Rats were administered NNC-711 and scopolamine by the intraperitoneal route in a final volume of saline that corresponded to 1 ml/kg. For passive avoidance studies, NNC-711 was administered in a dose range of 0 - 3.0 mg/kg 30 minutes prior to training time, while scopolamine (0.8 mg/kg) was administered at the 6-hour post-training time. In water maze studies, NNC-711 was administered at a dose of 0.5 or 1.5 mg/kg 30 min prior to the first trial on each of the testing days but was not administered prior to the retention trial. NC-711 demonstrated a dose-dependent reversal of scopolamine-induced amnesia in passive avoidance training. In the water maze study administration of NNC-711 prior to the training sessions significantly improved performance.

Stable cell lines for human GAT-1 were generated in LM(tk-) cells using the calcium phosphate method and selection in G-418. Cells were grown under standard conditions (37°C, 5% CO 2) in DMEM. Cells were grown in 24-well plates and washed 3 times with HEPES buffered saline and allowed to equilibrate at 37 deg-C. After 10 min the medium was removed and unlabled NO-711 in HBS was added (45 microL/well). Transport was initiated by adding 50/micro-L per well of a concentrated solution of [3H]GABA in HBS (final concentration = 50 nM). Plates were incubated at 37°C for 10 min, then washed rapidly 3 x with ice-cold HBS. Cells were solubilized with 0.05% sodium deoxycholate/0.1 N NaOH (0.25 ml/well), an aliquot neutralized with 1 N HC1, and radioactivity was determined by scintillation counting. Protein was quantified in an aliquot of the solubilized cells using a BIO-RAD protein assay kit. NNC-711 was found to have an IC50 of 0.04 micro-M against Human GAT-1.