G protein regulatory network shapes magnitude and kinetics of behavioral responses in an engineered opioid receptor model

G protein-coupled receptors (GPCRs) control essential neuronal functions. One GPCR with prominent effects on the nervous system and animal behavior is the mu-opioid receptor (MOR). GPCRs mediate their effects by engaging a gamut of G proteins, which are inhibited by Regulators of G protein signaling (RGS). At present, how different RGS proteins regulate the magnitude and temporal kinetics of G protein signaling to affect behavior remains unclear. Here, we use an engineered cross-species Caenorhabditis elegans model of MOR signaling (tgMOR) to test how multiple RGS proteins shape MOR signaling and behavioral responses to opioids. Our results indicate opioid-induced effects on locomotor behavior in tgMOR are primarily mediated by Gαo and are modified by opposing Gαq action. We further delineate that EGL-10 (RGS7) is a primary RGS that modulates the magnitude of MOR-meditated responses. In a differential effect, EAT-16 (RGS9) and its regulator RSBP-1 (R7BP) principally influence the timing of behavioral response onset. Thus, a multi-layered RGS network is required to shape the magnitude and kinetics of MOR signaling and ensuing behavioral responses to opioids. The G protein regulatory network revealed here might also have broader implications for other Gαo/i-coupled receptors.