Mechanisms of ligand-receptor interactions of the dopamine D2L receptor and their relation to functional selectivity

Binding and functional studies indicate that some agonists are capable of differentially activating pathways linked to a single receptor isoform. In cases where known mechanisms like other receptors or receptor reserve are ruled out, this phenomenon is referred to as “functional selectivity,” “agonist directed trafficking,” and a variety of other terms. Rather than viewing receptors in a traditional way as digital entities (i.e., one or more active or inactive states), functional selectivity posits that a receptor can be induced into unique conformations that may lead to different patterns of activation of functional responses linked to a single receptor. This dissertation elucidates the molecular mechanisms that explain why some ligands can differentially activate the effector pathways coupled to the human D2L receptor. The focus will be to discern the structural interactions of the hD2L receptor with such functionally selective ligands, and how these interactions result in selective activation of effector pathways. Computationally-selected point mutations of the D2L receptor were made, and detailed analysis of both binding and function made for a series of rationally selected ligands, emphasizing compounds that were relatively rigid and had fewer degrees of freedom in possible docking poses. Functional assays tested the intrinsic activity and potency for GTPγS binding, inhibition of forskolin-stimulated cAMP, phosphorylation of MAPK, and release of [3H]-arachidonic acid. Results from this dissertation provide evidence to support the hypothesis that ligand-selective interactions with specific residues of the receptor can induce conformational changes that lead to a characteristic pattern of activation of one of more signaling pathways. Developing a structure-based understanding of functional selectivity will provide general insight into the mechanisms of GPCR activation, and may also open the door to design of small molecules with desired patterns of functional effects at a single receptor