Effects of the triple monoamine uptake inhibitor amitifadine on pain-related depression of behavior and mesolimbic dopamine release in rats

Abstract

Pain-related depression of behavior and mood is a key therapeutic target in the treatment of pain. Clinical evidence suggests a role for decreased dopamine (DA) signaling in pain-related depression of behavior and mood. Similarly, in rats, intraperitoneal injection of dilute lactic acid (IP acid) serves as a chemical noxious stimulus to produce analgesic-reversible decreases in both (1) extracellular DA levels in nucleus accumbens (NAc) and (2) intracranial self-stimulation (ICSS), an operant behavior reliant on NAc DA. Intraperitonial acid-induced depression of ICSS is blocked by DA transporter (DAT) inhibitors, but clinical viability of selective DAT inhibitors as analgesics is limited by abuse potential. Drugs that produce combined inhibition of both DA and serotonin transporters may retain efficacy to block pain-related behavioral depression with reduced abuse liability. Amitifadine is a "triple uptake inhibitor" that inhibits DAT with approximately 5- to 10-fold weaker potency than it inhibits serotonin and norepinephrine transporters. This study compared amitifadine effects on IP acid-induced depression of NAc DA and ICSS and IP acid-stimulated stretching in male Sprague-Dawley rats. Amitifadine blocked IP acid-induced depression of both NAc DA and ICSS and IP acid-stimulated stretching. In the absence of the noxious stimulus, amitifadine increased NAc levels of both DA and serotonin, and behaviorally, amitifadine produced significant but weak abuse-related ICSS facilitation. Moreover, amitifadine was more potent to block IP acid-induced depression of ICSS than to facilitate control ICSS. These results support consideration of amitifadine and related monoamine uptake inhibitors as candidate analgesics for treatment of pain-related behavioral depression.

Figure 1

Effects of amitifadine on control ICSS in the absence of the noxious stimulus. Left column shows drug effects on full ICSS frequency–rate curves. Abscissae: Frequency of electrical brain stimulation in Log Hz. Ordinates: Percent maximum control reinforcement rate (%MCR). Filled points represent frequencies at which reinforcement rates were statistically different from vehicle rates as determined by a two-way ANOVA followed by a Holm–Sidak post hoc test, P < 0.05. Right column shows summary ICSS data expressed as percent pre-drug baseline number of stimulations per component delivered across all frequencies of brain stimulation. Abscissae: drug dose in mg/kg. Ordinates: Percent baseline total number of stimulations per component. Upward/downward arrows indicate significant drug-induced increase/decrease in ICSS relative to vehicle for at least one brain stimulation frequency as determined by analysis of full frequency–rate curves. All data show mean ± SEM for five to six rats. Statistical results for data in left panels are as follows: (A) Amitifadine potency at a 30 min pretreatment time (N=5): significant main effect of frequency [F_(9,36) = 27.9, P < 0.001], no significant main effect of dose [F_(4,16) = 0.7, P = 0.59], and a significant frequency X dose interaction [F_(36,144) = 2.7, P = < 0.0001]. (B) 3.2 mg/kg amitifadine timecourse (n = 6): significant main effect of frequency [F_(9,45) = 32.1, P < 0.0001] but not of time [F_(5,25) = 0.7, P = 0.61], and no significant interaction [F_(45,225) = 1.4, P = 0.05] (C) 10 mg/kg amitifadine timecourse (n = 6): significant main effect of frequency [F_(9,45) = 36.6, P < 0.0001], time [F_(5,25) = 4.8, P < 0.003] and significant frequency X time interaction [F_(45,225) = 5.8, P < 0.0001].

Figure 2

Effects of amitifadine on 1.8% acid-induced depression of ICSS. Left column shows effects on full ICSS frequency–rate curves. Abscissae: Frequency of electrical brain stimulation in Log Hz. Ordinates: Percent maximum control reinforcement rate (%MCR). Right columns show summary ICSS data expressed as percent pre-treatment baseline total number of stimulations per component delivered across all frequencies of brain stimulation. Abscissae: drug dose in mg/kg. Ordinates: Percent baseline number of stimulations per component. All other details as described for Figure 1. All data show mean ± SEM for five to six rats. Statistical results for data in left panels are as follows: (A) 1.8% Acid effect (n = 5): significant main effect of frequency [F_(9,36) = 19.4, P < 0.001], treatment [F_(1,4) = 25.1, P < 0.01], and significant frequency X treatment interaction [F_(9,36) = 3.2, P < 0.01]. (B) Amitifadine effects determined at 30 min post-treatment (n = 5): significant main effect of frequency [F_(9,36) = 53.8, P < 0.0001], no significant main effect of dose [F_(4,16) = 1.8, P = 0.17], and significant frequency X treatment interaction [F_(36,144) = 4.9, P = < 0.0001]. (C) Amitifadine effects determined at 100 min post-treatment (n = 6): significant main effect of frequency [F_(9,45) = 54.8, P < 0.0001], dose [F_(2,10) = 25.83, P < 0.0001] and significant frequency X treatment interaction [F_(18,90) = 4.7, P = < 0.0001].

