The effects of benactyzine (0.01-3.0 mg/kg) were examined on the frequency of alarm calls of squirrel monkeys in a laboratory setting. Under baseline conditions, few calls occurred, and neither saline nor a low dose (0.01 mg/kg) of benactyzine increased calling. Higher doses (0.03-3.0 mg/kg) of benactyzine significantly increased call rate (to 1-2 calls per s) in a dose-dependent manner. The rate-increasing effect of benactyzine on alarm calls appears to be related to a central antimuscarinic effect, as it could be partially blocked by 0.01 mg/kg physostigmine, completely blocked by 0.1 mg/kg physostigmine, but was not blocked at all by 0.1 mg/kg neostigmine. Neither of these cholinomimetics increased call rates when given alone. These findings show that benactyzine can increase alarm call rates in squirrel monkeys under defined laboratory conditions, and may serve as a useful pharmacological probe to study neurochemical mechanisms mediating the production of this type of vocalization. In the squirrel monkey, one such mechanism apparently involves a cholinergic substrate.