Dopamine D2 receptors in addiction-like reward dysfunction and compulsive eating in obese rats

Abstract

We found that development of obesity was coupled with emergence of a progressively worsening deficit in neural reward responses. Similar changes in reward homeostasis induced by cocaine or heroin are considered to be crucial in triggering the transition from casual to compulsive drug-taking. Accordingly, we detected compulsive-like feeding behavior in obese but not lean rats, measured as palatable food consumption that was resistant to disruption by an aversive conditioned stimulus. Striatal dopamine D2 receptors (D2Rs) were downregulated in obese rats, as has been reported in humans addicted to drugs. Moreover, lentivirus-mediated knockdown of striatal D2Rs rapidly accelerated the development of addiction-like reward deficits and the onset of compulsive-like food seeking in rats with extended access to palatable high-fat food. These data demonstrate that overconsumption of palatable food triggers addiction-like neuroadaptive responses in brain reward circuits and drives the development of compulsive eating. Common hedonic mechanisms may therefore underlie obesity and drug addiction.

Figure 1. Weight gain and reward dysfunction in rats with extended access to a cafeteria diet

(a) Mean (± SEM) weight gain in chow-only, restricted access and extended access rats (Access × Day interaction: F_39,702 = 7.9, P < 0.0001; *P < 0.05 compared with chow-only group, post-hoc test). (b) Mean (± SEM) percentage change from baseline reward thresholds (Access × Time interaction: F_78,1092 = 1.7, P < 0.0005; *P < 0.05 compared with chow-only group, post-hoc test).

Figure 2. Patterns of consumption in rats with extended access to a cafeteria diet

(a) Mean (± SEM) daily caloric intake in chow-only, restricted access and extended access rats (Access: F_1,324 = 100.6, P < 0.0001; Time: F_18,324 = 7.8, P < 0.0001; Access × Time interaction: F_18,324 = 4.6, P < 0.0001; *P < 0.05 compared with chow-only group, post-hoc test). (b) Mean daily caloric intake (± SEM) from chow (Access: F_2,504 = 349.1, P < 0.0001; Time: F_18,504 = 5.9, P < 0.0001; Access × Time interaction: F_36,504 = 3.52, P < 0.0001; *P < 0.05 compared with chow-only group, post-hoc test). (c) Mean daily caloric intake (± SEM) from fat (Access: F_2,486 = 118.7, P < 0.0001; Time: F_18,486 = 8.8, P < 0.0001; Access × Time interaction: F_36,486 = 6.2, P < 0.0001; *P < 0.05 compared with chow-only group, post-hoc test). (d) Comparison of mean (± SEM) total caloric intake, and calories consumed exclusively from chow, during the entire 40-day period of access (Access: F_2,54 = 25.0, P < 0.0001; Calorie source: F_2,54 = 1235.2, P < 0.0001; Access × Calorie source interaction: F_2,54 = 485.7, P < 0.0001; ***P < 0.001 compared with total calories in chow-only group, ###P < 0.001 compared with total calories in the same group of rats, post-hoc test).

Figure 3. Persistent reward dysfunction and hypophagia during abstinence in rats with extended access to a cafeteria diet

(a) Mean percentage change from baseline reward thresholds (± SEM) during abstinence from a palatable high-fat diet (Access: F_2,112 = 3.7, P < 0.05; Time: F_4,112 = 2.3, P > 0.05; *P < 0.05 compared with chow-only group, post-hoc test). (b) Mean caloric intake (± SEM) on the last day of access to the high-fat diet (Baseline) and during the 14 days of abstinence when only standard chow was available (Access: F_2,168 = 41.7, P < 0.0001; Time: F_6,168 = 65.6, P < 0.0001; Access × Time interaction: F_12,168 = 38.3, P < 0.0001; *P < 0.05 compared with chow-only group, post-hoc test). (c) Change in mean body weight (± SEM) compared with body weight on the last day of access to the high-fat diet (Baseline) and during the 14 days of abstinence when only standard chow was available (Access: F_1,126 = 37.2, P < 0.0001; Time: F_7,126 = 3.1, P < 0.01; Access × Time interaction: F_7,126 = 40.9, P < 0.0001; *P < 0.05 compared with chow-only group, post-hoc test). (d) Histological reconstruction of the location of BSR stimulating electrodes in the lateral hypothalamus of chow-only (△), restricted access () and extended access (●) rats.

