Cue-induced reinstatement of alcohol-seeking behavior is associated with increased ERK1/2 phosphorylation in specific limbic brain regions: blockade by the mGluR5 antagonist MPEP

Abstract

Relapse to alcohol use after periods of abstinence is a hallmark behavioral pathology of alcoholism and a major clinical problem. Emerging evidence indicates that metabotropic glutamate receptor 5 (mGluR5) antagonists attenuate relapse to alcohol-seeking behavior but the molecular mechanisms of this potential therapeutic effect remain unexplored. The extracellular signal-regulated kinase (ERK1/2) pathway is downstream of mGluR5 and has been implicated in addiction. We sought to determine if cue-induced reinstatement of alcohol-seeking behavior, and its reduction by an mGluR5 antagonist, is associated with changes in ERK1/2 activation in reward-related limbic brain regions. Selectively-bred alcohol-preferring (P) rats were trained to lever press on a concurrent schedule of alcohol (15% v/v) vs. water reinforcement. Following 9 days of extinction, rats were given an additional extinction trial or injected with the mGluR5 antagonist MPEP (0, 1, 3, or 10mg/kg) and tested for cue-induced reinstatement. Brains were removed 90-min later from the rats in the extinction and MPEP (0 or 10mg/kg) conditions for analysis of p-ERK1/2, total ERK1/2, and p-ERK5 immunoreactivity (IR). Cue-induced reinstatement of alcohol-seeking behavior was associated with a three to five-fold increase in p-ERK1/2 IR in the basolateral amygdala and nucleus accumbens shell. MPEP administration blocked both the relapse-like behavior and increase in p-ERK1/2 IR. p-ERK1/2 IR in the central amygdala and NAcb core was dissociated with the relapse-like behavior and the pharmacological effect of mGluR5 blockade. No changes in total ERK or p-ERK5 were observed. These results suggest that exposure to cues previously associated with alcohol self-administration is sufficient to produce concomitant increases in relapse-like behavior and ERK1/2 activation in specific limbic brain regions. Pharmacological compounds, such as mGluR5 antagonists, that reduce cue-induced ERK1/2 activation may be useful for treatment of relapse in alcoholics that is triggered by exposure to environmental events.

Figure 1

(A) Extinction of alcohol-reinforced lever pressing. Data are plotted as mean (± SEM) total responses per session on the ethanol and water levers on the last day of baseline (left) and through the 9 days of extinction training (right). Data represent an average of 5 treatment groups (N=31) shown in the lower panel. (B) Effect of the mGluR5 antagonist on cue-induced reinstatement of alcohol-seeking behavior. Data are shown as mean (± SEM) total responses on day 10 of the procedure. Bars show performance of a group that continued on extinction training (Ext) as compared to groups that received MPEP (0 – 10 mg/kg) prior to a cue-induced reinstatement test session. Data represent mean of n=6–7 rats from each drug dose / condition. * - significantly different from water on the same day; † - significantly different from Ext within the same reinforcement condition; ‡ - significantly different from MPEP (0 mg/kg) within the same reinforcement condition, p < 0.05, Tukey test.

Figure 2. p-ERK_1/2 IR in nucleus accumbens shell

(A) Mean (± S.E.M.) p-ERK_1/2 immunoreactivity (pixels/mm²) in the nucleus accumbens shell following extinction (Ext) or cue-induced reinstatement with MPEP (0 or 10 mg/kg) pretreatment. (B) Illustration of brain regions analyzed (adapted from Paxinos and Watson, 1998; nucleus accumbens core – AcbC; nucleus accumbens shell – AcbSh). (C) Representative photomicrographs of the cytological pattern of p-ERK_1/2 IR in the nucleus accumbens shell following Extinction or cue-induced reinstatement with MPEP (0 or 10 mg/kg) pretreatment. Data represent means of multiple brain sections from n=6 rats per condition. * - indicates significantly different from Ext; † - significantly different from MPEP (0 mg/kg), p<0.05, Tukey test.

Figure 3. p-ERK_1/2 IR in basolateral amygdala

(A) Mean (± S.E.M.) p-ERK_1/2 IR (pixels/mm²) in the basolateral amygdala following extinction (Ext) or cue-induced reinstatement with MPEP (0 or 10 mg/kg) pretreatment. (B) Illustration of brain regions analyzed (adapted from Paxinos and Watson, 1998; basolateral amygdala - BLA; central nucleus of the amygdala – CeA. (C) Representative photomicrographs of the cytological pattern of p-ERK_1/2 IR in the basolateral amygdala following Extinction or cue-induced reinstatement with MPEP (0 or 10 mg/kg) pretreatment. Data represent means of multiple brain sections from n=6 rats per condition. * - indicates significantly different from Ext; † - significantly different from MPEP (0 mg/kg), p<0.05, Tukey test.