Table 1.
In vitro uptake inhibitory potency (pIC50) and apparent binding affinity (pKi) of fluoxetine, duloxetine, atomoxetine and esreboxetine in rat cortical membrane or synaptosomal preparations, respectively (n = 3–12).
Figure 1.
Neither esreboxetine nor fluoxetine exhibited antinociceptive synergy with morphine in the rat formalin model.
(A) The selective NET inhibitor esreboxetine alone (Esrbx, IP, 10 mg/kg), failed to shift the morphine (Mor) dose-response curve (n = 6). Morphine alone: ED50 = 2.0 mg/kg (95% CI: 1.8–2.5); morphine+ esreboxetine (IP, 10 mg/kg): ED50 = 1.6 mg/kg (95% CI: 1.1–2.4). All data points are shown as mean ± SEM for each group and are expressed as percentage of controls. Inset (A) Esreboxetine (IP, 10 mg/kg) was associated with 82±5% NET and 3±5% SERT occupancy measured ex vivo at 75 min post-dose. All occupancy data represent mean (± SEM) for each group. (B) The selective SERT inhibitor fluoxetine alone (Flx, IP, 10 mg/kg), failed to shift the morphine dose-response curve (n = 6). Morphine+fluoxetine (IP, 10 mg/kg): ED50 = 1.6 mg/kg (95% CI: 1.2–2.2). Inset (B) Fluoxetine (IP, 10 mg/kg) was associated with 6±12% NET and 89±5% SERT occupancy measured ex vivo at 75 min post-dose.
Figure 2.
Atomoxetine exhibited antinociceptive synergy with morphine using a fixed-dose design in the rat formalin model.
(A) Both 3 and 10 mg/kg atomoxetine (Atx, IP) shifted the morphine (Mor) dose-response curve leftward in the rat formalin model (n = 6–16). Morphine alone: ED50 = 2.3 mg/kg (95% CI: 2.0–2.5); morphine+atomoxetine (IP, 3 mg/kg): ED50 = 1.1 mg/kg (95% CI: 0.8–1.6); and morphine+atomoxetine (IP, 10 mg/kg): ED50 = 0.6 mg/kg (95% CI: 0.4–0.8). All data points are shown as mean ± SEM for each group and are expressed as percentage of controls. Inset (A) Atomoxetine (IP) at 3 and 10 mg/kg was associated with 67±10% and 84±3% for NET and 35±9% and 64±5% for SERT occupancy measured ex vivo at 75 min post-dose, respectively. All occupancy data represent mean (± SEM) for each group. (B) A subefficacious dose of morphine 1 mg/kg (SC) left-shifted the atomoxetine dose-response curve (n = 6–16). Atomoxetine alone: ED50 = 27.8 mg/kg (95% CI: 22–36); and atomoxetine+morphine (SC, 1 mg/kg): ED50 = 2.5 mg/kg (95% CI: 1.3–4.7). (C) A fixed combination of NET selective inhibitor esreboxetine (Esrbx, IP, 10 mg/kg) and SERT selective inhibitor fluoxetine (Flx, IP, 1 mg/kg) left-shifted the morphine dose-response curve (n = 6–12). Morphine alone: ED50 = 2.3 mg/kg (95% CI: 2.0–2.5); morphine+esreboxetine (IP, 10 mg/kg)+fluoxetine (IP, 1 mg/kg): ED50 = 0.3 mg/kg (95% CI: 0.2–0.7).
Table 2.
Plasma unbound and brain unbound concentrations of atomoxetine in absence or presence of morphine; esreboxetine in absence or presence of morphine and/or fluoxetine; fluoxetine in absence or presence of morphine and/or esreboxetine; and duloxetine in absence or presence of morphine and ondansetron - at 75 min post-dosing.
Figure 3.
Atomoxetine exhibited antinociceptive synergy with morphine using a fixed-ratio design in the rat formalin model.