Figure 3

Effects of amitifadine on 5.6% acid-induced depression of ICSS. Left column shows effects on full ICSS frequency–rate curves. Abscissae: Frequency of electrical brain stimulation in Log Hz. Ordinates: Percent maximum control reinforcement rate (%MCR). Right column shows summary ICSS data expressed as percent baseline number of stimulations per component delivered across all frequencies of brain stimulation. Abscissae: drug dose in mg/kg. Ordinates: Percent baseline total number of stimulations per component. All other details as described for Figure 1. All data show mean ± SEM for five to six rats. Statistical results for data in left panels are as follows: (A) 5.6% Acid effect (n = 5): significant main effect of frequency [F_(9,36) = 83.3, P < 0.0001], treatment [F_(1,4) = 17.0, P < 0.05], and significant frequency X treatment interaction [F_(9,36) = 2.4, P < 0.05]. (B) Amitifadine effects determined at 100 min post-treatment (n = 6): significant main effect of frequency [F_(9,36) = 24.2, P < 0.0001], no significant main effect of dose [F_(2,8) = 2.4, P = 0.15], and a significant frequency X treatment interaction [F_(18,72) = 3.7, P = < 0.0001].

Figure 4

Effects of amitifadine on control levels of nucleus accumbens extracellular dopamine (left panel) and serotonin (right panel) in the absence of the noxious stimulus. Abscissae: Time in min after injection of amitifadine or vehicle. Ordinates: Percent baseline neurotransmitter levels. Filled points represent time points at which neurotransmitter levels were statistically different from vehicle levels as determined by a two-way ANOVA followed by a Holm–Sidak post hoc test, P < 0.05. All data show mean ± SEM for five to six rats. Statistical results for data in left panels are as follows: (A) Amitifadine effects on dopamine (n = 6): significant main effect of time [F_(30,600) = 15.1, P < 0.01] and treatment [F_(3,20) = 10.2, P < 0.001], and significant time X treatment interaction [F_(90,300) = 2.5, P = < 0.01]. (B) Amitifadine effects on serotonin (n = 5–7): significant main effect of time [F_(30,600) = 22.0, P < 0.001] and treatment [F_(3,20) = 3.8, P < 0.05], and significant time X treatment interaction [F_(90,300) = 3.6, P = < 0.001].

Figure 5

Effects of IP 5.6% acid on extracellular dopamine in the absence or presence of 3.2 mg/kg amitifadine. Abscissae: Time after injection of amitifadine or vehicle. Ordinates: Percent baseline neurotransmitter levels. “Inj 1” represents time of vehicle or amitifadine injection. “Inj 2” represents time of vehicle or acid injection. Filled points represent time points at which neurotransmitter levels were statistically different from vehicle levels as determined by a two-way ANOVA followed by a Holm–Sidak post hoc test, P < 0.05. All data show mean ± SEM for six rats. Statistical results for data in left panels are as follows: (A) 5.6% acid effects on dopamine levels after vehicle pretreatment (n = 6): significant main effect of time [F_(20,200) = 8.8, P < 0.001], treatment [F_(1,10) = 5.2, P < 0.05], and significant time X treatment interaction [F_(20,200) = 3.1, P = < 0.001]. (B) 5.6% acid effects on dopamine levels after amitifadine pretreatment (n = 6): significant main effect of time [F_(20,200) = 18.5, P < 0.001].

Figure 6

Effects of amitifadine on 1.8% acid-induced stretching. Abscissa: dose of amitifadine administered 30 min before lactic acid. Ordinate: number of stretches during 30-min observation period. Filled point represents dose at which stretching was significantly decreased compared to amitifadine vehicle pretreatment as determined by one-way ANOVA followed by Dunnetts test, p < 0.05. All data show mean ± SEM for five rats. One-way ANOVA indicated a significant effect of amitifadine [F_(3,12) = 7.8, P < 0.005]