Background & Aims
Sex and gender differences in the analgesic efficacy and side effects of opioids have been widely reported (Aubrun et al., 2005; Filllingim et al., 2005). Preclinical animal research is an essential resource for investigating the neurobiological mechanisms of opioids, including sex differences. So far, studies have mainly used rats to reveal a number of sex differences in the regional expression of endogenous opioids and µ-opioid receptors (MOR) (Sharp et al., 2022). Extending these studies using transgenic mouse lines would allow for powerful experiments that could probe the functional consequences of these differences. However, sex and strain differences in the behavioural effects of opioids in mice remains largely unknown. Therefore, the aim of this study was to characterise and compare sex differences in the analgesic and side effect profiles of morphine, and quantify regional MOR expression for the two most common strains of mice used in preclinical research – CD1 and C57BL/6.
Methods
24 male and 24 female CD1 and C57BL/6 mice were used for this study. Dose-response profiles to morphine analgesia (0, 5, 10 and 20mg/kg) for each sex and strain were established using the hot plate test (45 seconds at 51°C). Each mouse received each dose with a washout period of at least 3 days between doses, and hind paw withdrawal latency and hind paw licking was measured at 0, 30 and 60 minutes after administration. Morphine-induced hyperlocomotor activity was the primary side effect measure and was quantified using DeepLabCut software during the 60 minutes after vehicle or morphine administration. The regional expression of u-opioid receptors was quantified using western blots in the spinal cord and 8 brain regions – the rostral ventromedial medulla, the periaqueductal gray, the anterior cingulate cortex, the amygdala, hippocampus, the motor cortex, nucleus accumbens and caudate nucleus.
Results
Significant strain differences were observed in the analgesic effects of morphine at low doses but not at high doses. CD1 mice showed longer hind paw withdrawal latency than C57BL/6 mice at 5mg/kg (p=0.017, two-way ANOVA), but not at 10 or 20mg/kg. Small sex differences were observed in each strain only at 5mg/kg, with larger reductions in hind paw withdrawal licking seen in male CD1 mice (p=0.014) and female C57BL/6 mice (p=0.048, unpaired t-tests). In fact, in male C57BL/6 mice, 5mg/kg produced hyperlocomotor side effects without analgesia. Large strain differences, but no sex differences, in hyperlocomotor side effects were observed at all doses: 5mg/kg (p=0.004), 10mg/kg (p<0.0001) and 20mg/kg (p=0.02, two-way ANOVAs). Interestingly, linear regressions revealed that the degree of analgesia was positively correlated with the hyperlocomotor activity for CD1 males (p=0.03), but negatively correlated for CD1 females (p=0.015).
Conclusions
This is the first study to investigate both sex and strain differences in the analgesic and hyperlocomotor side effects of morphine in mice. In contrast to prior research in mice and rats, only small sex differences were observed in the analgesic effects of morphine, which could be related to the different pain assays used between studies. The dissociations between the analgesic effect and hyperlocomotor side effect of morphine across sexes and strains is likely due to differences in regional opioid receptor expression in the central nervous system. The stronger morphine-induced hyperlocomotor activity seen in C57BL/6 mice suggests this strain is better suited for investigating the neural mechanisms of opioid side-effects. The lack of strong behavioural sex differences in either strain suggests that research into the neurobiological sex differences in the processing of opioids may be best pursued using rats and people.
References
Aubrun F, Salvi N, Coriat P, Riou B. Sex- and age-related differences in morphine requirements for postoperative pain relief. Anesthesiology. 2005 Jul;103(1):156-60. doi: 10.1097/00000542-200507000-00023. PMID: 15983468.
Fillingim RB, Ness TJ, Glover TL, Campbell CM, Hastie BA, Price DD, Staud R. Morphine responses and experimental pain: sex differences in side effects and cardiovascular responses but not analgesia. J Pain. 2005 Feb;6(2):116-24. doi: 10.1016/j.jpain.2004.11.005. PMID: 15694878.
Sharp JL, Pearson T, Smith MA. Sex differences in opioid receptor mediated effects: Role of androgens. Neurosci Biobehav Rev. 2022 Mar;134:104522. doi: 10.1016/j.neubiorev.2022.104522. Epub 2022 Jan 4. PMID: 34995646; PMCID: PMC8872632.
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Presenting Author
Damien Boorman
Poster Authors
Damien Boorman
BA, BSc, PHD
University of Toronto Mississauga
Lead Author
Topics
- Gender/Sex Differences