Daily N-acetylcysteine pretreatment blunts cocaine-induced behavioral plasticity
In the present studies, we tested the hypothesis that N-acetylcysteine administered prior to each daily cocaine session would prevent the development of compulsive drug seeking. illustrates the impact of N-acetylcysteine pretreatment on daily cocaine self-administration under extended access conditions (6 hr/day). A comparison of daily infusions during each self-administration session produced a three-way interaction between cocaine treatment, N-acetylcysteine treatment, and self-administration session (ANOVA; F10,550=3.05, p=.001). Post hoc analyses indicated that cocaine self-administering rats pretreated with saline exhibited escalation, which was evident as an increase from day 1 in cocaine intake that was significant by the fifth session, and that this effect was prevented in cocaine self-administering rats pretreated with N-acetylcysteine (; Dunnetts T, p<.05). Further, N-acetylcysteine did not significantly alter the number of cocaine infusions obtained until escalation of intake was evident in cocaine rats pretreated with saline (; Dunnetts T, p<.05). The magnitude of escalation is evident when directly comparing the difference in intake on the last and first sessions (). Analysis of these data produced a significant interaction between N-acetylcysteine and cocaine treatments (ANOVA; F10,550=3.05, p=.001); post hoc tests confirmed that only cocaine rats pretreated with saline exhibited escalation (Dunnetts T, p<.05).
Figure 1 N-acetylcysteine pretreatment prevents escalation of drug intake during daily extended-access cocaine self-administration. Data depict (a) mean ± SEM daily cocaine intake (mg/kg, IV) across 11 maintenance self-administration sessions and (b) difference (more ...)
illustrates the impact of N-acetylcysteine administered prior to seven daily injections of cocaine on the establishment of behavioral sensitization. A comparison of total distance traveled on day 1 of the experiment resulted in a significant two-way interaction between time (18 ten-min intervals) and cocaine treatment (ANOVA: F17,782=14,03, p<.001). Post hoc analyses revealed that all rats treated with cocaine, regardless of N-acetylcysteine pretreatment, exhibited significantly higher levels of locomotor activity when compared to rats treated with saline (Dunnetts T, p<.05) indicating that N-acetylcysteine did not alter acute cocaine-induced locomotor activity (). Separate ANOVAs comparing total distance traveled on days 7 and 28 of the experiment each produced a significant three-way interaction between time (18 ten-min intervals), N-acetylcysteine treatment, and cocaine treatment (day 7: F17,782=1.92, p=.01; day 28: F17,782=2.69, p<.001). Post hoc analyses indicated that both cocaine groups exhibited higher levels of locomotor activity relative to saline controls on days 7 and 28 of the experiment (Dunnetts T, p<.05). Further, cocaine rats pretreated with saline exhibited higher levels of activity relative to those pretreated with N-acetylcysteine following the last of seven daily cocaine injections (day 7) and after a cocaine challenge given 21 days after the last N-acetylcysteine pretreatment (day 28; ; Dunnetts T, p<.05). To test for behavioral sensitization, we compared total distance across all three test days. This ANOVA produced an interaction between test day, N-acetylcysteine, and cocaine (F2,92=3.15, p=.05). Post hoc analyses indicated that rats pretreated with saline, but not N-acetylcysteine, exhibited cocaine-induced behavioral sensitization, evident as a significant increase in cocaine-induced activity on days 7 and 28 relative to day 1 (; Dunnets T, p<.05).
Figure 2 Daily N-acetylcysteine prevents cocaine-induced behavioral sensitization without altering acute locomotor activity. Total distance traveled (mean cm ± SEM) is depicted in 10-min intervals (a,b) or summed across the 2 hr session (c). Group designations (more ...)
illustrates the impact of repeated N-acetylcysteine pretreatment on self-administration under short-access conditions (2 hr/day) and cocaine-induced reinstatement of extinguished lever pressing. A comparison of infusions obtained during daily self-administration sessions with N-acetylcysteine and cocaine treatments as between subjects variables and self-administration session as a within subjects variable yielded only a main effect of cocaine (ANOVA: F1,37=115, p<.001), indicating that N-acetylcysteine pretreatment did not significantly alter self-administration under short-access conditions (). N-acetylcysteine administered during self-administration training also failed to alter the mean ± SEM number of extinction sessions needed to reach criteria relative to cocaine rats pretreated with saline (SEM: 11 ± 2 & 13.6 ± 1.7, respectively; p>.05). In contrast, a comparison of lever pressing during the reinstatement test day and the last three extinction sessions produced a three-way interaction between test day, N-acetylcysteine treatment, and cocaine treatment (ANOVA: F3,111=3.06, p=.03). Post hoc analyses revealed that a cocaine prime reinstated extinguished drug seeking, evident as a significant increase in lever pressing on the reinstatement test relative to the last extinction session, but only in rats that had been treated with saline during cocaine self-administration (Dunnetts T, p<.05); indeed, this group of rats exhibited significantly higher levels of responding on the reinstatement test day than all other groups (Dunnetts T, p<.05). In contrast, cocaine rats pretreated with N-acetylcysteine failed to reinstate cocaine-seeking (Dunnetts T, p<.05) despite testing 2–3 weeks after the last N-acetylcysteine treatment ().
