Cocaine-induced endocannabinoid release modulates behavioral and neurochemical sensitization in mice

Abstract

The endocannabinoid system has been implicated in the development of synaptic plasticity induced by several drugs abused by humans, including cocaine. However, there remains some debate about the involvement of cannabinoid receptors/ligands in cocaine-induced plasticity and corresponding behavioral actions. Here, we show that a single cocaine injection in Swiss-Webster mice produces behavioral and neurochemical alterations that are under the control of the endocannabinoid system. This plasticity may be the initial basis for changes in brain processes leading from recreational use of cocaine to its abuse and ultimately to dependence. Locomotor activity was monitored with photobeam cell detectors, and accumbens shell/core microdialysate dopamine levels were monitored by high-performance liquid chromatography with electrochemical detection. Development of single-trial cocaine-induced behavioral sensitization, measured as increased distance traveled in sensitized mice compared to control mice, was paralleled by a larger stimulation of extracellular dopamine levels in the core but not the shell of the nucleus accumbens. Both the behavioral and neurochemical effects were reversed by CB1 receptor blockade produced by rimonabant pre-treatments. Further, both behavioral and neurochemical cocaine sensitization were facilitated by pharmacological blockade of endocannabinoid metabolism, achieved by inhibiting the fatty acid amide hydrolase enzyme. In conclusion, our results suggest that a single unconditioned exposure to cocaine produces sensitization through neuronal alterations that require regionally specific release of endocannabinoids. Further, the present results suggest that endocannabinoids play a primary role from the earliest stage of cocaine use, mediating the inception of long-term brain-adaptive responses, shaping central pathways and likely increasing vulnerability to stimulant abuse disorders.

Keywords: Behavioral sensitization; cannabinoid CB1 receptors; cocaine addiction; endocannabinoids; in vivo dopamine microdialysis; nucleus accumbens.

Figure 1

Brain microdialysis probe placements in the NAC shell or core. Forebrain sections, redrawn from Paxinos and Franklin (2001), showing the limits of the positions of the dialyzing portions of the microdialysis probes (superimposed rectangles). On each section the anterior coordinate (measured from bregma) is indicated.

Figure 2. Cocaine-induced single trial behavioral sensitization is blunted by antagonism at CB1 receptors

Locomotor activity response to an acute challenge with cocaine (panel A, 10 mg/kg, or panel B, 20 mg/kg) in mice pretreated 24 hr and 40 minutes before the cocaine challenge with vehicle [A two-Way ANOVA yielded significant main effects of cocaine pre-exposure, F(1,67) = 11.235, p<0.002, and cocaine treatment, F(2,67) = 7.527, p<0.002, and a non-significant effect of their interaction: F(2,67) = 2.682, p=0.075] or increasing doses of rimonabant (RIM), 0.3 (hatched bars), 1.0 (light grey filled bars), and 3 mg/kg i.p., (dark grey filled bars), followed 40 minutes later with either saline (unfilled bars) or cocaine 20 mg/kg i.p. (solid black bars). [One-Way ANOVA of the effects of RIM 0.3-3.0 mg/kg pretreatments was significant at both the cocaine challenges, 10 mg/kg F(3,25) = 3.689, p<0.005), and 20 mg/kg F(3,32) = 3.042, p<0.005]. The results are expressed as the average distance traveled (cm/30 min) ± S.E.M. obtained for each experimental group of animals during the first 30 minutes after injection. Significant differences from control groups,* = p< 0.05 VS saline, or # = p<0.05 VS cocaine.

Figure 3. Cocaine-induced single trial behavioral sensitization is accompanied by increased stimulation of NAC core, but not NAC shell, DA transmission, which is blocked by antagonism at CB1 receptors

