Cue-induced reinstatement of seeking behavior in male rats is independent from the rewarding value of the primary reinforcer: Effect of mGluR5 blockade

Abstract

Environmental conditioning factors have a profound impact on alcohol-seeking behavior and the maintenance of alcohol use in individuals with alcohol dependence. Cues associated with alcohol, depending on the perceived value of the primary reinforcer, gain salience and can trigger relapse. This study investigates the correlation between the reward magnitude of the primary reinforcer and the reinstatement evoked by cues predictive of their availability in male rats. Rat self-administration procedures were used to test reinstatement, with reinforcers consisting of 10% alcohol, 10% sucrose, or 2% sodium chloride (NaCl) experienced under need-state conditions. The effect of MTEP ([(2-methyl-1,3-thiazol-4-yl) ethynyl] pyridine), a selective metabotropic glutamate receptor 5 (mGluR5) antagonist, on motivation and reinstatement behaviors was also evaluated. RESULTS: demonstrate that under Fixed Ratio 1 (FR1) schedule, the three reinforcers maintain operant responding with the following order of magnitude 10% sucrose >2% NaCl >10% alcohol > water. Under a progressive ratio (PR) schedule of reinforcement, rats exhibit a significantly higher breakpoint for 2% NaCl (under Na-depletion), followed by 10% sucrose and 10% alcohol. After extinction, a significant reinstatement is observed with the magnitude order of 10% sucrose >10% alcohol >2% NaCl. However, only re-exposure to alcohol-paired cues induced significant reinstatement of alcohol-seeking after 4 and 8 months. Treatment with MTEP significantly reduces reinstatement of responding across all reinforcers, with the strongest effect observed on alcohol-seeking. These findings suggest that mGluR5 plays a general role in controlling cue-reactivity, but the effect is prominent in the case of alcohol compared to natural rewards. In conclusion, the results demonstrate a remarkable dissociation between the rewarding magnitude of the primary reinforcer and its ability to trigger relapse upon presentation of a cue previously associated with it. Importantly, alcohol, despite having lower intrinsic motivational value compared to a natural reward (sucrose) or a consummatory stimulus experienced under need state conditions (NaCl), can elicit more robust and longer-term reinstatement of seeking responses. Finally, our data demonstrate a significant involvement of the mGluR5 system in the regulation of seeking behavior.

Keywords: Progressive ratio; Rats; Reinstatement; Rewards.

Figure 1:. Alcohol, Sucrose, NaCl, and Water Self-Administration under Progressive Ratio

The upper panel depicts the timeline of the experimental design, highlighting the different stages of the experiment with corresponding colors. (A) The self-administration phase utilized a fixed ratio 1 (FR1) schedule of reinforcement. The number of rewards obtained (+/−SEM) is shown for 10% alcohol (red circles), 10% sucrose (green circles), 2% NaCl (blue circles), and water (grey circles). (B) Represents the mean ± SEM of the three breakpoints (BP) achieved for each substance. The breakpoints are displayed for alcohol in the red column, sucrose in the green column, NaCl in the blue column, and water in the grey column. Breakpoints were measured during self-administration under a progressive ratio (PR) schedule of reinforcement. *p < 0.05, **p < 0.01 denote statistical significance compared to the water group. #p < 0.05, ##p < 0.01 denote statistical significance compared to the alcohol group.

Figure 2:. Assessment of Cue-Induced Alcohol-, Sucrose-, and NaCl-Seeking

The upper panel depicts the timeline of the experimental design, highlighting the different stages of the experiment with corresponding colors. (A) Cue-Induced Alcohol Seeking. (B) Cue-Induced Sucrose Seeking. (C) Cue-Induced NaCl seeking. Each figure represents the mean ± SEM of the active lever responses during immediate cue-induced reinstatement, cue-induced reinstatement 4 months later, and cue-induced reinstatement 8 months later. Each reinstatement cycle is preceded by an extinction phase. The discrimination phase shows that during the FR1 training phase, rats successfully learned to discriminate between the reward (sucrose, NaCl, or alcohol) in the S⁺/CS⁺ paired condition and the water in the S⁻/CS⁻ paired condition. White bars indicate extinction. Gray bars represent the S⁻/CS⁻ data.*p < 0.05, **p < 0.01 denote statistical significance compared to extinction.

Figure 3:. Correlation analysis for Alcohol, Sucrose, and NaCl.

This figure shows the correlation between the breakpoint (BP) achieved in the progressive ratio (PR) test (Experiment 1) and cue-induced reinstatement (Experiment 2) for: (A) alcohol, (B) sucrose, and (C) NaCl. No correlation was detected for the three reinstatements in all the substances tested. (D) Normalized data as the number of lever presses during reinstatement divided by (BP). Post-hoc tests indicated a significant difference between alcohol and sucrose (*p < 0.05) or NaCl (**p < 0.001), confirming that alcohol produces significantly stronger cue-induced relapse. (E) The Kaplan-Meier analysis showed that the alcohol group had a higher probability of eliciting long-term reinstatement compared to the other groups after the Mantel-Cox Test.

Figure 4:. Effect of MTEP on Cue-Induced Reinstatement

The upper panel depicts the timeline of the experimental design, highlighting the different stages of the experiment with corresponding colors. (A) MTEP on alcohol seeking. (B) MTEP on sucrose seeking. (C) MTEP on NaCl seeking. Reinstatement of responding was significant for alcohol, sucrose, and NaCl under the S⁺/CS⁺ but not under the S⁻/CS⁻ stimuli. Pretreatment with MTEP significantly reduced S⁺/CS⁺-induced reinstatement for all conditions. The effect of MTEP administration at 1.0 mg/kg, but not 0.3 mg/kg, significantly reduced cue-induced reinstatement for all drugs (p < 0.01) for alcohol and (p < 0.05) for sucrose and NaCl. *p < 0.05, **p < 0.01 denote statistical significance compared to the saline treatment.

Figure 5:. Effect of MTEP treatment on Alcohol, Sucrose and NaCl self-administration under PR

The upper panel depicts the timeline of the experimental design, highlighting the different stages of the experiment with corresponding colors. The lower panel indicates the effect of MTEP 1mg/kg on alcohol (red), sucrose (green) and NaCl (blue) self-administration under PR. No statistical significance was observed in all the three groups.