High and low doses of cocaine intake are differentially regulated by dopamine D2 receptors in the ventral tegmental area and the nucleus accumbens

Abstract

Dopamine D2 receptors (D2Rs) in the ventral tegmental area (VTA) and the nucleus accumbens (NAc) are associated with vulnerability to addiction; however, whether D2Rs in these two brain regions play differential roles in regulation of drug intake is unknown. Here, we compared the effect of decreased mRNA level of Drd2 in each region on cocaine self-administration in a dose-response function. Drd2 mRNA levels in rat VTA or NAc were knocked down by bilateral microinjection of lentivirus coding shRNAs against rat Drd2 or scrambled shRNA. Drd2 knockdown was persistent and stable between 20 and 90 days after lentiviral infection. Animals were trained to self-administer cocaine 20 days after Drd2 shRNA treatment. Compared to scrambled shRNA treated rats, Drd2 knockdown in the VTA increased cocaine self-administration at all tested doses (0.02-0.56 mg/kg/infusion) producing an upward shift (both the ascending and descending limb) in the dose-response curve of cocaine self-administration. In contrast, intra-NAc knockdown increased cocaine self-administration only on the ascending limb of the dose-response curve (0.02-0.07 mg/kg/infusion). These data suggest that D2Rs in the VTA, not in the NAc, regulate high-dose cocaine intake. The present study not only demonstrates that low levels of D2Rs in either region increase low doses of cocaine intake, but also reveals for the first time their dissociable roles in limiting high doses of cocaine self-administration.

Keywords: Cocaine self-administration; Dopamine D2 autoreceptors; Drd2 knockdown; Nucleus accumbens; Ventral tegmental area.

Fig. 1. Validation of Drd2 knockdown in rat PC12 cells

Rat PC12 cells plated in 24-well plates were infected by lentivirus containing Drd2 shRNA or scrambled shRNA for 3 days and then harvested for qPCR analysis. (a) The scrambled shRNA treatment had no effect on Drd2 mRNA level when compared to the control (no treatment). A one-way ANOVA revealed a significant main effect of treatment (F5,24=41.74, p<0.010). Bonferroni posthoc analysis indicated a significant reduction of Drd2 mRNA level in cells treated with one or the mixture of the four Drd2 shRNAs when compared to scrambled shRNA treatment (^**p<0.01). (b) Drd2 shRNAs did not alter Drd3 mRNA levels (n=5).

Fig. 2. Drd2 mRNA levels in scrambled shRNA and Drd2 shRNA treated animals

Lentivirus containing scrambled shRNA or shRNAs against rat Drd2 was bilaterally injected into the VTA or the NAc. (a) The Drd2 mRNA levels were examined 20- (Drd2KD20) and 90-days (Drd2KD90) post-treatment of Drd2shRNAs in the VTA by quantitative PCR and normalized by β-actin gene. Drd2 shRNAs significantly reduced the mRNA levels 20 days after infection and remained stable even after 90 days. (b) Cocaine self-administration itself reduced Drd2 mRNA levels in the scrambled control (Scramble) and knockdown (Drd2KD) animals in the VTA. (c) Drd2 shRNA in the NAc reduced the mRNA level and cocaine self-administration reduced Drd2 mRNA levels in both scrambled control and knockdown animals in the NAc. Data were presented as the mean Drd2 mRNA levels ± SEM (^*p<0.05, ^**p<0.01 vs control).

Fig. 3. The differential effects of Drd2 knockdown in the VTA and the NAc on cocaine self-administration

(a) The number of cocaine infusions did not differ between scrambled shRNA injection in the VTA and the NAc. Cocaine dose-infusion (b) and dose-intake (c) curves were generated for Drd2 knockdown in the VTA and in the NAc. Data were represented as the mean number of cocaine infusion/hr or the total cocaine intake (mg/kg/hr) ± SEM (^*p<0.05, ^**p<0.01 vs. control). Fig. 3c is in the semi-logarithmic scale.