Table 1.
Receptor and SERT occupancy assay conditions for each target
| 5-HT3 | 5-HT7 | 5-HT1B | 5-HT1A | SERT | |
|---|---|---|---|---|---|
| Preincubation buffer | 50 mM Tris HCl 150 mM NaCl | 50 mM Tris HCl 4 mM CaCl2 0.5 μM L-ascorbic acid | 170 mM Tris HCl 4 mM CaCl2 5.67 mM L-ascorbic acid | None | None |
| Preincubation time | 1 × 5 min | 1 × 3 min | 1 × 3 min | None | None |
| Assay buffer | 50 mM Tris HCl 150 mM NaCl 4 mM CaCl2 | 170 mM Tris HCl 4 mM CaCl2 0.5 uM L-ascorbic acid 100 μM pargyline | 170 mM Tris HCl 4 mM CaCl2 5.67 mM L-ascorbic acid 10 μM pargyline | 170 mM Tris HCl 4 mM CaCl2 5.67 mM L-ascorbic acid 10 μM pargyline | 50 mM Tris HCl 150 mM NaCl 5 mM KCl |
| Radioligand | [3H]LY278584 2 nM | [3H]SB269970a 5.9 nM | [3H]GR125743 1 nM | [3H]8-OH-DPATb 3 nM | [3H]escitalopram 4.5 nM |
| Non-specific binding agent | ondansetron 10 μM | SB269970 1 μM | SB216641 1 μM | WAY-100635 1 uM | Paroxetine 1 uM |
| Incubation time | 1 h | 1 h | 1 h | 1 h | 1 h |
| Region of Interest | amygdala, piriform cortex | Anterior paraventricular thalamic nucleus interanteromedial thalamic nucleus, other medial thalamic regions | Caudate, putamen, nucleus accumbens | Hippocampus | Lateral septum, medial septum, olfactory tubercle |
Specific details of each assay for each target are provided. See also du Jardin et al., 2013 and Pehrson et al., 2013.
a
[3H]SB269970 was supplied by Amersham (Piscataway, NJ, USA), and has a specific activity of 33 Ci/mmol and radioactivity concentration of 0.2 mCi/mL. Given a counting efficiency of 44%, the cpm/mL equivalent of 5.9 nM [3H]SB269970 is approximately 190 000.
b
[3H]8-OH-DPAT was supplied by Perkin Elmer (Waltham, MA, USA) and has a specific activity of 154.2 Ci/mmol and radioactivity concentration of 1.0 mCi/mL. Given a counting efficiency of 44%, the cpm/mL equivalent of 3 nM [3H]8-OH-DPAT is approximately 452 000.