FIGURE 1

General design of the study (a) and alcohol-seeking behavior during the final test session (b, c). (a) The experimental procedure consisted of the initiation phase, self-administration and abstinence periods, and ended with the final test session. During the final session, the animals were reexposed to the alcohol self-administration context ('context', the extinction group) or context and discrete cues ('cue') according to the within-session extinction/reinstatement paradigm (the reinstatement group). No alcohol was available during the final session. The animals were anesthetized and perfused 30 (for P-extracellular signal-regulated kinases (ERK), P-c-Jun, and P-JNK immunostaining) or 90 min (for c-Fos immunostaining) from the beginning of the final test session. indicates anesthesia. For other details, see Materials and Methods and Table 1. (b) Mean (SEM) number of non-reinforced responses on the active ('A') and inactive ('IA') lever during the last week of alcohol self-administration and during the final test session (the extinction session) is shown for the alcohol and control rats from the extinction group. The alcohol rats were trained to press the active lever for 8% ethanol and were reexposed to the alcohol self-administration context after 30-day abstinence. During the final session, alcohol was not available. The control animals were always exposed to the same stimuli but were never trained to lever press for any reward and had no access to alcohol. A: active lever (the alcohol rats received portion of alcohol during self-administration phase if pressed that lever). IA: inactive lever (pressing that lever had no programmed consequences). Note that non-reinforced responding on the previously active lever was limited to the alcohol/extinction rats (Table 1). Data were analyzed with a mixed factor ANOVA with repeated measures on time (for the alcohol animals: effect of time: F(2, 36)=54.23, p<0.001; effect of lever: F(1, 18)=38.27, p<0.001); for the control animals: effect of time: F(2, 28)=10.03, p<0.05; effect of lever: F(1, 14)=1.28, p>0.05); ^*** p<0.001 alcohol vs control, Tukey's test. (c) Mean (SEM) number of non-reinforced responses on the active ('A') and inactive ('IA') lever during the last week of alcohol self-administration and during the final test session (the extinction/reinstatement session) is shown for the alcohol and control rats from the reinstatement group. The alcohol rats were trained to press the active lever for 8% ethanol and were reexposed to the alcohol self-administration context and discrete cues after 30-day abstinence. During the final session, alcohol was not available. The control animals were always exposed to the same stimuli but were never trained to lever press for any reward and had no access to alcohol. During the final test session, the animals were exposed for 30 min to the self-administration context. The alcohol-paired discrete cues were repeatedly presented in the last 10 min of the session. Note that non-reinforced responding on the previously active lever was limited to the alcohol/reinstatement rats (Table 1). Data were analyzed with a mixed factor ANOVA with repeated measures on time (for the alcohol animals: effect of time: F(2, 44)=24.57, p<0.001; effect of lever: F(1, 22)=149.99, p<0.001; for the control animals: effect of time: F(2, 44)=1.71, p>0.05; effect of lever: F(1, 22)=0.90, p>0.05). ^*** p<0.001 alcohol vs control, Tukey's test.