Abstract The abused inhalant toluene has potent behavioral effects, but only recently has progress been made in understanding the molecular pathways that mediate the action of toluene in the brain. Toluene and ethanol induce similar behavioral effects and share some targets including NMDA and GABA receptors. In studies examining neuronal actions of ethanol, mice lacking the calcium-stimulated adenylyl cyclases (ACs), AC1 and AC8 (DKO), show increased sedation durations and impaired protein kinase A (PKA) phosphorylation following acute ethanol treatment. Therefore, using DKO mice, we compared the neurobehavioral responses following toluene exposure to that of ethanol exposure to determine if these abused substances share molecular mechanisms of action. In the present study, acute sensitivity to toluene- or ethanol-induced changes in locomotor activity was evaluated in DKO and wild type (WT) mice. Mice were exposed to toluene vapor (0, 500, 1000, 2000, 6000, or 8000ppm) for 30min in static exposure chambers equipped with activity monitors. Both WT and DKO mice demonstrated increased ambulatory distance during exposure to a 2000-ppm concentration of toluene compared to respective air-exposed (0ppm) controls. Significant increases in locomotor activity were also observed during an air-only recovery period following toluene exposure in WT and DKO mice that had been exposed to 2000ppm of toluene compared to respective air controls. Sedative effects of toluene were equivalent in WT and DKO mice, both during exposure and afterwards during recovery. Although no significant differences in locomotor activity were detected in DKO compared to WT mice at individual doses tested, a significant main effect of toluene was achieved, with DKO mice demonstrating a generalized reduction in locomotor activity during the post-toluene recovery period compared to WT mice (when analyzing all doses collectively). For comparison to toluene, additional WT and DKO mice were treated with 1.0 or 2.0g/kg ethanol (i.p.) and monitored for locomotor activation. In WT mice, both doses of ethanol increased distance traveled compared to saline controls. Conversely, DKO mice demonstrated no increase in locomotor activation at 1.0g/kg, with significantly reduced distances traveled at both doses compared to ethanol-treated WT mice. These behavioral activity results suggest that acute effects of ethanol and toluene are distinct in the mechanisms by which they induce acute sedating effects with respect to AC1 and AC8 activity, but may be similar in the mechanisms subserving locomotor stimulation.