Neural Circuit in the Dorsal Raphe Nucleus Responsible for Cannabinoid-Mediated Increases in 5-HT Efflux in the Nucleus Accumbens of the Rat Brain

Abstract

In vivo microdialysis was used in this study to reveal the role of cannabinoids in regulating serotonin (5-HT) efflux in the nucleus accumbens (NAcc) and dorsal raphe nucleus (DRN). The cannabinoid CB1 receptor agonists WIN55212-2 and CP55940 systematically administered to rats caused significant increases in 5-HT efflux in the NAcc but failed to have an effect in the DRN. To reveal mechanisms underlying regionally selective responses, we tested the hypothesis that cannabinoids have both direct and indirect effects on 5-HT efflux, depending on the location of CB1 receptors in the neural circuit between DRN and NAcc. We showed that the direct effect of cannabinoids caused a reduction in 5-HT efflux whereas the indirect effect resulted in an increase. Furthermore, the indirect effect was blocked by the GABA(A) receptor antagonist bicuculline in the DRN, suggesting that the action is likely due to a presynaptic inhibition on GABAergic activity that exerts a tonic influence on neuronal circuits regulating 5-HT efflux. Involvement of GABAergic neurons was confirmed by measuring changes in GABA efflux. Taken together, our study suggests that cannabinoids may have direct and indirect effects on the 5-HT regulatory circuits, resulting in regionally selective changes of 5-HT efflux in the brain.

Figure 1

Systemic effect of CB1 receptor agonists on 5-HT efflux in the DRN and NAcc. Data are expressed as percent of average baseline values (% mean ± s.e.m.). Mean baseline level of extracellular 5-HT was 0.62 ± 0.02 pg/sample in the DRN (n = 24) and 0.31 ± 0.01 pg/sample in the NAcc (n = 44). The arrow indicates the time of the injection. (a) Systemic injection of WIN 55,212-2 (2.5 and 5 mg/kg, i.p.) had no effect on extracellular 5-HT in the DRN. (b) Systemic injection of WIN 55,212-2 (2.5 and 5 mg/kg, i.p.) produced dose- and time-dependent effects on extracellular 5-HT in the NAcc. (c) Systemic injection of CP55940 (0.25 and 0.5 mg/kg, i.p.) produced dose- and time-dependent effects on extracellular 5-HT in the NAcc. *P < 0.05 and ***P < 0.001 versus vehicle examined by the repeated measures ANOVA followed by the post hoc Scheffe test.

Figure 2

Effects of CB1 and opioid receptor antagonists on WIN 55,212-2-induced increases in 5-HT in the nucleus accumbens. Data are expressed as percent of average baseline values (% mean ± s.e.m.). Mean baseline level of extracellular 5-HT was 0.39 ± 0.04 pg/sample in the NAcc (n = 45). The arrows indicate the time of injection of antagonists 30 min before WIN 55,212-2. The effect of WIN 55,212-2 on 5-HT efflux in the NAcc was blocked by SR 147778 (5 mg/kg i.p.; (a)) and AM251 (5 mg/kg, i.p; (b)), but not by naloxone (10 mg/kg, s.c.; (c)). *P < 0.05 and ***P < 0.001 versus vehicle examined by the repeated measures ANOVA followed by post hoc Scheffe test.

Figure 3

Effect of reverse dialysis infusion of WIN 55,212-2 on 5-HT efflux in the DRN (a) or NAcc (b). Data are expressed as percent of average baseline values (% mean ± s.e.m.). Note that 1 μM citalopram was included in the infusion medium. Mean baseline level of extracellular 5-HT was 3.35 ± 0.06 pg/sample in the DRN (n = 23) and 4.15 ± 0.79 pg/sample in the NAcc (n = 24). The open horizontal bars indicate the 30-min period of drug infusion. (a) WIN 55,212-2 (100, 300 and 1000 μM) produced a concentration-dependent increase in 5-HT efflux in the DRN. (b) WIN 55,212-2 (100, 300, and 1000 μM) produced a concentration-dependent decrease in 5-HT efflux in the NAcc. *P < 0.05, **P < 0.01 and ***P < 0.001 versus vehicle examined by the repeated measures ANOVA followed by post hoc Scheffe test.

Figure 4

Data are expressed as percent of average baseline values (% mean ± s.e.m.). Mean baseline level of extracellular 5-HT was 0.39 ± 0.06 pg/sample in the NAcc (n = 37). (a) Effect on 5-HT efflux in the NAcc of reverse dialysis infusion of WIN 55,212-2 in the DRN. The open horizontal bars indicate the 30-min period of drug infusion. WIN 55,212-2 (300 and 1000 μM) in the DRN produced a concentration-dependent increase in 5-HT efflux in the NAcc. *P < 0.05 and **P < 0.01 versus vehicle examined by the repeated measures ANOVA followed by post hoc Scheffe test. (b) Effect on the WIN 55,212-2-induced increases in 5-HT efflux in the NAcc mediated by the reverse dialysis infusion of bicuculline (100 μM) in the DRN. Systemic injection of WIN 55,212-2 at 5 mg/kg i.p. took place at time zero. Bicuculline pretreatment in the DRN blocked the systemic effect of WIN 55,212-2 on 5-HT efflux in the NAcc.

Figure 5

Effect of systemic cannabinoids on GABA efflux in the DRN. The arrow indicates the time of injection of 5 mg/kg WIN 55,212-2, 0.5 mg/kg CP55940, or vehicle at t = 0 minutes. Data are expressed as percent of average baseline values (% mean ± s.e.m.). Mean baseline level of extracellular GABA was 0.32 ± 0.06 pmol/sample. There was a significant reduction in GABA efflux in the DRN following systemic injection of WIN 55,212-2 or CP55945. *P < 0.05, **P < 0.01, and ***P < 0.001 versus vehicle examined by the repeated measures ANOVA followed by post hoc Scheffe test.