mTORC1 in the orbitofrontal cortex promotes habitual alcohol seeking
- University of California, San Francisco, United States
Figures
Figure 1 with 4 supplements
Inhibition of mTORC1 in the OFC reduces alcohol seeking.
(a–c) Intra-OFC infusion of rapamycin does not alter self-administration of alcohol. Rats underwent 7 weeks of IA-20%2BC and were then trained to self-administer 20% alcohol using a FR3 schedule. …
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Figure 1—source data 1
Cumulative lever presses at 3 min intervals for vehicle- and rapamycin-treated rats during a 30 min self-administration session (Figure 1a).
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig1-data1-v4.xlsx
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Figure 1—source data 2
Total lever presses and alcohol consumed (g/kg) during a 30 min self-administration session in vehicle- and rapamycin-treated rats (Figure 1b–c).
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig1-data2-v4.xlsx
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Figure 1—source data 3
Cumulative lever presses at 3 min intervals for vehicle- and rapamycin-treated rats during a 30 min extinction session (Figure 1d).
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Figure 1—source data 4
Total lever presses during a 30 min extinction session in vehicle- and rapamycin-treated rats (Figure 1e).
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig1-data4-v4.xlsx
Figure 1—figure supplement 1
Schematic drawing of cannulae placement.
Schematic illustration of coronal sections of the rat brain showing approximate bilateral placements of cannulae in the OFC according to Paxinos and Franklin (2001).
Figure 1—figure supplement 2
Inhibition of mTORC1 in the OFC does not alter alcohol consumption in a 2-bottle choice paradigm.
Rats with a history of IA-20%2BC were infused with vehicle (white) or rapamycin (50 ng/μl, red) into the OFC 3 hr before the beginning of the final 2BC session, and alcohol consumption was recorded …
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Figure 1—figure supplement 2—source data 1
Alcohol consumed (g/kg) by vehicle- and rapamycin-treated rats after 30 min or 24 hr of two-bottle choice.
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig1-figsupp2-data1-v4.xlsx
Figure 1—figure supplement 3
Inhibition of mTORC1 in the OFC does not impact inactive lever presses or the latency to the first and last active lever press.
Rats were trained to self-administer 20% alcohol using an FR3 schedule. Vehicle (white) or rapamycin (50 ng/μl, red) was infused into the OFC 3 hr before an operant self-administration session, and …
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Figure 1—figure supplement 3—source data 1
Total inactive lever presses and the latencies to the first and last active lever presses during a 30 min self-administration session in vehicle- and rapamycin-treated rats.
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig1-figsupp3-data1-v4.xlsx
Figure 1—figure supplement 4
Intra-OFC infusion of rapamycin does not alter locomotion.
Rats were trained to self-administer 20% alcohol using a FR3 schedule. Vehicle (white) or rapamycin (50 ng/μl, red) was infused into the OFC 3 hr before a 30 min extinction session, and …
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Figure 1—figure supplement 4—source data 1
Inter-response intervals during a 30 min self-administration session in vehicle- and rapamycin-treated rats.
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig1-figsupp4-data1-v4.xlsx
Figure 2 with 3 supplements
Inhibition of mTORC1 in the OFC attenuates habitual responding for alcohol.
(a) Timeline of experiment. Rats underwent 7 weeks of IA20%−2BC. Rats were then trained to operant self-administer 20% alcohol during three overnight FR1 sessions and were pseudo-randomly assigned …
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Figure 2—source data 1
Total lever presses during 10 min extinction sessions on non-devalued and devalued days in RR- and RI-trained rats (Figure 2c).
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig2-data1-v4.xlsx
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Figure 2—source data 2
Total lever presses during 10 min extinction sessions on non-devalued and devalued days in vehicle- and rapamycin-treated, RR-trained rats (Figure 2d).
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig2-data2-v4.xlsx
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Figure 2—source data 3
Total lever presses during 10 min extinction sessions on non-devalued and devalued days in vehicle- and rapamycin-treated, RI-trained rats (Figure 2e).
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig2-data3-v4.xlsx
Figure 2—figure supplement 1
Number of lever presses and alcohol consumed in RI and RR-trained animals.
Rats were trained to respond for alcohol in an operant self-administration paradigm under RI (red) or RR (green) schedule of reinforcement. Significance between treatment groups was determined using …
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Figure 2—figure supplement 1—source data 1
Total lever presses during 1 hr RR and RI training sessions (Figure 2—figure supplements 1a).
Alcohol consumed (g/kg) during 1 hr RR and RI training sessions (Figure 2—figure supplements 1b).
