AbstractThe intestinal hormone and neuromodulator cholecystokinin (CCK) receptors CCK1R and CCK2R act as a signaling hub in brain–gut axis, mediating digestion, emotion, and memory regulation. CCK receptors exhibit distinct preferences for ligands in different posttranslational modification (PTM) states. CCK1R couples to G_s and G_q, whereas CCK2R primarily couples to G_q. Here we report the cryo-electron microscopy (cryo-EM) structures of CCK1R–G_s signaling complexes liganded either by sulfated cholecystokinin octapeptide (CCK-8) or a CCK1R-selective small-molecule SR146131, and CCK2R–G_q complexes stabilized by either sulfated CCK-8 or a CCK2R-selective ligand gastrin-17. Our structures reveal a location-conserved yet charge-distinct pocket discriminating the effects of ligand PTM states on receptor subtype preference, the unique pocket topology underlying selectivity of SR146131 and gastrin-17, the conformational changes in receptor activation, and key residues contributing to G protein subtype specificity, providing multiple structural templates for drug design targeting the brain–gut axis.