Radioligand | α2-AR no. 1 | α2-AR no. 2 | α2-AR + Giα1 | α2-AR + Goα1 no. 1 | α2-AR + Goα1 no. 2 |
---|---|---|---|---|---|
[3H]RX821002 | |||||
K d(nM) | 1.3 ± 0.4 | 2.9 ± 0.6 | 2.1 ± 0.1 | 2.1 ± 0.1 | 1.7 ± 0.2 |
βmax(pmol/mg) | 6 ± 1.3 | 30 ± 4.3 | 31 ± 3.3 | 21 ± 1.7 | 11.3 ± 1.2 |
[3H]Clonidine | |||||
K d (nM) | 0.7 ± 0.2 | 0.7 ± 0.1 | 1.2 ± 0.1 | 0.5 ± 0.2 | 1.2 ± 0.01 |
βmax (pmol/mg) | 1.3 ± 0.5 | 2.1 ± 0.1 | 2.2 ± 0.2 | 2.5 ± 0.3 | 1.4 ± 0.2 |
[3H]UK14304 | |||||
K d (nM) | 0.7 ± 0.1 | 0.7 ± 0.2 | 0.7 ± 0.1 | 0.4 ± 0.2 | 0.7 ± 0.1 |
βmax (pmol/mg) | 3.2 ± 0.72-150 | 5.3 ± 1.32-150 | 7.0 ± 0.32-150 | 8.4 ± 0.72-150 | 4.3 ± 0.12-150 |
For each radioligand, parallel binding studies were conducted in the absence and presence of the nonhydrolyzable GTP analog Gpp(NH)p (100 μM). The binding of [3H]RX821002 was not influenced by the addition of Gpp(NH)p. In contrast, ∼80 to 85% of the specific binding of [3H]clonidine and [3H]UK14304 (at ligand concentrations of 2 nM) was Gpp(NH)p-sensitive. For both [3H]clonidine and [3H]UK14304, the specific binding obtained in the presence of Gpp(NH)p was subtracted from that obtained in the absence of Gpp(NH)p at each radioligand concentration to generate a saturation binding isotherm (see Fig. 4). The Gpp(NH)p-sensitive specific binding was evaluated by RADLIG to determine radioligand affinities and binding site densities. “No.” refers to independently isolated transfectants. Results are expressed as mean ± S.E. derived from two or three experiments with duplicate determinations.
↵2-150 p < .01 [3H]UK14304 versus [3H]clonidine (Student’s t test).