(A) The dose-response curve of a fixed-ratio of 3 parts atomoxetine (Atx, IP) to 1 part morphine (Mor, SC) leftward shifted relative to the atomoxetine dose-response curve alone (n = 6–12). All data points are shown as mean ± SEM for each group and are expressed as percentage of controls. (B) An isobologram for the combined effects of atomoxetine and morphine in a fixed ratio combination 3∶1. The ED50 value for morphine is plotted on the abscissa, and the ED50 value for atomoxetine is plotted on the ordinate. The solid line represents the line of additivity and the isobol point (observed ED50 value) is located to the left and below the theoretical additive ED50 value (with non-overlapping 95% CI). (C) The dose-response curve of a fixed-ratio of concomitant administration of 10 part atomoxetine (IP) to 1 part morphine (SC) leftward shifted relative to the atomoxetine dose-response curve alone (n = 6–16). All data points are shown as mean ± SEM for each group and are expressed as percentage of controls. (D) An isobologram for the combined effects of atomoxetine and morphine in a fixed ratio combination 10∶1. The isobol point (observed ED50 value) is located to the left and below the theoretical additive ED50 value (without overlapping 95% CI).
Figure 4.
The antinociceptive activity of atomoxetine in the rat formalin model was independent of µ-opioid receptor activation.
The µ-opioid receptor antagonist naloxone (Nal, IP, 5 mg/kg), at a dose which effectively blocked morphine (Mor)-induced analgesia in the rat formalin model, did not inhibit atomoxetine (Atx)-induced antinociception (n = 5–7). All values are shown as mean ± SEM for each group and are expressed as percentage of controls. Student’s t test, t (10) = 7.668, ***p<0.001.
Figure 5.
Antinociceptive synergy between atomoxetine and morphine did not reflect impaired motor coordination.
The white bars represent the % reduction in the flinching behavior compared to vehicle-treatment in the rat formalin model (n = 10–22), and the grey bars represent the change in latency for rats to fall from an accelerating rotating rod compared to vehicle treatment in the rat RotaRod test (n = 8). All data points are shown as mean ± SEM for each group and are expressed as percentage of controls. Data from one-way ANOVA are as follows: rat formalin model: F (4, 53) = 36.12, p<0.0001; RotaRod: F (4, 34) = 4.604, p = 0.004. Data from the post hoc Dunnett’s test follows: **p<0.01, q = 3.265; ***p<0.001, q = 9.258–9.370, compared to vehicle treatment.
Figure 6.
Duloxetine failed to exhibit antinociceptive synergy with morphine in the rat formalin model.
(A) Duloxetine (Dlx) at 5 mg/kg failed to shift the morphine (Mor) dose-response curve leftward. Morphine alone: ED50 = 2.3 mg/kg (95% CI: 2.0–2.5); morphine+duloxetine (IP, 5 mg/kg): ED50 = 2.0 mg/kg (95% CI: 1.3–3.0). All data points are shown as mean ± SEM for each group and are expressed as percentage of controls. Inset (A) Duloxetine (IP) at 5 mg/kg was associated with 62±5% for NET and 92±3% for SERT occupancy measured ex vivo at 75 min post-dose. All occupancy data represent mean (± SEM) for each group. (B) A subefficacious dose of morphine 1 mg/kg (SC) failed to left-shift the duloxetine dose-response curve (n = 6–12). Duloxetine alone: ED50 = 10.9 mg/kg (95% CI: 8–15); and duloxetine+morphine (SC, 1 mg/kg): ED50 = 7.7 mg/kg (95% CI: 4–16).
Figure 7.
Coadministration of ondansetron potentiates the antinociceptive response to duloxetine and morphine in the rat formalin model.
(A) Co-administration of the 5-HT3 receptor selective antagonist ondansetron (Ond, IP, 3 mg/kg) potentiates the antinociceptive response to duloxetine (Dlx, IP, 5 mg/kg) and morphine (Mor, SC, 1 mg/kg). Ondansetron alone, or in combination with either morphine or duloxetine, did not exhibit antinociceptive activity (n = 6–19). All data points are shown as mean ± SEM for each group and are expressed as percentage of controls. One-way ANOVA: F (7, 75) = 7.447, p<0.0001. Data from the post hoc Newman-Keuls test follows: ***p<0.001, q = 4.956–9.764 for duloxetine+morphine+ondansetron versus the other groups. (B) Co-administration of ondansetron (IP, 3 mg/kg) did not reveal antinociceptive synergy between the SERT selective reuptake inhibitor fluoxetine (Flx, IP, 10 mg/kg) and morphine (SC, 1 mg/kg). Ondansetron alone, or in combination with either morphine or fluoxetine, did not exhibit antinociceptive activity (n = 7). One-way ANOVA followed by post hoc Newman-Keuls: p>0.05 morphine versus fluoxetine+morphine; p>0.05 fluoxetine+morphine versus fluoxetine+morphine+ondansetron.