Antidepressant-like Activity of a Triple Combination of Venlafaxine, Codeine and Nor-Binaltorphimine in the Mouse Tail Suspension Test

About one-third of patients with major depressive disorder do not remit on conventional monoaminergic antidepressants. Venlafaxine (VEN) is a classic antidepressant that inhibits serotonin and norepinephrine reuptake. Venlafaxine does not bind opioid receptors, yet its antidepressant-like action requires intact µ-opioid signalling. The dynorphin/κ-opioid axis exerts an opposing, dysphoria-promoting influence. A possible question is whether combining serotonin and noradrenaline reuptake inhibition, µ-opioid receptor activation by codeine (COD) and sustained κ-opioid receptor blockade produces an antidepressant-like effect larger than that of the components given alone. Male NMRI mice were tested in the forced swim test (FST) (one experiment) and in the tail suspension test (TST) (three experiments) using a parallel-design (n = 10 to 17 per group; 277 animals), as follows: VEN 10 to 40 mg/kg in both tests—FST and TST; COD 10 to 40 mg/kg added to VEN 20 mg/kg in TST; and nor-binaltorphimine (norBNI) 10 mg/kg, given 24 h before, combined with VEN 20 mg/kg and COD 20 mg/kg and compared with norBNI alone, COD alone and norBNI + VEN, with amitriptyline (AMI) 20 mg/kg as the positive control in the TST. The three agents and two reference ligands were also docked into eight experimental human structures of the monoamine transporters and of the μ, κ and δ opioid receptors. VEN did not reduce FST immobility at any dose but produced a dose-dependent reduction in the TST, reaching −66.2% at 40 mg/kg. COD did not potentiate VEN 20 mg/kg at any of three COD doses. The triple combination, in the TST, reduced immobility by 65.7% relative to saline (Dunnett p = 0.013), an effect of similar magnitude to AMI with no significant difference detected between them, whereas norBNI alone, COD alone and norBNI + VEN did not reach significance. The design was underpowered for intermediate effects (the calculated n should be about 36 per group, unapproved because of ethical considerations). The triple combination was not significantly more effective than norBNI + VEN (p = 0.375), so the contribution of codeine remains undetermined. Docking reproduced the experimental pose of all eight co-crystallised ligands and was consistent with VEN occupying the transporter S1 site, COD the μ-opioid pocket and norBNI the κ receptor. A triple serotonin-adrenergic and opioid combination produced an antidepressant-like effect that none of the single treatments tested in the same cohort reached, but the fixed-dose design does not establish cooperativity between all three mechanisms. The docking results are consistent with, but do not prove, engagement of three separate sites.