Figure 3 Daily N-acetylcysteine prevents cocaine-induced reinstatement without altering cocaine self-administration. Data are depicted as mean ± SEM number of (a) infusions across daily self-administration sessions (2 hr/day) or (b) responses on the active (more ...)
N-acetylcysteine Pretreatment Prevents Cocaine-Induced Changes in Extracellular Glutamate Levels in the Nucleus Accumbens
illustrates extracellular basal glutamate levels sampled from the nucleus accumbens using in vivo microdialysis. An ANOVA used to compare glutamate levels (pmol/μl; ) with N-acetylcysteine and cocaine treatment as between subject measures and time (nine 20-min samples) as a repeated measure did not yield a significant interaction between the variables (F8,296=1.28, p=.25), nor a main effect of cocaine (F1,37=0.54, p=.47), N-acetylcysteine (F1,37=0.94, p=.34) or time (F8,296=1.73, p=.09). A comparison of basal glutamate (e.g., prior to cocaine injection; ) calculated as area under the curve yielded a significant interaction between N-acetylcysteine and cocaine treatments (ANOVA: F1,37=4.05, p=.05). Subsequent post hoc analyses revealed that a reduction in basal glutamate levels relative to saline controls was evident in rats pretreated with saline prior to daily cocaine self-administration sessions; this change in extracellular glutamate was not evident in cocaine rats that had been treated with N-acetylcysteine during self-administration training even when sampled 2–3 weeks after the last N-acetylcysteine treatment (Dunnets T; p<.05).
Figure 4 Daily N-acetylcysteine prevents long-term changes in extracellular glutamate levels in the nucleus accumbens produced by repeated cocaine. Basal glutamate levels (mean ± SEM) are depicted as pmol/μl across 20 min samples (a) or area under (more ...)
An ANOVA comparing cocaine-evoked glutamate (percent baseline; ) with N-acetylcysteine and cocaine treatment as between subject measures and time (nine 20-min samples) as a repeated measure yielded a significant three way interaction (F8,296=2.05, p=.04). Post hoc analyses indicated that cocaine increased extracellular glutamate relative to the last baseline sample in rats that had been pretreated with saline prior to cocaine self-administration (Dunnets T, p<.05; ); this effect was not obtained in rats pretreated with N-acetylcysteine during self-administration training (Dunnetts T, p<.05). Analyzing glutamate as area under the curve generated from samples collected after the cocaine injection also indicated that only cocaine rats pretreated with saline exhibited a significant increase in extracellular glutamate. An interaction between N-acetylcysteine and cocaine treatments group was obtained (F1,37=4.05, p=.05), with post hoc analyses indicating a significant increase in glutamate in rats pretreated with saline prior to cocaine self-administration relative to saline controls or cocaine rats pretreated with N-acetylcysteine (Dunnetts T, p<.05). While the area primarily sampled in this experiment is the nucleus accumbens, the microdialysis probe for some subjects also extended into the striatum dorsal to the nucleus accumbens or the olfactory tubercles ().
Figure 5 A schematic illustrating the placement of the 2 mm active membrane portion of the microdialysis probe for the rats included in the microdialysis study. The active regions of the microdialysis probes were primarily located in the nucleus accumbens, although (more ...)
N-acetylcysteine Pretreatment Prevents Cocaine-induced Plasticity involving System xc
illustrates the impact of daily N-acetylcysteine pretreatment on cocaine-induced plasticity involving system xc-. A comparison of 35S cystine transport produced an interaction between N-acetylcysteine and cocaine treatment conditions (F1,31=7.53, p=.01). Post hoc analyses revealed that rats pretreated with saline prior to cocaine self-administration exhibited reduced 35S cystine transport relative to saline controls (Dunnetts T, p<.05). Further, rats pretreated with N-acetylcysteine prior to cocaine self-administration failed to exhibit diminished cystine-glutamate exchange even though cystine uptake was measured 23–34 days after the last treatment of either cocaine or N-acetylcysteine (Dunnetts T, p<.05).
Figure 6 Daily N-acetylcysteine prevents cocaine-induced blunting of system xc- activity in the nucleus accumbens. Data depict 35S cystine transport restricted to system xc- activity in nucleus accumbens tissue punches. Group designations refer only to treatments (more ...)