Molecular, cellular, and structural mechanisms of cocaine addiction: a key role for microRNAs

Abstract

The rewarding properties of cocaine play a key role in establishing and maintaining the drug-taking habit. However, as exposure to cocaine increases, drug use can transition from controlled to compulsive. Importantly, very little is known about the neurobiological mechanisms that control this switch in drug use that defines addiction. MicroRNAs (miRNAs) are small non-protein coding RNA transcripts that can regulate the expression of messenger RNAs that code for proteins. Because of their highly pleiotropic nature, each miRNA has the potential to regulate hundreds or even thousands of protein-coding RNA transcripts. This property of miRNAs has generated considerable interest in their potential involvement in complex psychiatric disorders such as addiction, as each miRNA could potentially influence the many different molecular and cellular adaptations that arise in response to drug use that are hypothesized to drive the emergence of addiction. Here, we review recent evidence supporting a key role for miRNAs in the ventral striatum in regulating the rewarding and reinforcing properties of cocaine in animals with limited exposure to the drug. Moreover, we discuss evidence suggesting that miRNAs in the dorsal striatum control the escalation of drug intake in rats with extended cocaine access. These findings highlight the central role for miRNAs in drug-induced neuroplasticity in brain reward systems that drive the emergence of compulsive-like drug use in animals, and suggest that a better understanding of how miRNAs control drug intake will provide new insights into the neurobiology of drug addiction.

Figure 1

Subregions of the rodent anterior striatum involved in cocaine self-administration. After limited cocaine self-administration, the shell subregion of the nucleus accumbens (NAc) mediates the rewarding effects of cocaine, whereas the core of the NAc mediates cocaine's satiating effects and the conditioned reinforcing effects of drug-paired pavlovian stimuli. Extended cocaine self-administration recruits the dorsal striatum, whereas the NAc continues to play an important role in self-administration. The dorsal striatum comes to control escalation of cocaine intake, habitual cocaine seeking, and eventually continued cocaine seeking in the face of contingent foot-shock punishment. Neuroanatomical representation of the rat striatum is 1.2 mm anterior to Bregma.

Figure 2

(a) Expression profiling of microRNAs (miRNAs) in the dorsal striatum of control, restricted, or extended access rats. Each column represents data from a cohort of two rats, with a total of six rats per group. The bracket identifies miR-212 and miR-132, the expression of which was increased only in extended access rats. (b) Taqman assay verified that striatal miR-212 levels were increased in extended access rats. (c) Taqman assay verified that striatal miR-132 levels were increased in extended access rats. ^*P<0.05, ^**P<0.01, statistically significant compared with the extended access group. This figure is reproduced with permission from Hollander et al (2010).

Figure 3

(a) The right panel is representative immunochemistry staining ( × 10 magnification) from rats treated with a lentivirus vector to overexpress miR-212 (Lenti-miR-212) or a control lentivirus (Lenti-control). Red circles are locations at which viral infusions were targeted in the dorsal striatum. Green is green fluorescent protein (GFP) from virus; red is the astrocyte marker glial fibrillary acidic protein (GFAP). CC, corpus callosum; Ctx, cortex; DSt, dorsal striatum; LV, lateral ventricle. Yellow arrows highlight the injector track used to deliver virus. Inset is an × 80 confocal image of a virus-infected neuron. (b) Fluorescent in situ hybridization was used to visualize striatal miR-212 expression (shown in red) in Lenti-control and Lenti-miR-212 rats, verifying that miR-212 is constitutively expressed in the dorsal striatum and that the Lenti-miR-212 vector upregulates miR-212 expression. (c) Striatal miR-212 overexpression reverses the long-term trajectory of cocaine intake in rats with extended access. (d) A locked nucleic acid (LNA)-modified antisense oligonucleotide against miR-212 (LNA-anti-miR-212) delivered into the dorsal striatum increases cocaine intake in extended access rats. ^*P<0.05, ^**P<0.01. Data are presented as mean±SEM. This figure is adapted with permission from Hollander et al (2010).

