Interaction of major monoterpenes from Rosmarinus officinalis L. essential oil with the human serotonin transporter: molecular docking, radioligand binding, and functional uptake studies

Major depressive disorder remains a leading cause of disability worldwide, and current antidepressants are limited by delayed therapeutic onset, incomplete efficacy, and adverse effects. Although Rosmarinus officinalis L. essential oil and its major monoterpenes exhibit antidepressant-like and anxiolytic properties, their molecular mechanisms remain unclear. This study investigated whether the principal rosemary monoterpenes, α-pinene, camphor, and 1,8-cineole, directly interact with the human serotonin transporter (hSERT) and its functional effects. Molecular docking simulations targeting the orthosteric (S1) and allosteric (S2) binding sites were combined with [ 3 H]-paroxetine binding displacement assays and SERT-mediated uptake experiments based on the fluorescent serotonin analog FFN246 in HEK293 cells expressing YFP-tagged hSERT. Docking analyses predicted plausible binding poses for all three monoterpenes within both the S1 and S2 regions, although with less favorable docking scores than fluoxetine. Radioligand displacement assays provided experimental evidence that all three compounds interact with hSERT, producing concentration-dependent displacement of [ 3 H]-paroxetine, with α-pinene displaying the highest potency, followed by 1,8-cineole and camphor. Functional assays further showed that all monoterpenes produced significant but incomplete inhibition of transporter-mediated FFN246 uptake, even at high micromolar concentrations, indicating partial rather than complete transporter blockade. Together, these findings identify hSERT as a previously unrecognized molecular target of rosemary monoterpenes, suggesting that these compounds may act as partial inhibitors of serotonin transport. These results provide new molecular insights into the potential mechanism that may contribute to the reported neurobehavioral effects of rosemary, and support further investigation of these compounds in serotonergic signaling and neurotransmission.