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig2-figsupp1-data1-v4.xlsx
Figure 2—figure supplement 2
Schematic drawing of cannulae placement.
Schematic illustration of coronal sections of the rat brain showing approximate bilateral placements of cannulae in the OFC according to Paxinos and Franklin (2001).
Figure 2—figure supplement 3
Inhibition of mTORC1 in the OFC does not alter voluntary alcohol intake prior to devaluation.
Rats were trained to respond for alcohol in an operant self-administration paradigm under RI (red) or RR (green) schedule of reinforcement. Vehicle (white) or rapamycin (50 ng/μl, red) was infused …
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Figure 2—figure supplement 3—source data 1
Alcohol consumed (g/kg) during 30 min home cage alcohol exposure prior to devaluation extinction tests in vehicle- and rapamycin-treated rats.
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig2-figsupp3-data1-v4.xlsx
Figure 3 with 2 supplements
Habitual sucrose responding is not affected by mTORC1 inhibition in the OFC.
(a) Timeline of experiment. Rats underwent 7 weeks of IA20%−2BC and were then trained to operant self-administer 8% sucrose. Sucrose concentration was progressively reduced to 1% during three …
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Figure 3—source data 1
Lever presses during sucrose devaluation extinction test in vehicle- and rapamycin-treated rats (Figure 3c).
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig3-data1-v4.xlsx
Figure 3—figure supplement 1
Schematic drawing of cannulae placement.
Schematic illustration of coronal sections of the rat brain showing approximate bilateral placements of cannulae in the OFC according to Paxinos and Franklin (2001).
Figure 3—figure supplement 2
Inhibition of mTORC1 in the OFC does not alter voluntary sucrose intake prior to devaluation.
Rats were trained to respond for sucrose in an operant self-administration paradigm under RI schedule of reinforcement. Vehicle (white) or rapamycin (50 ng/μl, red) was infused into the OFC 3 hr …
Figure 4 with 2 supplements
GluN2B in the OFC is required for mTORC1 activation by alcohol.
(a–b) Alcohol increases GluN2B phosphorylation in the OFC. Rats underwent 7 weeks of IA-20%2BC resulting in an average of alcohol intake of 5.01 g/kg/24 hr (Table 2). The OFC was dissected at the …
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Figure 4—source data 1
Full, uncropped western blot films from experiments in Figure 4.
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig4-data1-v4.pptx
Figure 4—figure supplement 1
GluN2B-dependent activation of mTORC1 signaling.
Cartoon depicting the signaling cascade detailed in Figure 4. GluN2B activation results in calcium entry and in the activation CaMKII. CaMKII activation is measured by phosphorylation (P). Calcium …
Figure 4—figure supplement 2
Timeline of experiments depicted in Figure 4e–j.
Rats underwent 7 weeks of IA-20%2BC paradigm or drank water only for the same duration of time. Ro-25–6981 (Fischer et al., 1997) (10 μg/µl) or vehicle was bilitteraly infused into the OFC 15 min …
Figure 5 with 3 supplements
GluN2B in the OFC promotes alcohol seeking and habitual alcohol responding.
(a–c) Intra-OFC administration of Ro25-6981 decreases alcohol seeking. Rats underwent 7 weeks of IA-20%2BC, and were then trained to self-administer 20% alcohol. Vehicle (white) or Ro25-6981 (Ro25, …
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Figure 5—source data 1
Cumulative lever presses at 3 min intervals for vehicle- and Ro25-treated rats during a 30 min extinction session (Figure 5a).
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig5-data1-v4.xlsx
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Figure 5—source data 2
Total lever presses and lever pressing rate (presses/min) during a 30 min extinction session in vehicle- and Ro25-treated rats (Figure 5b–c).
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig5-data2-v4.xlsx
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Figure 5—source data 3
Total lever presses during 10 min extinction sessions on non-devalued and devalued days in vehicle- and Ro25-treated, RI-trained rats (Figure 5d).
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig5-data3-v4.xlsx
Figure 5—figure supplement 1
Schematic drawing of cannulae placement.
Schematic illustration of coronal sections of the rat brain showing approximate bilateral placements of cannulae in the OFC according to Paxinos and Franklin (2001).
Figure 5—figure supplement 2
Inhibition of GluN2B in the OFC does not alter locomotion.
Vehicle (white) or Ro25-6981 (10 μg/μl, blue) was infused in the OFC 15 min before a single 30 min extinction session and inter-response intervals (IRIs) were measured. Mean ± SEM of the relative …
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Figure 5—figure supplement 2—source data 1
Inter-response intervals during a 30 min extinction session in vehicle- and Ro25-treated rats.
