GRASP 2010 Sample Abstract

THE COUPLING OF CB1 RECEPTORS WITH BETA-ARRESTIN1 AND 2 Deryck Hill and Chris Breivogel Campbell University College of Pharmacy and Health Sciences, Buies Creek, NC 27506 USA

Background/Purpose: Previous studies in our laboratory have demonstrated that beta- arrestin2-/- mice are more sensitive than wild-type mice to the physiological and behavioral effects THC, while there appeared to be no difference between these genotypes in the effects other cannabinoid agonists like CP5540 or methanandamide. The present studies were conducted to find evidence for the biochemical basis for the agonist selective effects of beta- arrestin2 by: 1) characterizing relative amounts of beta-arrestin1 and beta-arrestin2 in various mouse brain regions, 2) determining if knocking out beta-arrestin2 affects beta-arrestin1 levels the same brain regions, and 3) assessing CB1 receptor association with beta-arrestin1 and beta- arrestin2 in response to THC and CP55940. Methods: Western blotting was employed to quantify the relative amounts of beta-arrestin1 and beta-arrestin2 protein within each of six (cerebellum, hippocampus, basal ganglia, brain stem, midbrain, and cortex) mouse brain areas. Co-immunoprecipitation of CB1 from CHAPS- solubilized cortex membrane homogenates was used to determine association of beta-arrestin1 and 2 with CB1 receptors from mice treated with vehicle, THC or CP55940. Results: This is the first demonstration of relative levels of beta-arrestin1 and 2 in mouse brain. We found that beta-arrestin1 was highest in cortex and hippocampus and beta-arrestin2 was highest in cortex. Thus, subsequent co-immunoprecipitation experiments were carried out using mouse brain cortex. Comparisons within the same brain regions between wild type and beta- arrestin2 knockout samples revealed no difference in the amount of beta-arrestin1 was produced by the absence of beta-arrestin2. Co-immunoprecipitation experiments are preliminary, but so far it has been observed that beta-arrestin1 does associate with CB1. Conclusions: There did not appear to be any physiological compensatory production of beta- arrestin1 in beta-arrestin2-/- mice. The data are too preliminary at this point to determine the association of beta-arrestin2 with CB1 whether this association is affect by agonist treatments.

Financial support was provided by Campbell University College of Pharmacy & Health Sciences