Dopamine D2 receptors in nucleus accumbens cholinergic interneurons increase impulsive choice

Abstract

Impulsive choice, often characterized by excessive preference for small, short-term rewards over larger, long-term rewards, is a prominent feature of substance use and other neuropsychiatric disorders. The neural mechanisms underlying impulsive choice are not well understood, but growing evidence implicates nucleus accumbens (NAc) dopamine and its actions on dopamine D2 receptors (D2Rs). Because several NAc cell types and afferents express D2Rs, it has been difficult to determine the specific neural mechanisms linking NAc D2Rs to impulsive choice. Of these cell types, cholinergic interneurons (CINs) of the NAc, which express D2Rs, have emerged as key regulators of striatal output and local dopamine release. Despite these relevant functions, whether D2Rs expressed specifically in these neurons contribute to impulsive choice behavior is unknown. Here, we show that D2R upregulation in CINs of the mouse NAc increases impulsive choice as measured in a delay discounting task without affecting reward magnitude sensitivity or interval timing. Conversely, mice lacking D2Rs in CINs showed decreased delay discounting. Furthermore, CIN D2R manipulations did not affect probabilistic discounting, which measures a different form of impulsive choice. Together, these findings suggest that CIN D2Rs regulate impulsive decision-making involving delay costs, providing new insight into the mechanisms by which NAc dopamine influences impulsive behavior.

Fig. 1. D2R upregulation in NAc CINs increases delay discounting.

A Top, low magnification image of AAV-DIO-D2-EGFP expression in CINs of the NAc (shown within dotted lines; scale = 500 µm). Bottom, higher magnification within the NAc core showing typical CIN morphology and co-localization of AAV-DIO-D2-EGFP and ChAT immunolabeling (a.c., anterior commissure; scale = 30 µm). B Schematic illustration of the delay discounting task. On free choice trials, two lever options are presented, each leading to a small or a large reward. The delay to the large reward is progressively increased across sessions (0–10 s), while the small reward is given with no delay. C In the absence of delays to either reward, EGFP and D2R-overexpressing mice similarly increased preference for the large reward option after 14 training sessions (shown here as blocks of 2 sessions). Delay main effect: F _{(6, 84)} = 2.608, p = 0.0229; virus effect: F _{(1, 14)} = 0.07516, p = 0.7880; virus x delay interaction: F _{(6, 84)} = 1.257, p = 0.2861. D As delays to the large reward were progressively increased, a 2-way RM ANOVA found a significant main effect of delay (F _{(5, 70)} = 43.47, p < 0.0001) as well as a significant virus x delay interaction: F _{(5, 70)} = 6.13, p < 0.0001, * denotes significant interaction. n = 8 mice/group, 5 females, 3 males per group. Data is expressed as the average of the last 2 days at each delay. E, F. D2R upregulation did not alter the median press latency during forced trials (virus effect: F _{(1, 14)} = 0.5845, p = 0.4572; virus x delay interaction: F _{(5, 70)} = 1.131, p = 0.3522) or during free choice trials (virus effect: F _{(1, 14)} = 0.4418, p = 0.5171; virus x delay interaction: F _{(5, 70)} = 0.8204, p = 0.5393). Values expressed as mean of median latencies ± S.E.M. G, H. No significant group changes were observed in either distance traveled in 5-min bins over a 90-min period in an open field using a different cohort of mice (virus effect: F _{(1, 18)} = 0.1850, p = 0.6722; virus x delay interaction: F _{(17, 306)} = 1.087, p = 0.3651), or in mean total distance traveled over 90-min (t ₍₁₈₎ = 0.4301, p = 0.6722). EGFP, n = 9 (3 females, 6 males); D2, n = 11 (5 females, 6 males).

Fig. 2. D2R upregulation in NAc CINs does not alter probabilistic discounting.

A Schematic illustration of probabilistic discounting task. On free choice trials, two lever options are presented, each leading to a small or a large reward. The probability of receiving the large reward is progressively decreased across sessions (100–20%), while the small reward is always given. B With increased training, both EGFP and D2R-overexpressing mice increased preference for the large reward when both options were equally probable to yield reward (100%) probability; session block effect: F _{(6, 84)} = 17.85, p < 0.0001; Tukey’s test showed significant differences between session blocks 1 and 7 for both groups (adjusted p values < 0.0001). No significant main effects of virus (F _{(1, 14)} = 0.08267, p = 0.7779) or virus x session block interactions (F _{(6, 84)} = 0.9243, p = 0.4818) were observed. C. A significant main effect of probability was detected (F_{(6, 84)} = 38.39, p < 0.0001), with both groups showing significant simple main effects within-group (100% vs. 33% or 20%, adjusted p values < 0.0001), but this effect was not significantly different following D2R upregulation (virus: F _{(1, 14)} = 0.8632, p = 0.3686; virus x probability: F _{(6, 84)} = 0.7642, p = 0.6001, n = 8 mice/group). EGFP: 3 males, 5 females; D2: 4 males, 4 females.

