ABSTRACTCocaine functions, in part, through agonist actions at sigma‐1 (σ1) receptors, while roles played by sigma‐2 (σ2) receptors are less established. Attempts to discriminate σ2receptor‐mediated effects of cocaine in locomotor hyperactivity assays have been hampered by the lack of potent and selective antagonists. Certain tetrahydroisoquinolinyl benzamides display high σ2receptor affinity, and excellent selectivity for binding to σ2over σ1receptors. The behavioral properties of this structural class of σ ligands have not yet been investigated. The present study evaluated 5‐bromo‐N‐[4‐(6,7‐dimethoxy‐3,4‐dihydro‐1H‐isoquinolin‐2‐yl)‐butyl)]‐2,3‐dimethoxy‐benzamide,1, a ligand shown by others to bind preferentially to σ2over σ1receptors, as well as dopamineD2andD3sites. First, we determined binding to monoamine transporters and opioid receptors, and noted 57‐fold selectivity for σ2receptors over the serotonin transporter, and >800‐fold selectivity for σ2receptors over the other sites tested. We then examined1in locomotor activity studies using male CD‐1® mice, and saw no alteration of basal activity at doses up to 31.6 µmol/kg. Cocaine produced a fivefold increase in locomotor activity, which was attenuated by 66% upon pretreatment of mice with1at 31.6 µmol/kg. In vivo radioligand binding studies also were performed, and showed no occupancy of σ1receptors or the dopamine transporter by1, or its possible metabolites, at the 31.6 µmol/kg dose. Thus, ligand1profiles behaviorally as a σ2receptor‐selective antagonist that is able to counteract cocaine's motor stimulatory effects.Synapse 68:73–84, 2014. © 2013 Wiley Periodicals, Inc.