11C-NMSP PET studies were taken 40 min after a bolus i.v. injection of approximately 20 mCi 11C-NMSP. To create the normal 11C-NMSP SPM image template, additional dynamic scans were obtained at 0–60 min after injection of 11C-NMSP under the resting state in healthy controls.

HEK293 cells stably expressing human dopamine D2 and D3 receptors were used for activity evaluation. Immediately prior to testing, N-Methylspiperone was freshly dissolved in 30% DMSO and 70% H2O to a stock concentration of 100 μM and diluted into 13 half-log serial dilutions using 30% DMSO vehicle. Final test concentrations ranged from 10 μM to 10 pM. Radioligand competition experiments were conducted in glass tubes containing 300 μL of fresh EBSS buffer with 0.2 mM sodium metabisulfite, 50 μL of diluted test compound, 100 μL of membranes (10 μg total protein). All compound dilutions were tested in triplicate and the reaction incubated for 1 h at room temperature. The reaction was terminated by filtration through Whatman GF/B filters, presoaked for 1 h in 0.5% polyethylenimine, using a Brandel R48 filtering manifold (Brandel Instruments, Gaithersburg, MD).