Structural basis of LY334370 recognition and selectivity at the 5-HT_1F receptor

The 5-HT_1F receptor is a serotonin receptor subtype highly expressed in trigeminal sensory neurons, where it modulates neuropeptide release and nociceptive signaling without inducing vasoconstriction. This makes it an important therapeutic target for migraine. LY334370 was developed as a first-generation selective 5-HT_1FR agonist and demonstrated efficacy in clinical studies. However, the molecular mechanism underlying 5-HT_1FR activation by LY334370 remains poorly understood. Here, we determined a 3.13 Å cryo-EM structure of the LY334370-bound 5-HT_1FR-miniGα_oA complex. Combined with functional analyses, this structure delineates the molecular determinants underlying LY334370 recognition. Comparison with BRL54443 indicates that LY334370 selectivity for 5-HT_1FR is driven by its optimal accommodation within the receptor-specific extended binding pocket. Furthermore, comparative analysis with the lasmiditan-bound 5-HT_1FR-Gα_i1 complex reveals distinct agonist binding modes and provides mechanistic insight into Gα_i/o subtype-specific coupling. Collectively, these findings elucidate the structural basis of 5-HT_1FR activation, ligand selectivity, and G protein coupling, providing a structural framework for the rational design of safer and more effective anti-migraine drugs.