The orphan G protein-coupled receptor 6 (GPR6) is highly expressed in the striatum and has been linked to multiple striatal pathologies. The identification of endogenous ligands and their mechanisms of action at GPR6 will help to elucidate the physiological and pathological roles of the receptor. In the current study, we tested the concentration-dependent effects of a variety of endocannabinoid-like N-acylamides on GPR6 signaling. Here, we demonstrate for the first time that N-arachidonoyl dopamine, N-docosahexaenoyl dopamine, N-oleoyl dopamine and N-palmitoyl dopamine exert inverse agonism at GPR6. This effect was concentration-dependent, with potencies in the micromolar range, and functionally selective for beta-arrestin2 recruitment. Structure-activity relationship studies demonstrate that both the N-acyl side chain and the dopamine head group are important for these ligands to act on GPR6. Our discovery of these N-acyl dopamines as endogenous inverse agonists for GPR6 moves us one step further in understanding the roles GPR6 play in neurodegenerative and neuropsychiatric disorders related to striatal dysfunction. (C) 2019 Elsevier Inc. All rights reserved.