S426/S430 phosphorylation accounts for Beta‐arrestin 2‐mediated desensitization for cannabinoid sensitivity and tolerance in mutant mice

Abstract

Desensitization of G protein‐coupled receptors, such as the cannabinoid receptor 1 (CB1R), is a key mechanism involved in the in the development of tolerance. CB1R desensitization is mediated via phosphorylation of residues S426 and S430 by a G protein‐coupled receptor kinase (GRK). Following GRK‐mediated phosphorylation at these residues, beta‐arrestin 2 (bArr2) is recruited to facilitate desensitization of the receptor. Previous work demonstrated that tolerance for cannabinoid drugs was reduced in S426A/S430A mutant mice and in bArr2‐KO mice. Since there are other phosphorylatable serine and threonine residues in the C‐terminal tail of CB1R, it was not clear whether bArr2 recruitment to S426 and S430 accounts for all bArr2‐mediated desensitization. To address this question, we assessed cannabinoid acute response and daily tolerance in S426A/S430A x bArr2‐KO double mutant mice. Both S426A/S430A and bArr2‐KO single mutant mice display increased sensitivity and delayed tolerance to the antinociceptive effects of delta‐9‐tetrahydrocannabinol (Δ9‐THC) and the synthetic cannabinoid, CP55,940. Double mutant mice did not differ from the S426A/S430A single mutant model in respect to antinociception, suggesting that the effect of complete bArr2 deletion did not enhance the effect of the S426A/S430A point mutants. However, the hypothermic effects of acute Δ9‐THC dosing were increased in male double mutant mice, relative to S426A/S430A and bArr2 single mutant mice. These results indicate that phosphorylation of S426 and S430 are likely responsible for bArr2‐mediated desensitization, with the exception of hypothermia induced by acute Δ9‐THC dosing. Therefore, these results also suggest sensitivity to cannabinoid‐induced hypothermia is agonist‐ and duration‐dependent.