Structural insights into the ligand binding and G(i) coupling of serotonin receptor 5-HT(5A)

5-hydroxytryptamine receptor 5A (5-HT(5A)) belongs to the 5-HT receptor family and signals through the G(i/o) protein. It is involved in nervous system regulation and an attractive target for the treatment of psychosis, depression, schizophrenia, and neuropathic pain. 5-HT(5A) is the only G(i/o)-coupled 5-HT receptor subtype lacking a high-resolution structure, which hampers the mechanistic understanding of ligand binding and G(i/o) coupling for 5-HT(5A). Here we report a cryo-electron microscopy structure of the 5-HT(5A)–G(i) complex bound to 5-Carboxamidotryptamine (5-CT). Combined with functional analysis, this structure reveals the 5-CT recognition mechanism and identifies the receptor residue at 6.55 as a determinant of the 5-CT selectivity for G(i/o)-coupled 5-HT receptors. In addition, 5-HT(5A) shows an overall conserved G(i) protein coupling mode compared with other G(i/o)-coupled 5-HT receptors. These findings provide comprehensive insights into the ligand binding and G protein coupling of G(i/o)-coupled 5-HT receptors and offer a template for the design of 5-HT(5A)-selective drugs.