Project Summary Dopamine receptors are the primary therapeutic targets for a variety of neurological and psychiatric disorders, including schizophrenia, Parkinson’s disease, and many other movement disorders. The five subtypes of dopamine receptors (D1-D5) are members of the superfamily of G protein-coupled receptors. This proposal is focused on the D2 receptor and, in particular, on two novel DRD2 allelic variants that are linked to hyperkinetic movement disorders. An important distinction between the two variants is that subjects with the mutation M374R have more severe dysfunction with an earlier onset than those with the mutation I212F. The three aims will use a variety of approaches to compare and contrast the effects of the two mutations on D2 receptor function. Our overall goal is to determine which changes to receptor function are pathogenic, and which are benign or even potentially neuroprotective. D2-I212F exhibits increased constitutive activation of G proteins, increased sensitivity to low concentrations of agonist, decreased arrestin binding, slow kinetics for regulation of potassium channels, and inefficient expression. We will determine if the more severe clinical outcome caused by the M374R mutation is explained by qualitatively similar changes that differ only in magnitude, or if there are qualitative differences with I212F. In Aim 1 we will assess arrestin binding and G protein activation by the two novel variants, expanding our characterization to all D2 receptor-activated Gα subtypes and exploring the role of receptor phosphorylation. In Aim 2 we will carry out in silico studies of receptor activation. In Aim 3 we will create a knock-in mouse model of heterozygous expression of D2-M374R. We will evaluate the functional properties of D2-M374R in midbrain dopamine neurons by slice electrophysiology, using first viral expression of D2-M374R, and then the knock-in mice as they become available. We will also evaluate gait abnormalities in the knock-in mice, and quantify D2 receptor expression in midbrain and neostriatum. The aims proposed here will quantify the effect of these novel polymorphisms on the function and expression of the D2 receptor in cells and in mouse brain and should provide a biological explanation for manifestation of neurological dysfunction in humans with these variants.