The study investigates the structural mechanisms underlying the loading of monoamine neurotransmitters, such as serotonin and dopamine, into synaptic vesicles by vesicular monoamine transporter 2 (VMAT2). VMAT2 is crucial for monoaminergic signaling and its dysfunction is linked to various neuropsychiatric disorders. The research employs cryo-electron microscopy (cryo-EM) to capture the structures of human apo VMAT2 and its complexes with serotonin, tetrabenazine (TBZ), and reserpine (RES). These structures reveal three distinct conformations: lumen-facing, occluded, and cytosol-facing. Notably, TBZ induces significant rearrangements in TM2 and TM7, extending beyond typical rocker-switch movements. These findings provide insights into ligand recognition and the proton-driven exchange cycle, which are essential for understanding VMAT2's function and designing improved pharmaceuticals targeting this transporter.The study investigates the structural mechanisms underlying the loading of monoamine neurotransmitters, such as serotonin and dopamine, into synaptic vesicles by vesicular monoamine transporter 2 (VMAT2). VMAT2 is crucial for monoaminergic signaling and its dysfunction is linked to various neuropsychiatric disorders. The research employs cryo-electron microscopy (cryo-EM) to capture the structures of human apo VMAT2 and its complexes with serotonin, tetrabenazine (TBZ), and reserpine (RES). These structures reveal three distinct conformations: lumen-facing, occluded, and cytosol-facing. Notably, TBZ induces significant rearrangements in TM2 and TM7, extending beyond typical rocker-switch movements. These findings provide insights into ligand recognition and the proton-driven exchange cycle, which are essential for understanding VMAT2's function and designing improved pharmaceuticals targeting this transporter.