The figure shows the time course of the effects of systemic administration of cocaine 10 mg/kg (top panels, A, C), or 20 mg/kg (lower panels, B,D) on extracellular levels of DA in dialysates from the NAC core (left panels) or shell (right panels) in control animals (● filled circles, number of mice and basal levels of extracellular DA expressed as femtomoles/10-μl sample ± S.E.M. were: n=5, 41.1 ± 3.9, and n=5, 42.9 ± 4.9 fmoles/sample, for cocaine 10 and 20 mg/kg, respectively in the core; n=6, 57.9 ± 6.7 fmoles/sample, and n=9, 48.4 ± 4.7 fmoles/sample, for cocaine 10 and 20 mg/kg, respectively in the shell), or in animals sensitized to cocaine (○ unfilled circles, n=7, 37.4 ± 2.0 fmoles/sample, and n=6, 30 ± 3.4 fmoles/sample, for cocaine 10 and 20 mg/kg, respectively in the core; n=9, 48.5 ± 3.0 fmoles/sample, and n=8, 51.3 ± 4.2 fmoles/sample, for cocaine 10 and 20 mg/kg, respectively in the shell), in animals pretreated with RIM 1 mg/kg and cocaine 20 mg/kg (◻ unfilled squares, n=6, 52.4 ± 3.6 fmoles/sample, and n=6, 51.8 ± 4.4 fmoles/sample, for cocaine 10 and 20 mg/kg, respectively in the core; n=9, 44.2 ± 3.7 fmoles/sample, and n=6, 29.1 ± 3.1 fmoles/sample, for cocaine 10 and 20 mg/kg, respectively in the shell) and in animals pretreated with RIM 3 mg/kg and cocaine (▵, unfilled triangles, n=6, 59.1 ± 5.7 fmoles/sample, and n=6, 52.1 ± 9.1 fmoles/sample, for cocaine 10 and 20 mg/kg, respectively in the core) or pretreated with RIM 3 mg/kg and saline (∎filled squares, n=4, 35.0 ± 7.7 fmoles/sample, for cocaine 20 mg/kg in the core) 24 hr before microdialysis tests. [Three-Way ANOVA (RIM pretreatment X cocaine treatment X time) showed in the core significant main effects of cocaine dose F(1,42)= 15.296, p<0.001, RIM pretreatment F (3, 42) = 3.168, p<0.05, and time F (6,252)= 91.066, p<0.001; also, there were significant interactions of cocaine dose × time F(18,252) = 19.735, p<0.001, RIM pretreatment × time F(18,252)= 2.297, p<0.05; non-significant interactions of RIM pretreatment × cocaine dose F (3.42)=0.518, NS; and pretreatment × cocaine dose × time interaction F= (18,252)= 0.486, NS. Three-Way ANOVA in the shell showed significant main effects of cocaine dose F(1,36)= 24.057, p<0.001, and time F (12,432)= 101.088, p<0.001, and a non-significant main effect of RIM pretreatment F (2, 36) = 0.139; also, there were significant interactions of cocaine dose × time F(12,432) = 19.735, p<0.001, non-significant interactions of pretreatment × cocaine dose F (2,36)=0.872; pretreatment × time F(24,432)= 1.007; and pretreatment × cocaine dose × time interaction F= (24,432)= 0.594]. Results are means with vertical bars representing S.E.M. of the amount of DA in 10 minutes dialysate samples, expressed as percentage of basal values.

Figure 4. Blockade of the FAAH enzyme by URB597 facilitates the occurrence of cocaine-induced single trial behavioral and NAC core neurochemical sensitization in animals pretreated with a non-sensitizing dose of cocaine

This figure shows on panel A that mice pretreated with a 10 mg/kg dose of cocaine 24 hr in advance, and then tested with a dose of cocaine, 20 mg/kg i.p., did not display behavioral sensitization, measured as increased distance traveled, as compared with mice that were pretreated with saline 24 hr before [one way ANOVA shows NS effect of sensitization treatment, F(1,14)=0.029, p=0.866]. However, increased locomotor activity, indicating behavioral sensitization, was observed in mice tested with cocaine 20 mg/kg i.p. when they received a 24 hr pretreatment with cocaine 10 mg/kg that was preceded by administration 1 hr pretreatment with either 1 or 3 mg/kg i.p. doses of URB597 (which are known to block the FAAH) [two way ANOVA shows significant main effects of URB pretreatment, F(2,52)=3.326, p<0.05, cocaine treatment, F(1,52)=7.586, p<0.01, and their interaction, F(2,52)=3.163, p<0.05]. Panel B shows dopamine microdialysis data from mice implanted with probes in the NAC core that, in agreement with locomotor activity data, display a larger DAergic response in this brain area, indicating sensitization, obtained in animals pretreated with the FAAH blocker, URB597 (URB) 3 mg/kg i.p. administered 1 hr before a non-sensitizing dose of cocaine, 24 hr before the testing dose of cocaine, 20 mg/kg i.p., compared to animals that received URB vehicle pretretments (pretreatment groups’ number of animals, n, and basal DA values, fmoles/sample ± S.E.M., were: Veh+Sal, ● filled circles, n=4, 63.18±8.6; Veh+cocaine 10 mg/kg, ○ unfilled circles, n=8, 53.2+5.0; URB+Sal, ∎filled squares, n=5, 64.7±3.9; URB+cocaine 10 mg/kg, ◻ unfilled squares, n=6, 52.4±7.9) [two way ANOVA for repeated measures shows significant main effects of URB pretreatment, F(1,27)=4.556, p<0.05, and time, F(3,81)=76.781, p<0.001, and significant URB pretreatment by time interaction, F(3,81)=2.847, p<0.05]. In microdialysis experiments, results are expressed as means with vertical bars representing S.E.M. of the amount of DA in 10 minutes dialysate samples, expressed as percentage of basal values. * = p< 0.05, Tukey’s post-hoc test.