Mechanism of Dopamine 2-like Receptor Signaling in The Blood Brain Barrier that Regulates Male Courtship in Drosophila Melanogaster

In Drosophila melanogaster, courtship behavior and the brain circuits that mediate it are well characterized. Blood Brain Barrier (BBB) is an important part of the brain. It is purely glial in Drosophila and produces sex-specific modulators of male courtship behavior. One of the sex-specific BBB transcripts encodes a dopamine receptor, D2R, a G protein coupled receptor. D2R plays a major role in many central nervous system functions in both invertebrates and vertebrates. In Drosophila, we have previously found that D2R is preferentially expressed in the SPG cells of the adult male BBB and plays an important role in the regulation of male courtship. However, the molecular pathways downstream of D2R that modulate courtship behavior were unknown. D2R is a member of the family of G protein-coupled receptors (GPCRs) that can signal through two transducers: G proteins and β-arrestin. This study aims to characterize the role of G protein or arrestin signaling of D2R in the BBB for proper male courtship. The question was addressed by employing a genetic and a pharmacological approach. Two unique molecular genetic tools were generated by site-directed mutagenesis, a G protein-biased and a β-arrestin-biased version of D2R. These receptor versions, which can preferentially activate either the G protein or the β-arrestin downstream pathways were examined for their ability to rescue the courtship defects of D2R mutants. In a complementary pharmacological approach, well-known synthetic agonists, the G protein-biased agonist, MLS1547 and the β-arrestin-biased agonist, UNC9994 were administered to flies to examine their potential to rescue the courtship defects of D2R hypomorphic flies. The results from the genetic approach demonstrate that arrestin-mediated, but not G protein-mediated, signaling downstream of D2R is essential for normal male courtship. In agreement with this finding, the courtship defect of D2R hypomorphic flies was rescued by the administration of the arrestin-biased agonist, UNC9994. Together, these findings demonstrate that arrestin-mediated signaling through the D2R receptor in the BBB is required for normal male courtship behavior.