Fig. 3. CIN D2R upregulation in NAc does not alter timing.

A Schematic representation of the temporal discrimination task. In each session, a single response on one of two lever options is rewarded based on the duration of the sample auditory stimulus. Two tone durations are presented in each session. B Mean proportion of correct responses on the corresponding lever in 2-s tone trials (short, left) or 8-s tone trials (long, right) across blocks of 2 sessions was not altered by D2R upregulation. A 3-way ANOVA showed no virus main effect, F _{(1, 65)} = 0.005903, p = 0.9390, and no significant virus x tone duration x session block (F_{(4, 65)} = 0.9819, p = 0.4237) or virus x session block (F _{(4, 65)} = 0.05092, p = 0.9950) or virus x tone duration (F _{(1, 65)} = 0.2230, p = 0.6383) interactions; n = 8 EGFP (7 females, 1 male), 7 D2 (6 females, 1 male). C The duration of tones was proportionally increased to 6 s (short, left) and 24 s (long, right). While there was a significant main effect of session block on discrimination of tone durations (F _{(7, 104)} = 21.52, p < 0.0001), no significant main effect of virus (F _{(1, 104)} = 1.890, p = 0.1721), or virus x tone duration x session block (F _{(7, 104)} = 0.2323, p = 0.9766), virus x session block (F_{(7, 104)} = 0.6384, p = 0.7232), or virus x tone duration (F _{(1, 104)} = 0.07523, p = 0.7844) interactions were found. D Mean lever press rate during peak trials in the final five sessions of the 24-s peak interval task. E–G Mean best-fitting parameters (derived from fitting to Gaussian function) for peak location (timing accuracy), peak width (timing precision), or peak height (peak response rate). No significant differences were observed in any of these parameters using unpaired t tests.

Fig. 4. Lack of CIN D2Rs decreases delay, but not probabilistic, discounting.

A ChAT-IRES-Cre mice were crossed to Drd2^loxP/loxP mice to obtain mice lacking D2Rs in CINs (CIN-D2KO). CIN-D2KO and control Drd2^loxP/loxP mice increased preference for the large reward with training in the absence of delays to either reward option (session block effect: F _{(6, 78)} = 9.206, p < 0.0001; Tukey’s test showed significant within session effects in both groups between session blocks 1 and 7 (adjusted p values < 0.001), but no effect of genotype: F _{(1, 13)} = 1.651, p = 0.2213, and no genotype x session block interaction: F _{(6, 78)} = 0.607, p = 0.7237). B Compared to Drd2^loxP/loxP mice, CIN-D2KO mice showed greater preference for the large reward at longer delays (decreased delay discounting). Delay effect: F _{(5, 65)} = 72.56, p < 0.0001; genotype x delay: F _{(5, 84)} = 4.756, p < 0.001, * denotes significant interaction. Drd2^loxP/loxP, n = 8 (5 females, 3 males); CIN-D2KO, n = 7 (4 females, 3 males). One Drd2^loxP/loxP did not reach the criterion of ≥50% large choice on Session Block 7 and was excluded from the analysis. C, D Two-way ANOVA showed a significant main effect of delay on median press latency during forced trials (F _{(5, 65)} = 2.628, p = 0.0317), but no genotype effect: F _{(1, 13)} = 0.05419, p = 0.8195; genotype x delay interaction: F _{(5, 65)} = 0.5731, (p = 0.7203) or during free choice trials (genotype effect: F _{(1, 13)} = 0.5917, p = 0.4555; genotype x delay interaction: F _{(5, 65)} = 0.4477, p = 0.8135). No delay effect was seen during free choice trials (F _{(5, 65)} = 1.349, p = 0.2554). Values expressed as mean of median latencies ± S.E.M. E, F No significant group differences were observed in either distance traveled in 5-min bins over a 90-min period in an open field using a different cohort of mice (genotype effect: F (1, 14) = 0.1786, p = 0.6790; virus x delay interaction: F _{(17, 238)} = 0.7946, p = 0.7946), or in mean total distance traveled over 90-min (t ₍₁₄₎ = 0.4226, p = 0.6790). n = 8 mice per group (4 females, 4 males/ group). G Using a third cohort of mice, no significant group differences were found in the preference between reward options during training (when both options were given at 100% probability). Session block effect: F _{(6, 72)} = 0.7816, p = 0.5871; genotype (F _{(1, 12)} = 0.3588, p = 0.5603); genotype x session block interactions (F _{(6, 72)} = 0.9471, p = 0.4672) were observed. H No significant group differences were observed in probabilistic discounting in same cohort as in G (probability effect): F _{(6, 72)} = 14.26, p < 0.0001; genotype x probability: F _{(6, 72)} = 0.9263, p = 0.4814, Drd2^loxP/loxP, n = 8 (4 females, 4 males); CIN-D2KO, n = 6 (3 females, 3 males), 1 CIN-D2KO did not reach the criteria of ≥ 50% large choice on Session Block 7 and was excluded from the analysis.