The serotonin (5-HT) system plays an important role in the pathophysiology and treatment of several major psychiatric disorders. Currently, no suitable positron emission tomography (PET) imaging paradigm is available to assess 5-HT release in the living human brain. [11C]AZ10419369 binds to 5-HT1B receptors and is one of the most 5-HT-sensitive radioligands available. This study applied 5-HT concentration enhancers which can be safely studied in humans, and examined their effect on [11C]AZ10419369 binding at clinically relevant doses, including amphetamine (1 mg/kg), 3,4-methylenedioxymethamphetamine (MDMA; 1 mg/kg) or 5-hydroxy-L-tryptophan (5-HTP; 5 mg/kg). Twenty-six PET measurements (14 for amphetamine, 6 for MDMA and 6 for 5-HTP) using a bolus and constant infusion protocol were performed in four cynomolgus monkeys before or after drug administration. Binding potential (BPND) values were determined with the equilibrium method (integral interval: 63-123 min) using cerebellum as the reference region. BPND values were significantly decreased in several examined brain regions after administration of amphetamine (range: 19-31%), MDMA (16-25%) or 5-HTP (13-31%). Reductions in [11C]AZ10419369 binding were greater in striatum than cortical regions after administration of 5-HTP, while no prominent regional differences were found for amphetamine and MDMA. In conclusion, [11C]AZ10419369 binding is sensitive to changes in 5-HT concentration induced by amphetamine, MDMA or 5-HTP. The robust changes in BPND, following pretreatment drugs administered at clinically relevant doses, indicate that the applied PET imaging paradigms hold promise to be successfully used in future human studies.