Figure 4

(a) Immunochemical detection of methyl CpG-binding protein 2 (MeCP2) in the dorsal striatum of drug-naive rats. Top left, MeCP2 (red) rarely colocalized with glial fibrillary acidic protein (GFAP; green), a marker for astrocytes. Top right, MeCP2 (red) almost exclusively colocalized with the neuronal nuclear marker NeuN (green). There was an increase in the number of MeCP2-positive cells in the dorsal striatum of rats with extended cocaine access (bottom right) compared to that in rats with restricted access (bottom left). (b) The stimulatory effects of cocaine on miR-212 expression in the dorsal striatum were dramatically increased in extended access rats treated with a lentivirus vector delivering a short hairpin interfering RNA to knockdown MeCP2 (Lenti-sh-MeCP2 rats). (c) Disruption of striatal miR-212 signaling using an antisense oligonucleotide (LNA-anti-miR-212) ‘rescues' low levels of cocaine intake in Lenti-sh-MeCP2 rats with extended daily access to cocaine. (d) MeCP2 likely inhibits miR-212 (and miR-132) expression by binding to methylated CpG dinucleotides in the promoter region of the miR-212/132 gene cluster, resulting in the recruitment of histone deacetylases (HDACs) and other co-repressor proteins. ^*P<0.05, ^***P<0.001. This figure is adapted with permission from Im et al (2010).

Figure 5

The complex interactions between miR-212, cAMP response element-binding protein (CREB), methyl CpG-binding protein 2 (MeCP2), and brain-derived neurotrophic factor (BDNF). The red circle implies that CREB signaling protects against the development of escalating cocaine intake, whereas the green circle implies that MeCP2-BDNF signaling promotes escalation of intake. Green arrows indicate a stimulatory relationship, whereas red lines indicate an inhibitory relationship. Cocaine is shown to activate both CREB and MeCP2-BDNF signaling, and the balance between these two pathways, coordinated by miR-212, likely regulates escalation of cocaine intake in extended access rats, and perhaps vulnerability to cocaine addiction.

Figure 6

Detailed overview of the mechanisms by which miR-212 in the dorsal striatum controls cocaine intake in rats with extended access to the drug. Prolonged access to cocaine in rats with extended access increases levels of phosphorylated cAMP response element-binding protein (p-CREB) in the dorsal striatum, likely through activation of dopamine D1 receptors, which increases miR-212 expression (green arrow). Upregulated miR-212 can subsequently influence cocaine intake through at least two mechanisms. First, mR-212 represses expression of SPRED1, a repressor of Raf1 kinase. As a result, Raf1 can phosphorylate and sensitize the activity of membrane-bound adenylate cyclases, thereby increasing intracellular levels of the second messenger cAMP. This results in the activation of protein kinase A (PKA) and other downstream kinases, increasing the phosphorylation of CREB and its association with coactivators such as CREB-regulated transcription coactivator (CRTC). In the activated state, CRTC is acetylated, which stabilizes its association with pCREB and drives the expression of genes with a cAMP response element in their promoter, one of which is miR-212. As CREB-responsive genes are protective against the development of escalating cocaine intake, this action reduces the motivation to further consume cocaine. A second mechanism by which miR-212 can influence cocaine intake is by repressing methyl CpG-binding protein 2 (MeCP2), whose expression levels are known to correlate closely with those of brain-derived neurotrophic factor (BDNF) in the striatum. As BDNF drives cocaine intake, miR-212 knockdown of MeCP2 reduces the motivation to consume cocaine. Interestingly, MeCP2 itself can repress miR-212 expression. Hence, miR-212 can boost striatal CREB and reduce striatal MeCP2 signaling, with these two transcription factors in turn feeding back to control miR-212 expression levels. Hence, homeostatic interactions between miR-212, CREB, and MeCP2 likely control vulnerability to develop compulsive-like cocaine intake in rats with extended access to the drug.