After baseline nociceptive testing, morphine sulphate (Sigma) was administered to mice subcutaneously (s.c.) 20 mg/kg twice per day on days 1–3 and 40 mg/kg twice per day on day 4 in 50–100 μl volumes of 0.9 % NaCl. This protocol has been used to demonstrate opioid tolerance, dependence, hyperalgesia and other maladaptive phenotypes in mice in previous experiments [13 (link), 18 (link)].
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