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. 2020 Jul;177(13):2923-2931.
doi: 10.1111/bph.15004. Epub 2020 Feb 17.

Morphine-induced respiratory depression is independent of β-arrestin2 signalling

Andrea Kliewer  1 , Alexander Gillis  2 , Rob Hill  3 , Frank Schmiedel  1 , Chris Bailey  4 , Eamonn Kelly  3 , Graeme Henderson  3 , Macdonald J Christie  2 , Stefan Schulz  1
Affiliations

Morphine-induced respiratory depression is independent of β-arrestin2 signalling

Andrea Kliewer et al. Br J Pharmacol. 2020 Jul.

Abstract

Background and purpose: GPCRs can signal through both G proteins and β-arrestin2. For the μ-opioid receptor, early experimental evidence from a single study suggested that G protein signalling mediates analgesia, whereas β-arrestin2 signalling mediates respiratory depression and constipation. Consequently, for more than a decade, much research effort has been focused on developing biased μ-opioid agonists that preferentially target G protein signalling over β-arrestin signalling, as it was believed that such drugs would be analgesics devoid of respiratory depressant activity. However, the prototypical compounds that have been developed based on this concept have so far failed in clinical and preclinical development.

Experimental approach: The present study was set up to re-examine opioid-induced respiratory depression in β-arrestin2 knockout mice. To this end, a consortium was formed consisting of three different laboratories located in different countries to evaluate independently opioid-induced respiratory depression.

Key results: Our consensus results unequivocally demonstrate that the prototypical μ-opioid agonist morphine (3.75-100 mg·kg-1 s.c. or 3-30 mg·kg-1 i.p.) as well as the potent opioid fentanyl (0.05-0.35 mg·kg-1 s.c.) do indeed induce respiratory depression and constipation in β-arrestin2 knockout mice in a dose-dependent manner indistinguishable from that observed in wild-type mice.

Conclusion and implications: Our findings do not support the original suggestion that β-arrestin2 signalling plays a key role in opioid-induced respiratory depression and call into question the concept of developing G protein-biased μ-opioid receptor agonists as a strategy for the development of safer opioid analgesic drugs.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Respiratory depressant effect of morphine in β‐arrestin2 knockout mice. (a) Data: Sydney, Australia. Time course of respiratory rate depression following injection of 6, 10, or 50 mg·kg−1 of morphine s.c. measured with whole‐body plethysmography chambers (Buxco/DSI Instruments; n = 6–11). Parameters were normalised to the pre‐drug baseline as 100%. For dose–response curves, maximum depression of respiratory rate data from experiments in which mice were injected with 3–100 mg·kg−1 were fitted to a logistic function. (b) Data: Bristol, United Kingdom. Depression of respiratory rate following injection of 3, 10, and 30 mg·kg−1 morphine i.p. injection presented as respiratory rate (n = 6) measured with whole‐body plethysmography chambers (EMKA Technologies). The percentage respiratory rate following morphine injection for each animal was normalised to the pre‐drug baseline as 100%; baseline respiration rates before morphine injection in mice breathing 5% CO2 in air were 472.2 ± 23.7 and 484.5 ± 16.1 in wild‐type (WT) and β‐arrestin2 knockout mice, respectively (n = 6). Data are the means ± SEM. (c) Data: Jena, Germany. Time course of respiratory rate measured with a nose‐out plethysmography system (Harvard Apparatus) 30 min after 3.5, 15, or 52.5 mg·kg−1 of morphine s.c. (n = 6). Furthermore, dose–response curves of mean respiratory suppression over 30 min in which mice were injected with 3.5–52.5 mg·kg−1 are reported as the means ± SEM; baseline were set as 0. In (a)–(c), there was no statistical difference between morphine respiratory depression in β‐arrestin2 knockout and WT mice as determined by two‐way ANOVA with Bonferroni post hoc test. (d) Data: Jena, Germany. Time course of respiratory rate in WT and β‐arrestin2 knockout mice measured with nose‐out plethysmography system 15 min after 0.05, 0.1 or 0.3 mg·kg−1 of fentanyl s.c. (n = 6). Dose–response curves of respiratory suppression over 30 min in which mice were injected with 0.05–0.3 mg·kg−1 are reported as the means ± SEM; baselines were set as 0. Two‐way ANOVA with Bonferroni post hoc test. (e) Brain lysates from WT and β‐arrestin2 knockout mice (n = 2) were analysed for expression of β‐arrestin2. Blots were stripped and probed with anti‐actin antibody to confirm equal loading. The positions of molecular mass markers are indicated on the left (in kDa). (f) Schematic drawing of intracellular signalling cascades postulating that opioid‐induced respiratory depression is mediated by G protein signalling
Figure 2
Figure 2
Opioid‐induced constipation in β‐arrestin2 knockout mice. Dose–response curves for accumulated faecal boli weight in the constipation test after (a) morphine and (b) fentanyl administration (n = 6–18). Data are the means ± SEM; *indicate statistically significant differences between drug and vehicle; n.s. indicates no statistically significant differences between genotypes; one‐way ANOVA with Bonferroni post hoc test. Data from Jena, Germany
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References

    1. Alexander, S. P. H. , Christopoulos, A. , Davenport, A. P. , Kelly, E. , Mathie, A. , Peter, J. A. , … CGTP Collaborators (2019). The concise guide to pharmacology 2019/2020: G protein‐coupled receptors. British Journal of Pharmacology, 176, S21–S141. 10.1111/bph.14748 - DOI - PMC - PubMed
    1. Alexander, S. P. H. , Roberts, R. E. , Broughton, B. R. S. , Sobey, C. G. , George, C. H. , & Stanford, S. C. (2018). Goals and practicalities of immunoblotting and immunohistochemistry: A guide for submission to the British Journal of Pharmacology. British Journal of Pharmacology, 175, 407–411. - PMC - PubMed
    1. Bohn, L. M. , Gainetdinov, R. R. , Lin, F. T. , Lefkowitz, R. J. , & Caron, M. G. (2000). μ‐Opioid receptor desensitization by β‐arrestin‐2 determines morphine tolerance but not dependence. Nature, 408, 720–723. 10.1038/35047086 - DOI - PubMed
    1. Curtis, M. J. , Bond, R. A. , Spina, D. , Ahluwalia, A. , Alexander, S. P. , Giembycz, M. A. , … McGrath, J. (2015). Experimental design and analysis and their reporting: New guidance for publication in BJP. British Journal of Pharmacology, 172, 3461–3471. 10.1111/bph.12856 - DOI - PMC - PubMed
    1. Harding, S. D. , Sharman, J. L. , Faccenda, E. , Southan, C. , Pawson, A. J. , Ireland, S. , … NC‐IUPHAR (2018). The IUPHAR/BPS Guide to PHARMACOLOGY in 2018: Updates and expansion to encompass the new guide to IMMUNOPHARMACOLOGY. Nucleic Acids Research, 46, D1091–D1106. 10.1093/nar/gkx1121 - DOI - PMC - PubMed

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