The study investigates the molecular mechanism of antihistamines' recognition and regulation of the histamine H₁ receptor (H₁R) to enhance the design of new antihistamines. The researchers determined the cryo-EM structures of H₁R in its apo form and bound to different antihistamines, identifying a secondary ligand-binding site in addition to the deep hydrophobic cavity. This secondary site, composed of ECL2, TM2, TM3, and TM7, is decorated with polar residues and may support the introduction of new derivative groups. The structures show that antihistamines exert inverse regulation by inserting a shared phenyl group into the deep cavity and blocking the movement of the toggle switch residue W428. The study also explores the recognition mechanisms of specific antihistamines, such as mepyramine, astemizole, and desloratadine, revealing detailed interactions with H₁R. These findings enrich our understanding of GPCR modulation and facilitate the design of novel antihistamines.The study investigates the molecular mechanism of antihistamines' recognition and regulation of the histamine H₁ receptor (H₁R) to enhance the design of new antihistamines. The researchers determined the cryo-EM structures of H₁R in its apo form and bound to different antihistamines, identifying a secondary ligand-binding site in addition to the deep hydrophobic cavity. This secondary site, composed of ECL2, TM2, TM3, and TM7, is decorated with polar residues and may support the introduction of new derivative groups. The structures show that antihistamines exert inverse regulation by inserting a shared phenyl group into the deep cavity and blocking the movement of the toggle switch residue W428. The study also explores the recognition mechanisms of specific antihistamines, such as mepyramine, astemizole, and desloratadine, revealing detailed interactions with H₁R. These findings enrich our understanding of GPCR modulation and facilitate the design of novel antihistamines.