Figure 4. Weight gain is inversely related to striatal D2R levels

(a) Chow-only, restricted access and extended access rats were sub-divided into two groups per access condition based on a median split of body weights; light (L) or heavy (H). (b) The entire striatal complex was collected from all rats and D2R levels in each group measured by Western blotting. The membrane-associated post-synaptic D2R band was resolved at 70 kDa, and the protein-loading control is displayed below (β-actin, 43 kDa). Full-length immunoblots are displayed in Supplementary Figure 12. (c) Relative amounts of D2R in the striatum of chow-only, restricted and extended access rats were quantified by densitometry (F_2,6=5.2, P < 0.05, main effect of Access; *P < 0.05 compared with Chow-only-L group).

Figure 5. Lentivirus-mediated knockdown of striatal D2R expression

(a) Graphical representation of the striatal areas in which Lenti-D2Rsh was overexpressed. Green circles in the left striatal hemisphere represent the locations at which viral infusions were targeted. Green staining in the right striatal hemisphere is a representative immunochemistry staining for GFP from the brain of a Lenti-D2Rsh rat. (b) Representative immunoblot of the decreased D2R expression in the striatum of Lenti-D2Rsh rats. Full-length immunoblots are displayed in Supplementary Figure 13. (c) Relative amounts of D2R in the striatum of Lenti-Control and Lenti-D2Rsh rats were quantified by densitometry (*P < 0.05 compared with the Lenti-Control group, post-hoc test). (d) Infection of glial cells in the striatum by the Lenti-D2Rsh vector was not detected. Green staining is GFP from virus; red is the astrocyte marker glial fibrillary acidic protein [GFAP]; cell nuclei are highlighted by DAPI staining in blue. The white arrows indicate a highly localized area of gliosis found only at the site of virus injection in the striatum and not in the surrounding tissues into which the virus has diffused. Even in this area none of the astrocytes were GFP-positive. The yellow arrows in the magnified image highlight the typical GFP-negative astrocytes that were detected. (e) High levels of neuronal infection in the striatum by the Lenti-D2Rsh vector were detected. Green staining is GFP from virus; red is the neuronal nuclear marker NeuN; cell nuclei are highlighted by DAPI staining in blue. The yellow arrows in the magnified image highlight GFP-positive and NeuN-positive neurons in the striatum. (f) A higher magnification image of a virally infected (GFP-positive) neuron in the striatum of Lenti-D2Rsh rats that displays the typical morphological features of medium spiny neurons.

Figure 6. Knockdown of striatal D2R increases vulnerability to reward dysfunction in rats with extended access to a cafeteria diet

(a) Mean (± SEM) percentage change from baseline reward thresholds in Lenti-Control and Lenti-D2Rsh rats that had extended access to the cafeteria diet for 14 consecutive days (Virus: F_1,156 = 5.9, P < 0.05; Time: F_13,156 = 2.2, P < 0.05; Virus × Time interaction: F_13,156 = 2.2, P < 0.05; #P < 0.05, interaction effect). (b) Mean (± SEM) percentage change from baseline reward thresholds in Lenti-Control and Lenti-D2Rsh rats that had chow-only access (c) Mean (± SEM) caloric intake during 14 days of chow-only or extended access animals (Access: F_2,28 = 135.6, ***P < 0.0001). (d) Mean (± SEM) weight gain during 14 days of chow-only or extended access animals (Access: F_2,28 = 96.4, P < 0.0001; ***P < 0.001, main effect of access).

Figure 7. Compulsive-like responding for palatable food

(a) Mean (± SEM) palatable diet consumption (kCal; ± SEM) in unpunished rats during the 30 min baseline sessions and on the test day when rats were exposed to a neutral conditioned stimulus that was not previously paired with noxious foot shock (Access: F_2,20 = 5.2, P < 0.05; #P < 0.05 compared with chow-only rats). (b) Mean (± SEM) palatable diet consumption (kCal; ± SEM) in punished rats during the 30 min baseline sessions and on the test day when rats were exposed to a conditioned stimulus that was previously paired with noxious foot shock (Access: F_2,21 = 3.9, P < 0.05; Cue: F_1,21 = 8.6, P < 0.01; Access × Cue interaction: F_2,21 = 4.7, P < 0.05; *P < 0.05 compared with intake during the baseline session, #P < 0.05 compared with chow-only rats). (c) Mean (± SEM) palatable diet consumption during the 30 min baseline sessions and on the test day in Lenti-Control and Lenti-D2Rsh rats that previously had chow-only or extended access to a cafeteria diet (Cue: F_1,26 = 29.7, P < 0.0001; *P < 0.05, **P < 0.01 compared with intake during the baseline sessions, post-hoc test). (d) Mean (± SEM) chow consumption during the 30 min baseline sessions and on the test day in Lenti-Control and Lenti-D2Rsh rats that previously had chow-only or extended access to a cafeteria diet (Cue: F_1,26 = 44.9, P < 0.0001; *P < 0.05, **P < 0.01 compared with intake during the baseline sessions, post-hoc test).