10 R.G. Pertwee
Table 3.Kivalues of cannabinoid receptor antagonists/inverse agonists for the in vitro displacement of
[^3 H]CP55940 from CB 1 -andCB 2 -specific binding sites
Ligand CB 1 CB 2 Reference
Kivalue (nM) Kivalue (nM)CB 1 -selective antagonists/inverse agonists
NESS 0327 0.00035a 21 a Ruiu et al. 2003
SR141716A 11.8 13,200 Felder et al. 1998
11.8 973 Felder et al. 1995
12.3 702 Showalter et al. 1996
5.6 >1,000 Rinaldi-Carmona et al. 1994
1.98b >1,000b Rinaldi-Carmona et al. 1994
1.8a 514 a Ruiu et al. 2003
AM281 12 b 4,200a Lan et al. 1999a
AM251 (compound 12) 7.49b 2,290a Lan et al. 1999b
LY320135 141 14,900 Felder et al. 1998
CB 2 -selective antagonists/inverse agonists
AM 630 5,152 31.2 Ross et al. 1999a
SR144528 437 0.60 Rinaldi-Carmona et al. 1998
305 b 0.30b Rinaldi-Carmona et al. 1998
>10,000 5.6 Ross et al. 1999a
70 a 0.28a Ruiu et al. 2003
50.3 1.99 Iwamura et al. 2001See Figs. 10 and 11 for the structures of the compounds listed in this table.
aBinding to mouse brain (CB 1 ) or spleen tissue (CB 2 ).
bBinding to rat cannabinoid receptors on transfected cells or on brain (CB 1 ) or spleen tissue (CB 2 ).
All other data from experiments with human cannabinoid receptor.
(reviewed in Howlett et al. 2002; Pertwee 1997, 1999a). Both assays can be per-
formed with membranes obtained from brain tissue or from cultured cells that
express CB 1 or CB 2 receptors either naturally or after transfection. In addition, the
cyclic AMP assay can be performed with whole cells, including primary cultures of
central neurons, and the [^35 S]GTPγS assay can be used in autoradiography exper-
iments with tissue sections (Breivogel et al. 1997; Selley et al. 1996; Sim et al. 1995).
The cyclic AMP assay is more sensitive than the [^35 S]GTPγS assay. Presumably
this is because modulation of cyclic AMP production takes place further along the
signalling cascade than [^35 S]GTPγS binding so that there is greater signal amplifi-
cation. For the [^35 S]GTPγS assay, it is important to include guanosine diphosphate
(GDP) and sodium chloride at appropriate concentrations (Breivogel et al. 1998;
Selley et al. 1996; Sim et al. 1995). GDP increases the ratio of agonist-stimulated
to basal [^35 S]GTPγS binding (signal-to-noise ratio) but also decreases the abso-
lute levels of both agonist-stimulated and basal [^35 S]GTPγS binding. In addition,
it magnifies the differences in efficacy exhibited in this assay by full and partial
agonists (Savinainen et al. 2001). The signal-to-noise ratio in this bioassay can be
further improved by including an adenosine A 1 receptor antagonist (Savinainen