Table II.
Ligand Binding Parameters in Membranes and Reconstituted Receptors
| Receptors | State | Ligand | K (nM)a,b | Hill coefficient |
|---|---|---|---|---|
| Wild type–h5-HT3AR | Cell membranec | [3H]GR65630 | 0.52 ± 0.02a | 1.1 ± 0.1 |
| Cell membranec | 5-HT | 4100 ± 330b | 1.1 ± 0.1 | |
| FLAG-h5-HT3AR-1D4 | Cell membranec | [3H]GR65630 | 0.68 ± 0.04a | 1.2 ± 0.1 |
| Cell membranec | 5-HT | 2200 ± 130b | 0.9 ± 0.1 | |
| Reconstituted in lipid bilayersde | [3H]GR65630 | 1.8 ± 0.2a | 1.8 ± 0.3 | |
| Control in cell membranesd | [3H]GR65630 | 1.8 ± 0.1a | 1.8 ± 0.2 | |
| FLAG-hGABAAα1β3R | Reconstituteddf | GABA | 64.1 ± 32.8b | 0.8 ± 0.3 |
| Control in cell membranesd | GABA | 62.3 ± 17.4b | 0.5 ± 0.1 | |
| Control in cell membranesc | [3H]muscimol | 9.3 ± 2.4a | 1.3 ± 0.5 |
a
Kd (equilibrium dissociation constant of the ligand) determined by titrating [3H]ligand.
b
Ki for ligands determined by displacement of 0.15 nM [3H]GR65630 or 2 nM [3H]muscimol (13% and 11% receptor occupancy, respectively). Inhibition constant (Ki) was calculated according to the equation Ki = IC50/(1 + (ligand concentration/Kd ligand)). The half maximal inhibitory concentration (IC50) was determined by nonlinear least squares fitting to the Hill equation.
c
Filter assay.
d
Precipitation assay used for reconstituted receptors.
e
Bilayer composition PC:PA:Chol, molar ratio 67:12:21.
f
Reconstituted in 11.5 mM cholate, 0.86 mM asolectin.