Mechanisms of vesicular monoamine transporter 2 modulation by amphetamines and lobeline

Amphetamine-type stimulants (ATSs) disrupt the brain monoaminergic system and strongly stimulate motivation, movement, and addiction. Vesicular monoamine transporter 2 (VMAT2) plays a vital role in monoaminergic neurotransmission by sequestering monoamines in presynaptic vesicles. How ATS binds to and regulates VMAT2 remains elusive. Here, we report the cryo-EM structures of human VMAT2 in complex with four ATSs (AMPH, METH, MDMA, and MDA) and the inhibitor lobeline, revealing the molecular mechanisms underlying ATS recognition and modulation. ATSs mimic monoamines and occupy the substrate-binding site of VMAT2, indicating that ATSs function as substrates and thus redistribute the vesicular monoamine pools to the cytosolic pools. Lobeline adopts an unexpected dual-binding mode and locks VMAT2 in a lumen-facing conformation, inhibiting monoamine and ATS uptake. Our dopamine release assay revealed that lobeline regulates AMPH-induced dopamine release by modulating both VMAT2 and DAT. This study provides important mechanistic insights into VMAT2 modulation by amphetamines and lobeline.