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig5-figsupp2-data1-v4.xlsx
Figure 5—figure supplement 3
Inhibition of GluN2B in the OFC does not alter voluntary alcohol intake prior to devaluation.
Rats were trained to respond for alcohol in an operant self-administration paradigm under RI schedule of reinforcement. Vehicle (white) or Ro25-6981 (5 μg/μl, blue) was infused into the OFC 15 min …
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Figure 5—figure supplement 3—source data 1
Alcohol consumed (g/kg) during 30 min home cage alcohol exposure prior to devaluation extinction tests in vehicle- and Ro25-treated rats.
- https://cdn.elifesciences.org/articles/51333/elife-51333-fig5-figsupp3-data1-v4.xlsx
Tables
Table 1
Mean consumption of sucrose, alcohol and water during outcome devaluation.
Rats undergoing alcohol seeking probe trials were pre-fed with sucrose (non-devalued test) or alcohol (devalued test), and water 30 min prior to a 10 min extinction session.
| Alcohol Devaluation | |||||||
|---|---|---|---|---|---|---|---|
| Training | RI | RR | RI | ||||
| Group | Vehicle | rapamycin | Vehicle | rapamycin | Vehicle | Ro25 | |
| Non-devalued day | 1% Sucrose (ml/kg) | 1.4 | 2.0 | 1.2 | 1.9 | 2.7 | 5.8 |
| ± | ± | ± | ± | ± | ± | ||
| 0.5 | 0.8 | 0.4 | 0.6 | 1.5 | 1.5 | ||
| Water (ml/kg) | 0.7 | 0.6 | 1.2 | 0.8 | 1.0 | 1.3 | |
| ± | ± | ± | ± | ± | ± | ||
| 0.3 | 0.1 | 0.7 | 0.1 | 0.2 | 0.3 | ||
| Devalued day | 20% Alcohol (g/kg) | 0.7 | 0.7 | 1.0 | 0.7 | 1.0 | 1.3 |
| ± | ± | ± | ± | ± | ± | ||
| 0.1 | 0.1 | 0.1 | 0.1 | 0.1 | 0.1 | ||
| Water (ml/kg) | 0.4 | 0.6 | 0.7 | 0.6 | 1.0 | 0.9 | |
| ± | ± | ± | ± | ± | ± | ||
| 0.1 | 0.1 | 0.1 | 0.1 | 0.4 | 0.2 | ||
| Sucrose Devaluation | |||||||
| Training | RI | ||||||
| Group | Vehicle | rapamycin | |||||
| Non-devalued day | 20% Alcohol (g/kg) | 0.9 | 0.8 | ||||
| ± | ± | ||||||
| 0.1 | 0.1 | ||||||
| Water (ml/kg) | 0.9 | 1.1 | |||||
| ± | ± | ||||||
| 0.1 | 0.1 | ||||||
| Devalued day | 1% Sucrose (ml/kg) | 4.5 | 5.1 | ||||
| ± | ± | ||||||
| 0.7 | 1.4 | ||||||
| Water (ml/kg) | 1.2 | 0.9 | |||||
| ± | ± | ||||||
| 0.2 | 0.1 | ||||||
Table 2
Individual alcohol drinking data from rodents used for biochemical experiments.
Individual alcohol drinking data of the final 4 sessions of IA-2BC20%. Alcohol intake is expressed as mean ± S.E.M.
| Figure 4a, b | Drinking paradigm IA20%−2BC 24h-withdrawal | Rat number | Last four drinking session (g/kg /24 ) |
|---|---|---|---|
| 1 | 5.97 | ||
| Animal numbers | 2 | 6.40 | |
| n = 6 | 3 | 4.00 | |
| 4 | 5.84 | ||
| 5 | 4.23 | ||
| 6 | 3.60 | ||
| Mean ± S.E.M. | 5.01 ± 0.49 | ||
| Figure 4e,f and g | Drinking paradigm IA20%−2BC 24h-withdrawal | Rat number | Last four drinking session (g/kg/24 h) |
| Animal numbers n = 12 | 1 | 5.35 | |
| 2 | 5.52 | ||
| 3 | 5.54 | ||
| 4 | 4.84 | ||
| 5 | 6.41 | ||
| 6 | 3.16 | ||
| 7 | 4.37 | ||
| 8 | 3.45 | ||
| 9 | 3.48 | ||
| 10 | 6.24 | ||
| 11 | 2.71 | ||
| 12 | 5.72 | ||
| Mean ± S.E.M. | 4.73 ± 0.37 |
