Ligand | Target receptor | % Control | Test Conditions |
---|---|---|---|
Binding to glutamate receptors other than AMPA receptors | |||
4 nM [3H]kainate | High-affinity kainate | 99.0 ± 2.2 | 5 mM CX516, 30 min, 25°C |
40 nM [3H]kainate | Low-affinity kainate | 99.6 ± 1.5 | 5 mM CX516, 30 min, 25°C |
10 nM [3H]MK-801 | N-Methyl-d-aspartate | 93.5 ± 2.7 | 2 mM CX516, 5 min, 25°C |
Glutamate transport into synaptoneurosomes | |||
25 nM [3H]glutamate | 99.0 ± 1.6 | 2 mM CX516, 10 min, 25°C |
Rat brain membranes in HEPES/Tris buffer were incubated with the radioligand and under the assay conditions indicated in the table. Incubations were terminated by filtration through GF/C filters after addition of 4 ml of an ice-cold solution of 100 mM NaCl, 10 mM Tris/HCl, pH 7.4. Incubations with [3H]MK-801 contained 100 μM glutamate to facilitate access to the binding site in the channel domain. Synaptoneurosomes in ACSF buffer were prepared from rat cortex (see Materials and Methods) and used to measure [3H]glutamate sequestration within 4 h after preparation. This sequestration was completely eliminated by the detergent saponin (0.025%; not shown) and reduced by 90% by the transport inhibitor threo-hydroxy-aspartate (250 μM; not shown), which is indicative of glutamate uptake through sodium-coupled transport.