title: β-Arrestin-2 knockout prevents development of cellular μ-opioid receptor tolerance but does not affect opioid-withdrawal-related adaptations in single PAG neurons.
abstract: OBJECTIVE
  Tolerance to the behavioural effects of morphine is blunted in β-arrestin-2 knockout mice, but opioid withdrawal is largely unaffected. The cellular mechanisms of tolerance have been studied in some neurons from β-arrestin-2 knockouts, but tolerance and withdrawal mechanisms have not been examined at the cellular level in periaqueductal grey (PAG) neurons, which are crucial for central tolerance and withdrawal phenomena.
  
  
  METHODS
  μ-Opioid receptor (MOPr) inhibition of voltage-gated calcium channel currents (ICa ) was examined by patch-clamp recordings from acutely dissociated PAG neurons from wild-type and β-arrestin-2 knockout mice treated chronically with morphine (CMT) or vehicle. Opioid withdrawal-induced activation of GABA transporter type 1 (GAT-1) currents was determined using perforated patch recordings from PAG neurons in brain slices.
  
  
  RESULTS
  MOPr inhibition of ICa in PAG neurons was unaffected by β-arrestin-2 deletion. CMT impaired coupling of MOPrs to ICa in PAG neurons from wild-type mice, but this cellular tolerance was not observed in neurons from CMT β-arrestin-2 knockouts. However, β-arrestin-2 knockouts displayed similar opioid-withdrawal-induced activation of GAT-1 currents as wild-type PAG neurons.
  
  
  CONCLUSIONS
  In β-arrestin-2 knockout mice, the central neurons involved in the anti-nociceptive actions of opioids also fail to develop cellular tolerance to opioids following chronic morphine. The results also provide the first cellular physiological evidence that opioid withdrawal is not disrupted by β-arrestin-2 deletion. However, the unaffected basal sensitivity to opioids in PAG neurons provides further evidence that changes in basal MOPr sensitivity cannot account for the enhanced acute nociceptive response to morphine reported in β-arrestin-2 knockouts.
  
  
  BACKGROUND
  This article is part of a themed section on Opioids: New Pathways to Functional Selectivity. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue-2.
authors.affiliation: Australian School of Advanced Medicine, Macquarie University, Sydney, NSW, Australia.
authors.forename: M
authors.initials: M
authors.lastname: Connor
authors.forename: E E
authors.initials: EE
authors.lastname: Bagley
authors.forename: B C
authors.initials: BC
authors.lastname: Chieng
authors.forename: M J
authors.initials: MJ
authors.lastname: Christie
mesh_terms: D000701:Analgesics, Opioid / Q000494:pharmacology; D000818:Animals; D019390:Arrestins / Q000235:genetics / Q000502:physiology*; D004361:Drug Tolerance / Q000502:physiology*; D050485:GABA Plasma Membrane Transport Proteins / Q000502:physiology; D008297:Male; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D009020:Morphine / Q000494:pharmacology; D009474:Neurons / Q000502:physiology; D010487:Periaqueductal Gray / Q000502:physiology*; D017450:Receptors, Opioid, mu / Q000502:physiology*; D013375:Substance Withdrawal Syndrome / Q000503:physiopathology*; D000071560:beta-Arrestin 2; D000071557:beta-Arrestins
chemical_list: D000701:Analgesics, Opioid; C000605258:Arrb2 protein, mouse; D019390:Arrestins; D050485:GABA Plasma Membrane Transport Proteins; D017450:Receptors, Opioid, mu; C493414:Slc6a1 protein, mouse; D000071560:beta-Arrestin 2; D000071557:beta-Arrestins; D009020:Morphine
keywords: arrestins; morphine; opioid tolerance; opioid withdrawal; periaqueductal grey; μ-opioid receptor