, respectively. 2 Binding constants of 26 compounds for the two rat cerebral cortex α2-adrenoceptor subtypes were determined by simultaneous computer modelling of control and guanfacine-masked drug competition curves as well as plain guanfacine competition curves using [3H]-MK912 as labelled ligand (i.e. a ‘3-curve assay’). Of the tested drugs WB4101, corynanthine, rauwolscine, yohimbine, ARC 239 and prazosin were found to be clearly α2C-selective with selectivities ranging from 16 to 30 fold whereas guanfacine, oxymetazoline, BRL 44408 and BRL 41992 were found to be α2A-selective with selectivities ranging from 9 to 22 fold. 3 The Kds of compounds obtained for the cerebral cortex α2C-adrenoceptors showed an almost 1:1 correlation with the corresponding Kds for α2-adrenoceptors expressed by the pA2d-gene (the rat ‘α2-C4’ adrenoceptor) in CHO-cells. The cerebral cortex α2A-adrenoceptors did not correlate well with the pA2d α2-adrenoceptor Kds. 4 In the rat spinal cord [3H]-MK 912 bound to α2A- and α2C-adrenoceptor sites with similar affinities as in the cerebral cortex and with densities 172 and 7.4 fmol mg−1 protein, respectively. Drug affinities for some compounds showing major selectivity for α2A- and α2C-adrenoceptors were fully compatible with the notion that the spinal cord sites were α2A- and α2C-adrenoceptors.