New methods based on fluorescently labeled agonists, genetically encoded fluorescent sensors, and advanced microscopy techniques, such as fluorescence resonance energy transfer (FRET) and highly inclined thin illumination (HILO), allow direct monitoring of signaling, internalization, and intracellular trafficking of G protein-coupled receptors (GPCRs) and their ligands in living cells with high temporal and spatial resolution. These methods have been essential in revealing that GPCRs can continue signaling via production of the soluble second messenger cyclic AMP after internalization into the endosomal compartment.