In drug reversal studies NGD94-1 was administered at 1 or 5 mg/kg intramuscularly 45 min prior to testing. Haloperidol was given at 0.025 mg/kg intramuscularly 30 min prior to testing

The dopamine D4 receptor is a Gprotein-coupled receptor and is negatively linked to adenylate cyclase, so that agonists at D4 receptors will inhibit forskolin-stimulated cAMP production. CHO cells stably expressing the cloned human D4.2 receptor were plated in 24-well plates 1 to 2 days before the assay, and were grown to confluence in Ham’s media with 10% FBS. On the day of the assay, each well was washed three times with serum-free Ham’s media containing 0.1 M HEPES. After a 30-min preincubation with NGD 94-1 and 50 mkM IBMX (phosphodiesterase inhibitor) at 37°C, cells were incubated with 5 mM forskolin for 15 min at 37°C. To stop the reaction, the plates were washed three times with cold PBS. Each well was incubated with 0.1 mM HCl for 20 min at room temperature. An aliquot of each sample was transferred to 12 * 75 mm polypropylene tubes, and the acid was neutralized with the addition of a solution containing 0.1 mM HEPES-0.1 mM K2CO3. The remaining acid was removed from the plates, and the cells were lysed with 0.5% Triton X-100. The protein content of each well was then determined with the Bio-Rad protein assay (Hercules, CA). The cAMP content in the neutralized extracts was determined with a cAMP RIA kit (NEN-DuPont, Boston, MA). The samples were quantitated using a gamma counter with an efficiency of 80 to 85%