ABSTRACT: We have identified a conserved threonine residue in the second intracellular (i2) loop of the 5-HT1A receptor that when mutated to alanine prevents coupling to Gβγ-mediated signaling, while preserving Gαi-induced actions. 48 In this review, we investigate the characteristics and potential role of the i2 domain in the coupling of the 5-HT1A receptor and other receptors to G proteins. The i2 domain, as well as portions of the i3 domain, is predicted to form an amphipathic α-helix with a positively charged face and a hydrophobic face. Mutagenesis experiments support a model in which the hydrophobic faces of these α-helical domains form an intracellular binding “pocket” for interaction with G proteins. Embedded in the hydrophobic face, Thr149 is crucial for signaling through Gβγ subunits, perhaps via interaction with its hydroxyl side-chain. Mutation of other residues of the i2 domain of Gi-coupled receptors is required to substantiate the importance of the α-helical i2 domain in receptor-Gβγ signaling. If confirmed in other receptors, these results support a general model in which activated receptor and Gβγ subunits remain associated to interact with effectors in a receptor-specific manner.